2024-03-29T00:44:00Z
http://ir.lib.uwo.ca/do/oai/
oai:ir.lib.uwo.ca:physpharmpub-1000
2009-12-12T02:27:43Z
publication:physpharm
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:paed
publication:pmid
publication:faculties
publication:paedpub
18505471
Connexin Expression and Gap Junctional Coupling in Human Cumulus Cells: Contribution to Embryo Quality
Wang, Hong-Xing
Tong, Dan
El-Gehani, Faraj
Tekpetey, Francis R.
Kidder, Gerald M.
Article
2008-05-24T07:00:00Z
Gap junction
Conductance
Connexin43
Pregnancy
Journal of Cellular and Molecular Medicine
Journal of Cellular and Molecular Medicine
13
5
972
984
10.1111/j.1582-4934.2008.00373.x
Medical Physiology
Obstetrics and Gynecology
Gap junctional coupling among cumulus cells is important for oogenesis since its deficiency in mice leads to impaired folliculogenesis. Multiple connexins (Cx), the subunits of gap junction channels, have been found within ovarian follicles in several species but little is known about the connexins in human follicles. The aim of this study was to determine which connexins contribute to gap junctions in human cumulus cells and to explore the possible relationship between connexin expression and pregnancy outcome from in vitro fertilization (IVF). Cumulus cells were obtained from IVF patients undergoing intracytoplasmic sperm injection (ICSI). Connexin expression was examined by RT-PCR and confocal microscopy. Cx43 was quantified by immunoblotting and gap junctional coupling was measured by patch-clamp electrophysiology. All but five of 20 connexin mRNAs were detected. Of the connexin proteins detected, Cx43 forms numerous gap junction-like plaques but Cx26, Cx30, Cx30.3, Cx32, and Cx40 appeared to be
restricted to the cytoplasm. The strength of gap junctional conductance varied between
patients and was significantly and positively correlated with Cx43 level, but neither was correlated with patient age. Interestingly, Cx43 level and intercellular conductance were positively correlated with embryo quality as judged by cleavage rate and morphology, and were significantly higher in patients who became pregnant than in those who did not. Thus, despite the presence of multiple connexins, Cx43 is a major contributor to gap junctions in human cumulus cells and its expression level may influence pregnancy outcome after ICSI.
https://ir.lib.uwo.ca/physpharmpub/4
oai:ir.lib.uwo.ca:physpharmpub-1003
2009-05-23T02:13:47Z
publication:physpharm
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:paed
publication:pmid
publication:faculties
publication:paedpub
19038973
Identification of WNT/β-CATENIN Signaling Pathway Components in Human Cumulus Cells
Wang, Hong-Xing
Tekpetey, Francis R.
Kidder, Gerald M.
Article
2009-01-01T08:00:00Z
Ovarian follicle
Paracrine signaling
Folliculogenesis
Gene expression
Signal transduction
Assisted reproduction
Molecular Human Reproduction
Molecular Human Reproduction
15
1
11
17
Medical Physiology
Obstetrics and Gynecology
Signaling via the conserved WNT/β-CATENIN pathway controls diverse developmental processes. To explore its potential role in the ovary, we investigated the expression of WNTs, frizzled (FZD) receptors and other pathway components in human cumulus cells obtained from oocytes collected for in vitro fertilization. Proteins were detected in cultured cells using immunofluorescence microscopy. Protein–protein interactions were analyzed by means of immunoprecipitation. WNT2, FZD2, FZD3 and FZD9 were identified but WNT1, WNT4 and FZD4 were not detected. WNT2 is co-expressed with FZD2, FZD3 and FZD9. Co-immunoprecipitation using WNT2 antibody demonstrated that WNT2 interacts with both FZD3 and FZD9, but only FZD9 antibody precipitated WNT2. We also identified DVL (disheveled), AXIN, GSK-3β (glycogen synthase kinase-3β) and β-CATENIN. β-CATENIN is concentrated in the plasma membranes. DVL co-localizes with FZD9 and AXIN in the membranes, but GSK-3β has little co-localization with AXIN and β-CATENIN. Interestingly, β-CATENIN is highly co-localized with FZD9 and AXIN. CDH1 (E-cadherin) was also detected in the plasma membranes and cytoplasm, co-localized with β-CATENIN, and CDH1 antibody precipitated β-CATENIN. The results suggest that WNT2 could act through its receptor FZD9 to regulate the β-CATENIN pathway in cumulus cells, recruiting β-CATENIN into plasma membranes and promoting the formation of adherens junctions involving CDH1.
Published in: Molecular Human Reproduction 2009 15(1):11-17; doi:10.1093/molehr/gan070
https://ir.lib.uwo.ca/physpharmpub/1
oai:ir.lib.uwo.ca:physpharmpub-1002
2009-05-23T02:06:54Z
publication:anatomypub
publication:physpharm
publication:anatomy
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:paed
publication:pmid
publication:faculties
publication:paedpub
19259389
Oogenesis Defects in a Mutant Mouse Model of Oculodentodigital Dysplasia
Tong, Dan
Colley, Deanne
Thoo, Renee
Li, Tony Y.
Plante, Isabelle
Laird, Dale W.
Bai, Donglin
Kidder, Gerald M.
Article
2009-03-01T08:00:00Z
Connexin43
Gap junction
Granulosa cell
Folliculogenesis
ODDD
Primordial germ cell
Oocyte maturation
Fertilization
Disease Models & Mechanisms
Disease Models & Mechanisms
2
3/4
157
167
Medical Physiology
Obstetrics and Gynecology
The essential role of connexin43 (Cx43) during oogenesis has been demonstrated by
the severe germ cell deficiency and arrested folliculogenesis observed in Cx43
knockout mice. Recently, another mutant mouse strain became available (Gja1Jrt/+) that
carries the dominant loss-of-function Cx43 mutation, Cx43G60S. Gja1Jrt/+ mice display
features of the human disease, oculodentodigital dysplasia (ODDD), caused by
mutations in the GJA1 gene. We have used this new mutant strain to study how a
disease-linked Cx43 mutant affects oogenesis. We found that female mutant mice are
subfertile with significantly reduced mating success and small litters. The
phosphorylated species of the Cx43 protein are reduced in the mutant ovaries in
association with impaired trafficking and assembly of gap junctions in the membranes
of granulosa cells, confirming that the mutant protein acts dominantly on its wild-type
counterpart. Correspondingly, although starting with normal abundance of germ cells,
ovaries of the mutant mice contain significantly fewer pre-ovulatory follicles and do
not respond to superovulation by gonadotropins, which is at least partially due to
reduced proliferation and increased apoptosis of granulosa cells. We conclude that the
Gja1Jrt mutation has a dominant negative effect on Cx43 function in the ovary,
rendering the females subfertile. Given these findings, closer examination of
reproductive function in ODDD human females is warranted.
Published in: Dis Model Mech. 2009 Mar–Apr; 2(3-4): 157–167. Published online 2009 February 23. doi: 10.1242/dmm.000935. PMCID: PMC2650217
https://ir.lib.uwo.ca/physpharmpub/2
oai:ir.lib.uwo.ca:physpharmpub-1018
2009-08-18T23:43:22Z
publication:med
publication:physpharm
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:medpub
17214902
Mouse Preimplantation Embryo Responses to Culture Medium Osmolarity Include Increased Expression of CCM2 and p38 MAPK Activation
Fong, Barry
Watson, Patricia H.
Watson, Andrew J.
Article
2007-01-10T08:00:00Z
Animals
Blastocyst
Cells
Cultured
Culture Media
Enzyme Activation
Female
Fluorescent Antibody Technique
Indirect
Gene Expression
Mice
Microfilament Proteins
Microscopy
Confocal
Osmolar Concentration
Phosphorylation
Pregnancy
Reverse Transcriptase Polymerase Chain Reaction
p38 Mitogen-Activated Protein Kinases
BMC Developmental Biology
BMC Developmental Biology
7
2
Medical Physiology
Obstetrics and Gynecology
Background: Mechanisms that confer an ability to respond positively to environmental osmolarity are fundamental to ensuring embryo survival during the preimplantation period. Activation of p38 mitogen-activated protein kinase (MAPK) occurs following exposure to hyperosmotic treatment. Recently, a novel scaffolding protein called Osmosensing Scaffold for MEKK3 (OSM) was linked to p38 MAPK activation in response to sorbitol-induced hypertonicity. The human ortholog of OSM is cerebral cavernous malformation 2 (CCM2). The present study was conducted to investigate whether CCM2 is expressed during mouse preimplantation development and to determine whether this scaffolding protein is associated with p38 MAPK activation following exposure of preimplantation embryos to hyperosmotic environments.
Results: Our results indicate that Ccm2 along with upstream p38 MAPK pathway constituents (Map3k3, Map2k3, Map2k6, and Map2k4) are expressed throughout mouse preimplantation development. CCM2, MAP3K3 and the phosphorylated forms of MAP2K3/MAP2K6 and MAP2K4 were also detected throughout preimplantation development. Embryo culture in hyperosmotic media increased p38 MAPK activity in conjunction with elevated CCM2 levels.
Conclusion: These results define the expression of upstream activators of p38 MAPK during preimplantation development and indicate that embryo responses to hyperosmotic environments include elevation of CCM2 and activation of p38 MAPK.
Published in: BMC Developmental Biology, 2007, 7:2. doi: 10.1186/1471-213X-7-2
https://ir.lib.uwo.ca/physpharmpub/19
oai:ir.lib.uwo.ca:obsgynpub-1000
2009-08-20T00:20:45Z
publication:physpharm
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
12646061
Responsiveness of Bovine Cumulus-Oocyte-Complexes (COC) to Porcine and Recombinant Human FSH, and the Effect of COC Quality on Gonadotropin Receptor and Cx43 Marker Gene mRNAs During Maturation In Vitro
Calder, Michele D.
Caveney, Anita N.
Smith, Lawrence C.
Watson, Andrew J.
Article
2003-02-11T08:00:00Z
Animals
Cattle
Connexin 43
Culture Media
Serum-Free
Dose-Response Relationship
Drug
Estradiol
Female
Follicle Stimulating Hormone
Humans
Oocytes
Oogenesis
Ovarian Follicle
RNA
Messenger
Receptors
FSH
Receptors
LH
Recombinant Proteins
Swine
Reproductive Biology and Endocrinology
Reproductive Biology and Endocrinology
1
14
Medical Physiology
Obstetrics and Gynecology
Substantially less development to the blastocyst stage occurs in vitro than in vivo and this may be due to deficiencies in oocyte competence. Although a large proportion of bovine oocytes undergo spontaneous nuclear maturation, less is known about requirements for proper cytoplasmic maturation. Commonly, supraphysiological concentrations of FSH and LH are added to maturation media to improve cumulus expansion, fertilization and embryonic development. Therefore, various concentrations of porcine FSH (pFSH) and recombinant human FSH (rhFSH) were investigated for their effect on bovine cumulus expansion in vitro. Expression of FSHr, LHr and Cx43 mRNAs was determined in cumulus-oocyte complexes to determine whether they would be useful markers of oocyte competence. In serum-free media, only 1000 ng/ml pFSH induced marked cumulus expansion, but the effect of 100 ng/ml pFSH was amplified in the presence of 10% serum. In contrast, cumulus expansion occurred with 1 ng/ml rhFSH in the absence of serum. FSHr mRNA was highest at 0-6 h of maturation, then abundance decreased. Similarly, Cx43 mRNA expression was highest from 0-6 h but decreased by 24 h of maturation. However, the relative abundance of LHr mRNA did not change from 6-24 h of maturation. Decreased levels of FSHr, LHr and Cx43 mRNAs were detected in COCs of poorer quality. In conclusion, expansion of bovine cumulus occurred at low doses of rhFSH in serum-free media. In summary, FSHr, LHr and Cx43 mRNA abundance reflects COC quality and FSHr and Cx43 mRNA expression changes during in vitro maturation; these genes may be useful markers of oocyte developmental competence.
Published in: Reproductive Biology and Endocrinology, 2003, 1:14. doi: 10.1186/1477-7827-1-14
https://ir.lib.uwo.ca/obsgynpub/1
oai:ir.lib.uwo.ca:epidempub-1011
2009-09-26T00:26:51Z
publication:epidem
publication:obsgyn
publication:obsgynpub
publication:paed
publication:pmid
publication:faculties
publication:paedpub
publication:epidempub
15720709
Is There Value in Using Physician Billing Claims along with Other Administrative Health Care Data to Document the Burden of Adolescent Injury? An Exploratory Investigation with Comparison to Self-reports in Ontario, Canada
Potter, Beth K.
Manuel, Douglas
Speechley, Kathy N.
Gutmanis, Iris A.
Campbell, M. Karen
Koval, John J.
Article
2005-02-18T08:00:00Z
Activities of Daily Living
Adolescent
Adult
Cost of Illness
Female
Health Services Research
Hospitalization
Humans
Insurance Claim Review
Male
Office Visits
Ontario
Patient Credit and Collection
Population Surveillance
Prevalence
Self Disclosure
Wounds and Injuries
BMC Health Services Research
BMC Health Services Research
5
15
Biostatistics
Epidemiology
Health and Medical Administration
Background: Administrative health care databases may be particularly useful for injury surveillance, given that they are population-based, readily available, and relatively complete. Surveillance based on administrative data, though, is often restricted to injuries that result in hospitalization. Adding physician billing data to administrative data-based surveillance efforts may improve comprehensiveness, but the feasibility of such an approach has rarely been examined. It is also not clear how injury surveillance information obtained using administrative health care databases compares with that obtained using self-report surveys. This study explored the value of using physician billing data along with hospitalization data for the surveillance of adolescent injuries in Ontario, Canada. We aimed i) to document the burden of adolescent injury using administrative health care data, focusing on the relative contribution of physician billing information; and ii) to explore data quality issues by directly comparing adolescent injuries identified in administrative and self-report data.
Methods: The sample included adolescents aged 12 to 19 years who participated in the 1996-1997 cross-sectional Ontario Health Survey, and whose survey responses were linked to administrative health care datasets (N = 2067). Descriptive analysis was used to document the burden of injuries as a proportion of all physician care by gender and location of care, and to examine the distribution of both administratively-defined and self-reported activity-limiting injuries according to demographic characteristics. Administratively-defined and self-reported injuries were also directly compared at the individual level.
Results: Approximately 10% of physician care for the sample was identified as injury-related. While 18.8% of adolescents had self-reported injury in the previous year, 25.0% had documented administratively-defined injury. The distribution of injuries according to demographic characteristics was similar across data sources, but congruence was low at the individual level. Possible reasons for discrepancies between the data sources included recall errors in the survey data and errors in the physician billing data algorithm.
Conclusion: If further validated, physician billing data could be used along with hospital inpatient data to make an important and unique contribution to adolescent injury surveillance. The limitations inherent in different datasets highlight the need to continue rely on multiple information sources for complete injury surveillance information.
Published in: BMC Health Services Research, 2005, 5:15. doi: 10.1186/1472-6963-5-15
https://ir.lib.uwo.ca/epidempub/11
oai:ir.lib.uwo.ca:epidempub-1012
2009-09-26T00:33:18Z
publication:epidem
publication:obsgyn
publication:obsgynpub
publication:paed
publication:pmid
publication:faculties
publication:paedpub
publication:epidempub
16343342
Socioeconomic Status and Non-fatal Injuries among Canadian Adolescents: Variations across SES and Injury Measures
Potter, Beth K.
Speechley, Kathy N.
Koval, John J.
Gutmanis, Iris A.
Campbell, M. Karen
Manuel, Douglas
Article
2005-12-12T08:00:00Z
Adolescent
Adult
Canada
Cross-Sectional Studies
Family Characteristics
Female
Health Surveys
Humans
Income
Male
Observer Variation
Recreation
Residence Characteristics
Social Class
Wounds and Injuries
BMC Public Health
BMC Public Health
5
132
Biostatistics
Epidemiology
Obstetrics and Gynecology
Pediatrics
Background: While research to date has consistently demonstrated that socioeconomic status (SES) is inversely associated with injury mortality in both children and adults, findings have been less consistent for non-fatal injuries. The literature addressing SES and injury morbidity among adolescents has been particularly inconclusive. To explore potential explanations for these discrepant research findings, this study uniquely compared the relationship across different measures of SES and different causes of injury (recreation versus non-recreation injuries) within a sample of Canadian adolescents.
Methods: The sample included adolescent participants (aged 12 to 19 years) in the Canadian 1996-1997 cross-sectional National Population Health Survey (n = 6967). Five SES measures (household income, two neighbourhood-level proxy measures, two parental indicators) were examined in relation to three injury outcomes (total, recreation, and non-recreation injuries) using multivariable logistic regression.
Results: Among males, a clear relationship with injury was observed only for a parental SES index, which was positively associated with total and recreation injuries (odds ratios for the highest versus lowest SES category of 1.9 for total and 2.5 for recreation injuries). Among females, there was some evidence of a positive relationship between SES and injuries, particularly for a neighbourhood-level education measure with total and recreation injuries (odds ratios of 1.7 for total and 2.0 for recreation injuries).
Conclusion: The results suggest that differences related to the measures of SES chosen and the causes of injury under study may both contribute to discrepancies in past research on SES and non-fatal injuries among adolescents. To clarify the potential SES-injury relationship among youth, the findings emphasize a need for a greater understanding of the meaning and relevance of different SES measures for adolescents, and for an exploration of the pathways through which SES may be related to injury risk.
Published in: BMC Public Health, 2005, 5:132. doi: 10.1186/1471-2458-5-132
https://ir.lib.uwo.ca/epidempub/12
oai:ir.lib.uwo.ca:obsgynpub-1001
2009-12-21T18:54:38Z
publication:obsgyn
publication:obsgynpub
publication:faculties
Communication Skills Assessed at OSCE Are Not Affected by Participation in the Adolescent Healthy Sexuality Program
Penava, D. A.
Stanojevic, S.
Article
2002-01-01T08:00:00Z
Communication
Adolescents
OSCE
Standardized Patients
Healthy Sexuality
Medical Education Online
Medical Education Online
7
1
6
Obstetrics and Gynecology
Purpose: We proposed that first year medical students who voluntarily participated in
the Healthy Sexuality adolescent program would perform better than their peers on an adolescent
counseling station at the year-end OSCE (Objective Structured Clinical Examination). In addition we compared medical students’ communication skills at the time of the program as assessed by self, peers and participating adolescents.
Methods: Nineteen first year medical students voluntarily participated in the ongoing Healthy
Sexuality program. Adolescent participants, medical student peer participants and medical students assessed communication comp onents on a 7-point Likert scale at the end of the program. At the year-end OSCE, all first year medical students at the University of Western Ontario were assessed at an adolescent counseling station by a standardized patient (SP) and a physician exa miner. Statistical analysis examined differences between the two groups.
Results: Students who participated in the Healthy Sexuality program did not perform better than
their colleagues on the year-end OSCE. A statistically significant correlation between physician examiner and SP evaluations was found (r = 0.62). Adolescent participants communication skills assessments in the Healthy Sexuality Program demonstrated no significant correlation with medical student assessments (self or peer).
Conclusions: Voluntary intervention with adolescents did not result in improved communication skills at the structured year-end examination. Further investigation will be directed towards delineating differences between SP and physician examiner assessments.
https://ir.lib.uwo.ca/obsgynpub/2
oai:ir.lib.uwo.ca:obsgynpub-1002
2010-07-25T21:46:23Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
19155375
Recruiting Medical Students to Rural Practice: Perspectives of Medical Students and Rural Recruiters
Jutzi, Leah
Vogt, Kelly
Drever, Erin
Nisker, Jeff
Article
2009-01-01T08:00:00Z
Adult
Attitude of Health Personnel
Career Choice
Family Practice
Female
Humans
Male
Motivation
Ontario
Personnel Loyalty
Personnel Selection
Questionnaires
Rural Health Services
Students
Medical
Young Adult
Canadian Family Physician
Canadian Family Physician
55
1
72
73
Medicine and Health Sciences
Obstetrics and Gynecology
OBJECTIVE: To explore the strategies used by rural recruitment programs and their perceived influence on medical students.
DESIGN: Two original questionnaires delivered electronically, one to medical students and the other to recruiters in rural Ontario communities.
SETTING: Ontario, Canada.
PARTICPANTS: All 525 medical students enrolled in the Schulich School of Medicine & Dentistry at the University of Western Ontario in London and physician recruiters in 71 rural communities in Ontario were invited to participate in the study.
MAIN OUTCOME MEASURES: The factors that influence medical students to consider rural practice, strategies used by recruiters, and student perceptions of the ethical appropriateness of both.
RESULTS: The questionnaire was completed by 42.1% of medical students. Lifestyle considerations were an important influence for 93.1% of students. Themes from the qualitative analysis included the ethical appropriateness of financial considerations, economic forces, perceived disadvantages of rural practice, competition between communities, and lack of altruism. Responses were received from recruiters in 43.7% of communities; of those, 92.9% offered financial incentives to attract prospective physicians.
CONCLUSION: Financial and lifestyle considerations are important influences on medical students' choice to practise in rural communities. Most medical students felt incentive programs offered by rural communities were ethically appropriate.
https://ir.lib.uwo.ca/obsgynpub/3
oai:ir.lib.uwo.ca:obsgynpub-1003
2009-12-28T00:12:29Z
publication:anatomypub
publication:physpharm
publication:anatomy
publication:biochempub
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:paed
publication:pmid
publication:biochem
publication:faculties
publication:paedpub
19369450
Gonadotropin-releasing Hormone-regulated Chemokine Expression in Human Placentation
Cavanagh, P. Craig
Dunk, Caroline
Pampillo, Macarena
Szereszewski, Jacob M.
Taylor, Jay E.
Kahiri, Caroline
Han, Victor
Lye, Stephen
Bhattacharya, Moshmi
Babwah, Andy V.
Article
2009-07-01T07:00:00Z
Angiogenic Proteins
Buserelin
Cell Line
Transformed
Chemokine CXCL2
Chemokine CXCL6
Chemokines
Chemokines
CXC
Chemotaxis
Leukocyte
Culture Media
Conditioned
Female
Fluorescent Antibody Technique
Gene Expression Profiling
Gonadotropin-Releasing Hormone
Hormone Antagonists
Humans
Interleukin-8
Jurkat Cells
Killer Cells
Natural
Neovascularization
Physiologic
Oligonucleotide Array Sequence Analysis
Oligopeptides
Placentation
Pregnancy
Receptors
Interleukin-8A
Receptors
Interleukin-8B
Receptors
LHRH
Reverse Transcriptase Polymerase Chain Reaction
Signal Transduction
T-Lymphocytes
Time Factors
Trophoblasts
American Journal of Physiology-Cell Physiology
American Journal of Physiology-Cell Physiology
297
1
C17
C27
10.1152/ajpcell.00013.2009
Anatomy
Medical Physiology
Obstetrics and Gynecology
Pediatrics
Placental expression of gonadotropin-releasing hormone (GnRH)-I and II, as well as their cognate receptor, coincides with a period of extensive remodeling of the maternal-fetal interface, near the end of the first trimester of pregnancy. To further define the role of GnRH in human placentation, we performed a microarray screen of HTR-8/SVneo trophoblasts to identify GnRH-regulated genes and their roles in placentation. This screen revealed that GnRH regulates the expression of four angiogenic chemokines: CXCL2, CXCL3, CXCL6, and CXCL8. The microarray data were subsequently confirmed by an extensive Q-PCR time-course analysis. CXCL8, a representative chemokine, was selected for further analysis and shown to be strongly expressed by trophoblasts at the maternal-fetal interface of the human placenta, as well as to accumulate in a GnRH-dependent manner in trophoblast-conditioned media in culture. Trophoblasts were subsequently shown to recruit lymphocytes (Jurkat T cells and primary peripheral blood T and uterine natural killer cells) in chemotaxis assays and this was shown to be GnRH dependent. Furthermore, this recruitment was shown to occur via the release of CXCR1/CXCR2 interacting chemokines, such as the CXCLs investigated in this study. This novel regulation of chemokines by GnRH signaling demonstrates the role of GnRH in regulating the recruitment of lymphocytes to the decidua and the possibility of a direct effect on spiral artery remodeling via the release of proangiogenic chemokines and secondary effects via release of angiogenic factors by recruited lymphocytes.
https://ir.lib.uwo.ca/obsgynpub/4
oai:ir.lib.uwo.ca:obsgynpub-1004
2010-01-10T03:06:29Z
publication:biochempub
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:biochem
publication:faculties
19805400
Dual Effects of Superovulation: Loss of Maternal and Paternal Imprinted Methylation in a Dose-dependent Manner
Market-Velker, Brenna A.
Zhang, Liyue
Magri, Lauren S.
Bonvissuto, Anne C.
Mann, Mellissa R. W.
Article
2010-01-01T08:00:00Z
Superovulation
ovarian stimulation
assisted reproductive technology
Human Molecular Genetics
Human Molecular Genetics
19
1
36
51
10.1093/hmg/ddp465
Medical Biochemistry
Obstetrics and Gynecology
Superovulation or ovarian stimulation is currently an indispensable assisted reproductive technology (ART) for human subfertility/infertility treatment. Recently, increased frequencies of imprinting disorders have been correlated with ARTs. Significantly, for Angelman and Beckwith-Wiedemann Syndromes, patients have been identified where ovarian stimulation was the only procedure used by the couple undergoing ART. In many cases, increased risk of genomic imprinting disorders has been attributed to superovulation in combination with inherent subfertility. To distinguish between these contributing factors, carefully controlled experiments are required on spontaneously ovulated, in vivo-fertilized oocytes and their induced-ovulated counterparts, thereby minimizing effects of in vitro manipulations. To this end, effects of superovulation on genomic imprinting were evaluated in a mouse model, where subfertility is not a confounding issue. This work represents the first comprehensive examination of the overall effects of superovulation on imprinted DNA methylation for four imprinted genes in individual blastocyst stage embryos. We demonstrate that superovulation perturbed genomic imprinting of both maternally and paternally expressed genes; loss of Snrpn, Peg3 and Kcnq1ot1 and gain of H19 imprinted methylation were observed. This perturbation was dose-dependent, with aberrant imprinted methylation more frequent at the high hormone dosage. Superovulation is thought to primarily affect oocyte development; thus, effects were expected to be limited to maternal alleles. Our study revealed that maternal as well as paternal H19 methylation was perturbed by superovulation. We postulate that superovulation has dual effects during oogenesis, disrupting acquisition of imprints in growing oocytes, as well as maternal-effect gene products subsequently required for imprint maintenance during pre-implantation development.
https://ir.lib.uwo.ca/obsgynpub/5
oai:ir.lib.uwo.ca:paedpub-1017
2010-02-28T07:04:54Z
publication:biochempub
publication:obsgyn
publication:obsgynpub
publication:paed
publication:pmid
publication:biochem
publication:faculties
publication:paedpub
20159591
ATRX Partners with Cohesin and MeCP2 and Contributes to Developmental Silencing of Imprinted Genes in the Brain
Kernohan, Kristin D.
Jiang, Yan
Tremblay, Deanna C.
Bonvissuto, Anne C.
Eubanks, James H.
Mann, Mellissa R. W.
Bérubé, Nathalie G.
Article
2010-02-16T08:00:00Z
DNA
DEVBIO
HUMDISEASE
Developmental Cell
Developmental Cell
18
2
191
202
http://dx.doi.org/10.1016/j.devcel.2009.12.017
Medical Biochemistry
Obstetrics and Gynecology
Pediatrics
Human developmental disorders caused by chromatin dysfunction often display overlapping clinical manifestations, such as cognitive deficits, but the underlying molecular links are poorly defined. Here, we show that ATRX, MeCP2, and cohesin, chromatin regulators implicated in ATR-X, RTT, and CdLS syndromes, respectively, interact in the brain and colocalize at the H19 imprinting control region (ICR) with preferential binding on the maternal allele. Importantly, we show that ATRX loss of function alters enrichment of cohesin, CTCF, and histone modifications at the H19 ICR, without affecting DNA methylation on the paternal allele. ATRX also affects cohesin, CTCF, and MeCP2 occupancy within the Gtl2/Dlk1 imprinted domain. Finally, we show that loss of ATRX interferes with the postnatal silencing of the maternal H19 gene along with a larger network of imprinted genes. We propose that ATRX, cohesin, and MeCP2 cooperate to silence a subset of imprinted genes in the postnatal mouse brain.
https://ir.lib.uwo.ca/paedpub/18
oai:ir.lib.uwo.ca:obsgynpub-1005
2011-02-28T00:41:53Z
publication:physpharm
publication:biochempub
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:paed
publication:pmid
publication:biochem
publication:faculties
publication:paedpub
20171025
IL-6 and TNFalpha across the Umbilical Circulation in Term Pregnancies: Relationship with Labour Events
Duncombe, Greg
Veldhuizen, Ruud A. W.
Gratton, Robert J.
Han, Victor K. M.
Richardson, Bryan S.
Article
2010-02-01T08:00:00Z
IL-6
TNFalpha
Umbilical circulation
Term labour
Early Human Development
Early Human Development
86
2
113
117
http://dx.doi.org/10.1016/j.earlhumdev.2010.01.027
Medical Physiology
Obstetrics and Gynecology
Pediatrics
<p>OBJECTIVE: We have determined venous and arterial cord blood levels for IL-6 and TNFalpha at the time of delivery to assess gestational tissue versus fetal sources in labouring and non-labouring patients at term, and the relationship to labour events. METHODS: Fifty-five patients were studied (elective cesarean section n=24, and labouring n=31) with blood sampling from a clamped segment of cord after delivery of the fetus and from the cord at its insertion into the placenta after delivery of the placenta, with subsequent measurement of blood gases, pH, IL-6 and TNFalpha. RESULTS: Umbilical cord levels for IL-6 were increased by 4 fold in low risk labouring patients, and a further 6 fold when showing histologic chorioamnionitis, but with no evident effect of nuchal cord with 'variable' fetal heart rate decelerations, fetal acidemia, nor of labour duration. IL-6 levels from the cord at its insertion into the placenta were generally higher than those from the respective umbilical levels indicating that placental release of IL-6 into cord blood must be occurring. However, a consistent venoarterial difference for IL-6 and thereby a net flux from the placenta could not be demonstrated. TNFalpha levels for both patient groups were uniformly low for all of the cord measurements with no significant differences noted. CONCLUSION: Umbilical cord levels for IL-6 are increased in low risk labouring patients at term in the absence of evident infection which likely involves both gestational tissue and fetal contributions. Cord levels for IL-6 are further increased in low risk labouring patients showing histologic chorioamnionitis which might then contribute to newborn morbidity in these pregnancies.</p>
https://ir.lib.uwo.ca/obsgynpub/6
oai:ir.lib.uwo.ca:obsgynpub-1006
2010-04-05T20:52:01Z
publication:anatomypub
publication:anatomy
publication:obsgyn
publication:oncpub
publication:obsgynpub
publication:pmid
publication:faculties
publication:onc
20187934
Constitutive Activation of BMP Signalling Abrogates Experimental Metastasis of OVCA429 Cells via Reduced Cell Adhesion
Shepherd, Trevor G.
Mujoomdar, Michelle L.
Nachtigal, Mark W.
Article
2010-02-26T08:00:00Z
ovarian cancer
metastasis
BMP signalling
OVCA429 cell
cell adhesion
Journal of Ovarian Research
Journal of Ovarian Research
3
5
http://dx.doi.org/10.1186/1757-2215-3-5
Anatomy
Obstetrics and Gynecology
Oncology
BACKGROUND: Activation of bone morphogenetic protein (BMP)4 signalling in human ovarian cancer cells induces a number of phenotypic changes in vitro, including altered cell morphology, adhesion, motility and invasion, relative to normal human ovarian surface epithelial cells. From these in vitro analyses, we had hypothesized that active BMP signalling promotes the metastatic potential of ovarian cancer.
METHODS: To test this directly, we engineered OVCA429 human ovarian cancer cells possessing doxycycline-inducible expression of a constitutively-active mutant BMP receptor, ALK3QD, and administered these cells to immunocompromised mice. Further characterization was performed in vitro to address the role of activated BMP signalling on the EOC phenotype, with particular emphasis on epithelial-mesenchymal transition (EMT) and cell adhesion.
RESULTS: Unexpectedly, doxycycline-induced ALK3QD expression in OVCA429 cells reduced tumour implantation on peritoneal surfaces and ascites formation when xenografted into immunocompromised mice by intraperitoneal injection. To determine the potential mechanisms controlling this in vivo observation, we followed with several cell culture experiments. Doxycycline-induced ALK3QD expression enhanced the refractile, spindle-shaped morphology of cultured OVCA429 cells eliciting an EMT-like response. Using in vitro wound healing assays, we observed that ALK3QD-expressing cells migrated with long, cytoplasmic projections extending into the wound space. The phenotypic alterations of ALK3QD-expressing cells correlated with changes in specific gene expression patterns of EMT, including increased Snail and Slug and reduced E-cadherin mRNA expression. In addition, ALK3QD signalling reduced beta1- and beta3-integrin expression, critical molecules involved in ovarian cancer cell adhesion. The combination of reduced E-cadherin and beta-integrin expression correlates directly with the reduced EOC cell cohesion in spheroids and reduced cell adhesion to the extracellular matrix substrates fibronectin and vitronectin that was observed.
CONCLUSIONS: We propose that the key steps of ovarian cancer metastasis, specifically cell cohesion of multicellular aggregates in ascites and cell adhesion for reattachment to secondary sites, may be inhibited by overactive BMP signalling, thereby decreasing the ultimate malignant potential of ovarian cancer in this model system.
https://ir.lib.uwo.ca/obsgynpub/7
oai:ir.lib.uwo.ca:philosophypub-1146
2010-04-20T01:46:45Z
publication:med
publication:epidem
publication:obsgyn
publication:philosophy
publication:obsgynpub
publication:pmid
publication:rotman
publication:rotmanpub
publication:institutes
publication:faculties
publication:philosophypub
publication:medpub
publication:epidempub
18611303
Ethical Issues Associated With the Introduction of New Surgical Devices, or Just Because We Can, Doesn’t Mean We Should
Ross, Sue
Robert, Magali
Harvey, Marie-Andrée
Farrell, Scott
Schulz, Jane
Wilkie, David
Lovatsis, Danny
Epp, Annette
Easton, Bill
McMillan, Barry
Schachter, Joyce
Gupta, Chander
Weijer, Charles
Article
2008-06-01T07:00:00Z
Clinical ethics
Risk Assessment
Safety
Journal of Obstetrics and Gynaecology Canada
Journal of Obstetrics and Gynaecology Canada
30
6
508
513
Bioethics and Medical Ethics
Philosophy
Surgical devices are often marketed before there is good evidence of their safety and effectiveness. Our paper discusses the ethical issues associated with the early marketing and use of new surgical devices from the perspectives of the six groups most concerned. Health Canada, which is responsible for licensing new surgical devices, should amend their requirements to include rigorous clinical trials that provide data on effectiveness and safety for each new product before it is marketed. Industry should comply with all Health Canada requirements to obtain licenses for new products. Until Health Canada requires effectiveness and safety data, industry should cooperate with physicians in appropriate studies before releasing new products and should make balanced presentations of all the available evidence. Surgeons should, before using a new surgical device, assess the evidence on its effectiveness and safety and ensure they are properly trained and competent in using the device. Surgeons should provide their patients with an evaluation of the available evidence and inform them about possible complications and the surgeon's level of experience with the new device. Patients, who should be given an honest evaluation of the available evidence, possible complications, and the surgeon's experience, should be encouraged to evaluate the evidence and information to their own satisfaction to ensure that fully informed consent is given. Health institutions, responsible for regulating practice within their walls, should review new devices for safety, effectiveness, and economic impacts, before allowing their use. They should also limit the use of new surgical devices to surgeons trained and competent in the new technology. Professional societies should provide guidance on the early adoption of new surgical devices and technologies. We urge all those involved in the development, licensing, and use of new surgical devices to aim for higher ethical standards to protect the health and safety of patients requiring surgery. The lowest acceptable ethical standard would require device manufacturers to provide surgeons with accurate and timely information on the efficacy and safety of their products, allowing surgeons and patients to evaluate the evidence (and the significance of information not yet available) before surgery.
https://ir.lib.uwo.ca/philosophypub/147
oai:ir.lib.uwo.ca:obsgynpub-1007
2010-06-21T00:03:30Z
publication:physpharm
publication:biochempub
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:biochem
publication:faculties
20453524
Cerebral Protein Synthesis in the Ovine Fetus Near Term with Amino Acid Infusion and Insulin-Induced Hypoaminoacidemia.
Maclachlan, James N.
McCallum, Jeremy D.
Smith, Norman
Matushewski, Brad
Richardson, Bryan S.
Article
2010-05-04T07:00:00Z
Fetal sheep
Cerebral protein synthesis
Amino acids
Insulin
Neonatology
Neonatology
98
4
297
304
http://dx.doi.org/10.1159/000291416
Medical Biochemistry
Medical Pharmacology
Medical Physiology
Obstetrics and Gynecology
Background: Studies during early development have shown that the precursor availability of amino acids directly affects protein synthesis both at the whole-body level and for select organ tissues, although this has not been studied for the brain.
Objective: We utilized a mixed amino acid infusate and an insulin euglycemic clamp technique in the ovine fetus near term, with increases and decreases in circulating amino acid levels of approximately 30 to 40% on average, and determined the impact on cerebral protein synthesis. Methods: Fetal sheep received a 6-hour infusion of Primene(R) 10% (amino acid infusate group) or a co-infusion of insulin and 10% dextrose (insulin/dextrose infusate group) together with a continuous infusion of L-[1-(13)C]-leucine. Measurements were obtained for fetal plasma leucine enrichment at steady-state and brain tissue intracellular free and protein-bound leucine enrichment at necropsy, followed by the determination of cerebral protein fractional synthetic rates (FSR).
Results: Protein FSR for the cerebral cortex averaged approximately 58 and approximately 39%/day when using the intracellular free and plasma enrichment values for the precursor pool measurements, respectively, providing for maximal and minimal FSR values, and with little difference between the amino acid and insulin/dextrose groups, although significantly higher than respective values for the cerebellum.
Conclusion: Accordingly, there was no evidence of a differential effect of increases versus decreases in circulating amino acids on cerebral protein synthesis as studied, which may be attributed to the saturable nature of the blood-brain barrier transporters for amino acids.
https://ir.lib.uwo.ca/obsgynpub/8
oai:ir.lib.uwo.ca:obsgynpub-1008
2010-06-21T00:20:06Z
publication:biochempub
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:biochem
publication:faculties
20452320
Multiple Epigenetic Modifiers Induce Aggressive Viral Extinction in Extraembryonic Endoderm Stem Cells
Golding, Michael C.
Zhang, Liyue
Mann, Mellissa R. W.
Article
2010-05-07T07:00:00Z
Epigenetic Modifiers
Extraembryonic Endoderm Stem Cells
Cell Stem Cell
Cell Stem Cell
6
5
457
467
http://dx.doi.org/10.1016/j.stem.2010.03.014
Medical Biochemistry
Obstetrics and Gynecology
To prevent insertional mutagenesis arising from retroviral reactivation, cells of embryonic origin possess a unique capacity to silence retroviruses. Given the distinct modes of X chromosome inactivation between embryonic and extraembryonic lineages, we investigated paradigms of viral extinction. We show that trophectoderm stem cells do not silence retroviral transcription, whereas extraembryonic endoderm stem cells aggressively extinguish proviral transcription, even more rapidly than do embryonic stem cells. By using a short hairpin RNA library, we identified epigenetic modifiers of retroviral extinction in extraembryonic endoderm stem cells. Multiple chromatin remodeling and polycomb repressor complex proteins act to modulate integrated, as well as endogenous, retroviral element silencing, with a subset of factors displaying differential effects between stem cell types. Furthermore, our data suggest that small RNAs play a role in this process through interactions with the Argonaute family. Our results further the understanding of mechanisms regulating retroviral transcription in different stem cell lineages.
https://ir.lib.uwo.ca/obsgynpub/9
oai:ir.lib.uwo.ca:philosophypub-1333
2010-07-24T00:35:21Z
publication:womenspub
publication:womens
publication:obsgyn
publication:philosophy
publication:obsgynpub
publication:rotman
publication:rotmanpub
publication:institutes
publication:faculties
publication:philosophypub
The 'Healthy' Embryo: Social, Biomedical, Legal and Philosophical Perspectives
Nisker, Jeff
Baylis, Françoise
Karpin, Isabel
McLeod, Carolyn
Mykitiuk, Roxanne
Book
2010-01-01T08:00:00Z
Human embryo
Human reproductive technology
Bioethics
Bioethics and Medical Ethics
Feminist, Gender, and Sexuality Studies
Philosophy
Public attention on embryo research has never been greater. Modern reproductive medicine technology and the use of embryos to generate stem cells ensure that this will continue to be a topic of debate and research across many disciplines. This multidisciplinary book explores the concept of a 'healthy' embryo, its implications on the health of children and adults, and how perceptions of what constitutes child and adult health influence the concept of embryo 'health'. The concept of human embryo health is considered from preconception to pre-implantation genetic diagnosis to recent foetal surgical approaches. Burgeoning capacities in both genetic and reproductive science and their clinical implications have catalysed the necessity to explore the concept of a 'healthy' embryo. The authors are from five countries and 13 disciplines in the social sciences, humanities, biological sciences and medicine, ensuring that the book has a broad coverage and approach.
Dr. Carolyn McLeod was a co-editor of this book. It is not available online here. If you are affiliated with The University of Western Ontario, please use the Shared Library Catalogue's <a href="http://alpha.lib.uwo.ca/" >Classic Search</a> to check whether the book is available in Western Libraries.<br>
If you are not affiliated with The University of Western Ontario, search <a href="http://www.worldcat.org/" >WorldCat</a> to find out where you can get access to the book.
https://ir.lib.uwo.ca/philosophypub/331
oai:ir.lib.uwo.ca:obsgynpub-1009
2010-07-25T21:39:56Z
publication:womenspub
publication:womens
publication:obsgyn
publication:philosophy
publication:obsgynpub
publication:pmid
publication:rotman
publication:rotmanpub
publication:institutes
publication:faculties
publication:philosophypub
16921478
Choice in Fertility Preservation in Girls and Adolescent Women with Cancer
Nisker, Jeffrey
Baylis, Françoise
McLeod, Carolyn
Article
2006-10-01T07:00:00Z
Cryopreservation
Fertility
Fertilization in Vitro
Infertility
Neoplasms
Oocytes
Cancer
Cancer
107
7 Suppl
1686
1689
http://dx.doi.org/10.1002/cncr.22106
Bioethics and Medical Ethics
Feminist, Gender, and Sexuality Studies
Obstetrics and Gynecology
Oncology
Philosophy
With the cure rate for many pediatric malignancies now between 70% and 90%, infertility becomes an increasingly important issue. Strategies for preserving fertility in girls and adolescent women occur in two distinct phases. The first phase includes oophorectomy (usually unilateral) and cryopreservation of ovarian cortex slices or individual oocytes; ultrasound-guided needle aspiration of oocytes, with or without in vitro maturation (IVM), followed by cryopreservation; and ovarian autografting to a distant site. The second phase occurs if the woman chooses to pursue pregnancy, and includes IVM of the oocytes, followed by in vitro fertilization (IVF) and transfer of any created embryos to the woman's uterus (or to a surrogate's uterus if the cancer patient's uterus has been surgically removed or the endometrium destroyed by radiotherapy). For ovarian autografting, the woman would undergo menotropin ovarian stimulation and retrieval of matured oocytes (likely by laparotomy, but possibly by ultrasound-guided needle aspiration if the ovary is positioned in an inaccessible location). The ethical challenges with each of these phases are many of fertility preservation and include issues of informed choice (consent or refusal). The lack of proven benefit with these strategies and the associated potential physical and psychological harms require careful attention to the key elements of informed choice, which include decisional capacity, disclosure, understanding and voluntariness, and to the benefits of in-depth counseling to promote free and informed choice at a time that is emotionally difficult for the decision makers.
https://ir.lib.uwo.ca/obsgynpub/10
oai:ir.lib.uwo.ca:philosophypub-1359
2010-08-02T05:23:23Z
publication:womenspub
publication:womens
publication:obsgyn
publication:philosophy
publication:obsgynpub
publication:rotman
publication:rotmanpub
publication:institutes
publication:faculties
publication:philosophypub
Nothing Extreme about Protecting Fresh Embryos
Baylis, Françoise
McLeod, Carolyn
Nisker, Jeff
Sherwin, Susan
Response or Comment
2007-01-16T08:00:00Z
Embryos
The Globe and Mail
The Globe and Mail
A15
A15
Bioethics and Medical Ethics
Feminist, Gender, and Sexuality Studies
Philosophy
https://ir.lib.uwo.ca/philosophypub/356
oai:ir.lib.uwo.ca:obsgynpub-1010
2010-08-05T05:59:08Z
publication:biochempub
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:biochem
publication:faculties
20547750
Domain-specific Response of Imprinted Genes to Reduced DNMT1
Weaver, Jamie R.
Sarkisian, Garnik
Krapp, Christopher
Mager, Jesse
Mann, Mellissa R. W.
Bartolomei, Marisa S.
Article
2010-08-01T07:00:00Z
Imprinted Genes
Reduced DNMT1
Molecular and Cellular Biology
Molecular and Cellular Biology
30
16
3916
3928
http://dx.doi.org/10.1128/MCB.01278-09
Biochemistry
Molecular Biology
Obstetrics and Gynecology
Imprinted genes are expressed in a monoallelic, parent-of-origin-specific manner. Clusters of imprinted genes are regulated by imprinting control regions (ICRs) characterized by DNA methylation of one allele. This methylation is critical for imprinting; a reduction in the DNA methyltransferase DNMT1 causes a widespread loss of imprinting. To better understand the role of DNA methylation in the regulation of imprinting, we characterized the effects of Dnmt1 mutations on the expression of a panel of imprinted genes in the embryo and placenta. We found striking differences among imprinted domains. The Igf2 and Peg3 domains showed imprinting perturbations with both null and partial loss-of-function mutations, and both domains had pairs of coordinately regulated genes with opposite responses to loss of DNMT1 function, suggesting these domains employ similar regulatory mechanisms. Genes in the Kcnq1 domain were less sensitive to the absence of DNMT1. Cdkn1c exhibited imprinting perturbations only in null mutants, while Kcnq1 and Ascl2 were largely unaffected by a loss of DNMT1 function. These results emphasize the critical role for DNA methylation in imprinting and reveal the different ways it controls gene expression.
https://ir.lib.uwo.ca/obsgynpub/11
oai:ir.lib.uwo.ca:biochempub-1117
2010-08-13T23:54:10Z
publication:biochempub
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:biochem
publication:faculties
20594350
A Novel Mechanism of Cell Growth Regulation by Cell Cycle and Apoptosis Regulatory Protein (CARP)-1
Jiang, Yan
Puliyappadamba, Vineshkumar T
Zhang, Liyue
Wu, Wenjuan
Wali, Anil
Yaffe, Michael B.
Fontana, Joseph A.
Rishi, Arun K.
Article
2010-07-01T07:00:00Z
Cell growth regulation
Cell Cycle and Apoptosis Regulatory Protein
(CARP)-1
Journal of Molecular Signaling
5
7
http://dx.doi.org/10.1186/1750-2187-5-7
Biochemistry
Obstetrics and Gynecology
BACKGROUND: CARP-1/CCAR1, a perinuclear phospho-protein, regulates signaling by adriamycin, steroids, or growth factors. However, intracellular events that regulate CARP-1-dependent cell growth are not fully understood.
RESULTS: Here we investigated whether CARP-1 is involved in signaling induced by the protein kinase A inhibitor H89. Treatments of human breast cancer cells with H89 resulted in apoptosis that involved enhanced CARP-1 threonine phosphorylation and expression. Depletion of CARP-1, on the other hand, abrogates apoptosis induced by H89. CARP-1 binds with signal transducer TAZ and over-expression of TAZ inhibits apoptosis by CARP-1. CARP-1 (651-759) interacts with a novel, N-terminal epitope of TAZ. H89 treatment stimulates threonine phosphorylation of CARP-1 (651-759), while substitution of threonine667 to alanine interferes with its binding with TAZ and apoptosis by H89. In addition, expression of wild type or CARP-1 (651-759) causes loss of c-myc expression due, in part, to suppression of c-myc transcription.
CONCLUSIONS: CARP-1 threonine667 regulates H89-dependent signaling by a novel pathway that involves modulation of CARP-1 interaction with TAZ and transcriptional down-regulation of c-myc.
https://ir.lib.uwo.ca/biochempub/115
oai:ir.lib.uwo.ca:obsgynpub-1011
2010-09-08T00:42:15Z
publication:biochempub
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:biochem
publication:faculties
20704727
Novel Cis-trans Interactions Are Involved in Post-transcriptional Regulation of Cyclin-dependent Kinase Inhibitor p21WAF1/CIP1 mRNA
Zhang, Liyue
Wali, Anil
Fontana, Joseph A.
Dawson, Marcia I.
Rishi, Arun K.
Article
2010-08-12T07:00:00Z
p21WAF1/CIP1 mRNA
Journal of Molecular Signaling
Journal of Molecular Signaling
5
12
http://dx.doi.org/10.1186/1750-2187-5-12
Biochemistry, Biophysics, and Structural Biology
Obstetrics and Gynecology
BACKGROUND: A variety of pathways target CDKI p21WAF1/CIP1 expression at transcriptional, post-transcriptional as well as translational levels. We previously found that cell growth suppressing retinoid CD437 enhanced expression of p21WAF1/CIP1 and DNA damage inducible GADD45 proteins in part by elevating their mRNA stability.
RESULTS: Here, we investigated molecular mechanisms of CD437-dependent post-transcriptional regulation of p21WAF1/CIP1 expression. By utilizing MDA-MB-468 HBC cells expressing chimeric rabbit beta-globin-p21WAF1/CIP1 transcripts we mapped multiple CD437-responsive sequences located within positions 1195 to 1795 of the 3'-untranslated region of p21WAF1/CIP1 mRNA. Several cytoplasmic proteins present in MDA-MB-468, MCF-7 HBC as well as HL-60R leukemia cells bound specifically, in vitro, with these CD437-responsive sequences. CD437 treatment of cells resulted in elevated binding of ~85 kD and ~55 kD cytoplasmic proteins with putative CD437-responsive sequences. A 12 nt RNA sequence (5'-UGUGGUGGCACA-3') present within CD437-responsive region of p21WAF1/CIP1 mRNA displayed specific and elevated binding with the above noted proteins. Treatment of cells with ActD or CHX prior to CD437 exposure did not abrogate RNA-protein interactions. However, treatment of cytoplasmic protein extracts with proteinase K or alkaline phosphatase resulted in loss of RNA-protein interactions.
CONCLUSIONS: CD437 regulates cell growth in part by regulating stability of p21WAF1/CIP1 mRNA that involves specific RNA-protein interactions that are phosphorylation-dependent, while not requiring nascent transcription or protein synthesis.
https://ir.lib.uwo.ca/obsgynpub/12
oai:ir.lib.uwo.ca:kinpub-1009
2011-03-18T23:18:33Z
publication:kin
publication:anatomy
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:paed
publication:pmid
publication:faculties
publication:anatomypub
publication:kinpub
publication:rwkex
publication:epidem
publication:epidempub
publication:paedpub
21184681
Increased Incidence of Glucose Disorders during Pregnancy Is Not Explained by Pre-pregnancy Obesity in London, Canada
Davenport, Margie H.
Campbell, M. Karen
Mottola, Michelle F.
Article
2010-12-24T08:00:00Z
Glucose disorders
Pregnancy
Pre-pregnancy obesity
London
Ontario
Canada
BMC Pregnancy and Childbirth
BMC Pregnancy and Childbirth
10
85
http://dx.doi.org/10.1186/1471-2393-10-85
Epidemiology
Kinesiology
Medical Anatomy
Obstetrics and Gynecology
Pediatrics
<p>BACKGROUND: The increasing incidence of impaired glucose tolerance (IGT), gestational diabetes (GDM) and type 2 diabetes (T2D) during pregnancy was hypothesized to be associated with increases in pre-pregnancy body mass index (BMI). The aims were to 1) determine the prevalence of IGT/GDM/T2 D over a 10 year period; 2) examine the relationship between maternal overweight/obesity and IGT/GDM/T2D; and 3) examine the extent to which maternal metabolic complications impact maternal and fetal pregnancy outcomes.</p>
<p>METHODS: Data arose from a perinatal database which contains maternal characteristics and perinatal outcome for all singleton infants born in London, Canada between January 1, 2000 and December 31, 2009. Univariable and multivariable odds ratios (OR) were estimated using logistic regression with IGT/GDM/T2 D being the outcome of interest.</p>
<p>RESULTS: A total of 36,597 women were included in the analyses. Population incidence of IGT, GDM and T2 D rose from 0.7%, 2.9% and 0.5% in 2000 to 1.2%, 4.2% and 0.9% in 2009. The univariable OR for IGT, GDM and T2 D were 1.65, 1.52 and 2.06, respectively, over the ten year period. After controlling for maternal age, parity and pre-pregnancy BMI the OR did not decrease. Although there was a positive relationship between pre-pregnancy BMI and prevalence of IGT/GDM/T2 D, this did not explain the time trends in the latter. Diagnosis of IGT/GDM/T2 D increased the risk of having an Apgar score <7 at 5>minutes, which was partially explained by gestational hypertension, high placental ratio, gestational age and large for gestational age babies.</p>
<p>CONCLUSIONS: We found a significant increase in the incidence of IGT/GDM/T2 D for the decade between 2000-2009 which was not explained by rising prevalence of maternal overweight/obesity.</p>
https://ir.lib.uwo.ca/kinpub/10
oai:ir.lib.uwo.ca:physpharmpub-1049
2011-05-24T01:15:21Z
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:physpharm
publication:biochempub
publication:biochem
18519738
P2X7 Receptors on Osteoblasts Couple to Production of Lysophosphatidic Acid: A Signaling Axis Promoting Osteogenesis
Panupinthu, Nattapon
Rogers, Joseph T.
Zhao, Lin
Solano-Flores, Luis Pastor
Possmayer, Fred
Sims, Stephen M.
Dixon, S. Jeffrey
Article
2008-06-02T07:00:00Z
Animals
Chondrocytes
Cyclooxygenase Inhibitors
Dinoprostone
Gene Expression Regulation
Lipids
Lysophospholipids
Transgenic Mice
Biological Models
Osteoblasts
Osteogenesis
Rats
Receptors
Purinergic P2
Receptors
Purinergic P2X7
Signal Transduction
Skull
The Journal of Cell Biology
The Journal of Cell Biology
181
5
859
871
http://dx.doi.org/10.1083/jcb.200708037
Medical Biochemistry
Medical Physiology
Obstetrics and Gynecology
Pharmacy and Pharmaceutical Sciences
<p>Nucleotides are released from cells in response to mechanical stimuli and signal in an autocrine/paracrine manner through cell surface P2 receptors. P2rx7-/- mice exhibit diminished appositional growth of long bones and impaired responses to mechanical loading. We find that calvarial sutures are wider in P2rx7-/- mice. Functional P2X7 receptors are expressed on osteoblasts in situ and in vitro. Activation of P2X7 receptors by exogenous nucleotides stimulates expression of osteoblast markers and enhances mineralization in cultures of rat calvarial cells. Moreover, osteogenesis is suppressed in calvarial cell cultures from P2rx7-/- mice compared with the wild type. P2X7 receptors couple to production of the potent lipid mediators lysophosphatidic acid (LPA) and prostaglandin E2. Either an LPA receptor antagonist or cyclooxygenase (COX) inhibitors abolish the stimulatory effects of P2X7 receptor activation on osteogenesis. We conclude that P2X7 receptors enhance osteoblast function through a cell-autonomous mechanism. Furthermore, a novel signaling axis links P2X7 receptors to production of LPA and COX metabolites, which in turn stimulate osteogenesis.</p>
https://ir.lib.uwo.ca/physpharmpub/49
oai:ir.lib.uwo.ca:physpharmpub-1054
2011-05-24T01:58:02Z
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:physpharm
publication:biochempub
publication:biochem
17135244
P2X7 Nucleotide Receptors Mediate Blebbing in Osteoblasts through a Pathway Involving Lysophosphatidic Acid
Panupinthu, Nattapon
Zhao, Lin
Possmayer, Fred
Ke, Hua Z.
Sims, Stephen M.
Dixon, S. Jeffrey
Article
2007-02-02T08:00:00Z
Adenosine Triphosphate
Animals
Newborn Animals
Genetic Crosses
Intracellular Signaling Peptides and Proteins
Lysophospholipids
Mice
Mice
Inbred C57BL
Mice
Inbred DBA
Osteoblasts
Phospholipases
Protein-Serine-Threonine Kinases
Rats
Receptors
Purinergic P2
Receptors
Purinergic P2X7
rho-Associated Kinases
The Journal of Biological Chemistry
The Journal of Biological Chemistry
282
5
3403
3412
http://dx.doi.org/10.1074/jbc.M605620200
Medical Biochemistry
Medical Physiology
Obstetrics and Gynecology
Pharmacy and Pharmaceutical Sciences
<p>Extracellular nucleotides, released in response to mechanical or inflammatory stimuli, signal through P2 receptors in many cell types, including osteoblasts. P2X7 receptors are ATP-gated cation channels that can induce formation of large membrane pores. Disruption of the gene encoding the P2X7 receptor leads to decreased periosteal bone formation and insensitivity of the skeleton to mechanical stimulation. Our purpose was to investigate signaling pathways coupled to P2X7 activation in osteoblasts. Live cell imaging showed that ATP or 2 ',3 '-O-(4-benzoylbenzoyl)-ATP (BzATP), but not UTP, UDP, or 2-methylthio-ADP, induced dynamic membrane blebbing in calvarial osteoblasts. Blebbing was observed in calvarial cells from wildtype but not P2X7 knock-out mice. P2X7 receptors coupled to activation of phospholipase D and A2, inhibition of which suppressed BzATP-induced blebbing. Activation of these phospholipases leads to production of lysophosphatidic acid (LPA). LPA caused dynamic blebbing in osteoblasts from both wild-type and P2X7 knock-out mice, similar to that induced by BzATP in wildtype cells. However, LPA-induced blebbing was more rapid in onset and was not affected by inhibition of phospholipase D or A2. Blockade or desensitization of LPA receptors suppressed blebbing in response to LPA and BzATP, without affecting P2X7-stimulated pore formation. Thus, LPA functions downstream of P2X7 receptors to induce membrane blebbing. Furthermore, inhibition of Rho-associated kinase abolished blebbing induced by both BzATP and LPA. In summary, we propose a novel signaling axis that links P2X7 receptors through phospholipases to production of LPA and activation of Rho-associated kinase. This pathway may contribute to P2X7-stimulated osteogenesis during skeletal development and mechanotransduction.</p>
https://ir.lib.uwo.ca/physpharmpub/54
oai:ir.lib.uwo.ca:obsgynpub-1012
2011-05-30T18:20:37Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:biochempub
publication:apmaths
publication:biochem
publication:apmathspub
20702853
Side-by-Side Comparison of Five Commercial Media Systems in a Mouse Model: Suboptimal In Vitro Culture Interferes with Imprint Maintenance
Market-Velker, B. A.
Fernandes, A. D.
Mann, M. R. W.
Article
2010-12-01T08:00:00Z
Animals
Blastocyst
Culture Media
DNA Methylation
Ectogenesis
Embryo Culture Techniques
Female
Fertilization
Fertilization in Vitro
Genomic Imprinting
Kruppel-Like Transcription Factors
Male
Mice
Mice
Inbred C57BL
RNA
Untranslated
Reproductive Techniques
Assisted
Statistics as Topic
Superovulation
snRNP Core Proteins
Biology of Reproduction
Biology of Reproduction
83
6
938
950
http://dx.doi.org/10.1095/biolreprod.110.085480
Applied Mathematics
Biochemistry, Biophysics, and Structural Biology
Obstetrics and Gynecology
<p>Assisted reproductive technologies (ARTs) are becoming increasingly prevalent and are generally considered to be safe medical procedures. However, evidence indicates that embryo culture may adversely affect the developmental potential and overall health of the embryo. One of the least studied but most important areas in this regard is the effects of embryo culture on epigenetic phenomena, and on genomic imprinting in particular, because assisted reproduction has been linked to development of the human imprinting disorders Angelman and Beckwith-Wiedemann syndromes. In this study, we performed side-by-side comparisons of five commercial embryo culture systems (KSOMaa, Global, Human Tubal Fluid, Preimplantation 1/Multiblast, and G1v5PLUS/G2v5PLUS) in relation to a best-case (in vivo-derived embryos) and a worst-case (Whitten culture) scenario. Imprinted DNA methylation and expression were examined at three well-studied loci, H19, Peg3, and Snrpn, in mouse embryos cultured from the 2-cell to the blastocyst stage. We show that embryo culture in all commercial media systems resulted in imprinted methylation loss compared to in vivo-derived embryos, although some media systems were able to maintain imprinted methylation levels more similar to those of in vivo-derived embryos in comparison to embryos cultured in Whitten medium. However, all media systems exhibited loss of imprinted H19 expression comparable to that using Whitten medium. Combined treatment of superovulation and embryo culture resulted in increased perturbation of genomic imprinting, above that from culture alone, indicating that multiple ART procedures further disrupt genomic imprinting. These results suggest that time in culture and number of ART procedures should be minimized to ensure fidelity of genomic imprinting during preimplantation development.</p>
https://ir.lib.uwo.ca/obsgynpub/13
oai:ir.lib.uwo.ca:epidempub-1053
2011-08-31T23:21:46Z
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:rwkex
publication:epidem
publication:epidempub
21672236
The Effects of Publishing Emergency Department Wait Time on Patient Utilization Patterns in a Community with Two Emergency Department Sites: A Retrospective, Quasi-experiment Design
Xie, Bin
Youash, Sabrina
Article
2011-06-14T07:00:00Z
Emergency department
Wait time
Patient utilization pattern
International Journal of Emergency Medicine
International Journal of Emergency Medicine
4
29
http://dx.doi.org/10.1186/1865-1380-4-29
Biostatistics
Epidemiology
Obstetrics and Gynecology
<p>BACKGROUND: Providing emergency department (ED) wait time information to the public has been suggested as a mechanism to reduce lengthy ED wait times (by enabling patients to select the ED site with shorter wait time), but the effects of such a program have not been evaluated. We evaluated the effects of such a program in a community with two ED sites.</p>
<p>METHODS: Descriptive statistics for wait times of the two sites before and after the publication of wait time information were used to evaluate the effects of the publication of wait time information on wait times. Multivariate logistical regression was used to test whether or not individual patients used published wait time to decide which site to visit.</p>
<p>RESULTS: We found that the rates of wait times exceeding 4 h, and the 95th percentile of wait times in the two sites decreased after the publication of wait time information, even though the average wait times experienced a slight increase. We also found that after controlling for other factors, the site with shorter wait time had a higher likelihood of being selected after the publication of wait time information, but there was no such relationship before the publication.</p>
<p>CONCLUSIONS: These findings were consistent with the hypothesis that the publication of wait time information leads to patients selecting the site with shorter wait time. While publishing ED wait time information did not improve average wait time, it reduced the rates of lengthy wait times.</p>
https://ir.lib.uwo.ca/epidempub/52
oai:ir.lib.uwo.ca:oncpub-1078
2011-08-31T23:46:25Z
publication:anatomy
publication:obsgyn
publication:oncpub
publication:obsgynpub
publication:pmid
publication:faculties
publication:anatomypub
publication:onc
21699706
On the Path to Translation: Highlights from the 2010 Canadian Conference on Ovarian Cancer Research
Thériault, Brigitte L.
Shepherd, Trevor G.
Review
2011-06-23T07:00:00Z
Knowledge translation
Ovarian Cancer
Journal of Ovarian Research
Journal of Ovarian Research
4
10
http://dx.doi.org/10.1186/1757-2215-4-10
Anatomy
Obstetrics and Gynecology
Oncology
<p>Ovarian cancer continues to be the most lethal of the gynaecologic malignancies due to the lack of early detection, screening strategies and ineffective therapeutics for late-stage metastatic disease, particularly in the recurrent setting. The gathering of researchers investigating fundamental pathobiology of ovarian cancer and the clinicians who treat patients with this insidious disease is paramount to meeting the challenges we face. Since 2002, the Canadian Conference on Ovarian Cancer Research, held every two years, has served this essential purpose. The objectives of this conference have been to disseminate new information arising from the most recent ovarian cancer research and identify the most pressing challenges we still face as scientists and clinicians. This is best accomplished through direct encounters and exchanges of innovative ideas among colleagues and trainees from the realms of basic science and clinical disciplines. This meeting has and continues to successfully facilitate rapid networking and establish new collaborations from across Canada. This year, more guest speakers and participants from other countries have extended the breadth of the research on ovarian cancer that was discussed at the meeting. This report summarizes the key findings presented at the fifth biennial Canadian Conference on Ovarian Cancer Research held in Toronto, Ontario, and includes the important issues and challenges we still face in the years ahead to make a significant impact on this devastating disease.</p>
https://ir.lib.uwo.ca/oncpub/78
oai:ir.lib.uwo.ca:physpharmpub-1064
2011-09-06T01:02:39Z
publication:anatomy
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:anatomypub
publication:physpharm
21738726
GPR54 (KISS1R) Transactivates EGFR to Promote Breast Cancer Cell Invasiveness
Zajac, Mateusz
Law, Jeffrey
Cvetkovic, Dragana Donna
Pampillo, Macarena
McColl, Lindsay
Pape, Cynthia
Di Guglielmo, Gianni M.
Postovit, Lynne M.
Babwah, Andy V.
Bhattacharya, Moshmi
Article
2011-06-28T07:00:00Z
Developmental Biology
Molecular Biology
Physiology
Diabetes and Endocrinology
Oncology
PLoS ONE
PLoS ONE
6
6
21599
21599
http://dx.doi.org/10.1371/journal.pone.0021599
Cell and Developmental Biology
Endocrinology, Diabetes, and Metabolism
Medical Physiology
Oncology
<p>Kisspeptins (Kp), peptide products of the Kisspeptin-1 (KISS1) gene are endogenous ligands for a G protein-coupled receptor 54 (GPR54). Previous findings have shown that KISS1 acts as a metastasis suppressor in numerous cancers in humans. However, recent studies have demonstrated that an increase in KISS1 and GPR54 expression in human breast tumors correlates with higher tumor grade and metastatic potential. At present, whether or not Kp signaling promotes breast cancer cell invasiveness, required for metastasis and the underlying mechanisms, is unknown. We have found that kisspeptin-10 (Kp-10), the most potent Kp, stimulates the invasion of human breast cancer MDA-MB-231 and Hs578T cells using Matrigel-coated Transwell chamber assays and induces the formation of invasive stellate structures in three-dimensional invasion assays. Furthermore, Kp-10 stimulated an increase in matrix metalloprotease (MMP)-9 activity. We also found that Kp-10 induced the transactivation of epidermal growth factor receptor (EGFR). Knockdown of the GPCR scaffolding protein, β-arrestin 2, inhibited Kp-10-induced EGFR transactivation as well as Kp-10 induced invasion of breast cancer cells via modulation of MMP-9 secretion and activity. Finally, we found that the two receptors associate with each other under basal conditions, and FRET analysis revealed that GPR54 interacts directly with EGFR. The stability of the receptor complex formation was increased upon treatment of cells by Kp-10. Taken together, our findings suggest a novel mechanism by which Kp signaling via GPR54 stimulates breast cancer cell invasiveness.</p>
https://ir.lib.uwo.ca/physpharmpub/62
oai:ir.lib.uwo.ca:obsgynpub-1013
2011-09-06T02:06:30Z
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:medpub
publication:med
publication:physpharm
20046880
SNAI1 and SNAI2 Are Asymmetrically Expressed at the 2-Cell Stage and Become Segregated to the TE in the Mouse Blastocyst
Bell, Christine E.
Watson, Andrew J.
Article
2009-12-31T08:00:00Z
Animals
Blastocyst
Blastomeres
Ectoderm
Embryonic Development
Fluorescent Antibody Technique
Gene Expression Regulation
Developmental
Immune Sera
Mice
NIH 3T3 Cells
Protein Transport
Reproducibility of Results
Reverse Transcriptase Polymerase Chain Reaction
Transcription Factors
Zygote
PLoS ONE
PLoS ONE
4
12
8530
8530
http://dx.doi.org/10.1371/journal.pone.0008530
Cell and Developmental Biology
Medical Physiology
Obstetrics and Gynecology
<p>SNAI1 and SNAI2 are transcription factors that initiate Epithelial-to-Mesenchymal cell transitions throughout development and in cancer metastasis. Here we show novel expression of SNAI1 and SNAI2 throughout mouse preimplantation development revealing asymmetrical localization of both SNAI1 and SNAI2 in individual blastomeres beginning at the 2-cell stage through to the 8-cell stage where SNAI1 and SNAI2 are then only detected in outer cells and not inner cells of the blastocyst. This study implicates SNAI1 and SNAI2 in the lineage segregation of the trophectoderm and inner cell mass, and provides new insight into these oncogenes.</p>
https://ir.lib.uwo.ca/obsgynpub/14
oai:ir.lib.uwo.ca:physpharmpub-1069
2011-11-14T00:19:36Z
publication:physpharmpub
publication:obsgyn
publication:obsgynpub
publication:paed
publication:pmid
publication:faculties
publication:physpharm
publication:paedpub
20107203
WNT2 Regulates DNA Synthesis in Mouse Granulosa Cells through Beta-catenin
Wang, Hong-Xing
Li, Tony Y.
Kidder, Gerald M.
Article
2010-05-01T07:00:00Z
Animals
Cell Proliferation
DNA
Female
Gene Knockdown Techniques
Glycogen Synthase Kinase 3
Granulosa Cells
Mice
Ovarian Follicle
Second Messenger Systems
Signal Transduction
Wnt2 Protein
beta Catenin
Biology of Reproduction
Biology of Reproduction
82
5
865
875
http://dx.doi.org/10.1095/biolreprod.109.080903
Medical Physiology
Obstetrics and Gynecology
Pediatrics
<p>WNTs are secreted extracellular signaling molecules that transduce their signals by binding to G protein-coupled receptors of the frizzled (FZD) family. They control diverse developmental processes, such as cell fate specification, cell proliferation, cell differentiation, and apoptosis. Although WNT signaling has been shown to be essential for development of the ovary, its mechanistic role in folliculogenesis within the adult ovary has not been studied extensively. Therefore, the objective of this study was to investigate the regulation and function of WNT2 signaling in mouse granulosa cells. Immunostaining identified WNT2 as being expressed in granulosa cells throughout folliculogenesis, but with varying signal strength: in sequential sections, WNT2 immunoreactivity was strongest in healthy antral follicles but weak in atretic follicles. Knockdown of WNT2 expression using transfected short interfering RNA decreased DNA synthesis in granulosa cells, whereas WNT2 overexpression using a recombinant viral vector enhanced it. WNT2 knockdown led to accumulation of glycogen synthase kinase-3beta (GSK3B) in the cytoplasm but reduced the expression of beta-catenin. Conversely, WNT2 overexpression reduced the expression of GSK3B in the cytoplasm and induced beta-catenin translocation from the membrane into the nucleus. Beta-catenin knockdown also inhibited DNA synthesis in granulosa cells and neutralized the effect of WNT2 overexpression. WNT2/beta-catenin signaling had a slight effect on the apoptosis of granulosa cells. Taken together, the data indicate that WNT2 regulates beta-catenin localization in granulosa cells, and WNT2/beta-catenin signaling contributes to regulating their proliferation.</p>
https://ir.lib.uwo.ca/physpharmpub/66
oai:ir.lib.uwo.ca:biologypub-1021
2012-02-24T02:12:22Z
publication:obsgyn
publication:obsgynpub
publication:paed
publication:pmid
publication:faculties
publication:biologypub
publication:biochempub
publication:biology
publication:biochem
publication:paedpub
22185339
Retinoic Acid Is a Key Regulatory Switch Determining the Difference between Lung and Thyroid Fates in Xenopus laevis
Wang, Jean H.
Deimling, Steven J.
D'Alessandro, Nicole E.
Zhao, Lin
Possmayer, Fred
Drysdale, Thomas A.
Article
2011-12-20T08:00:00Z
Retinoic acid
Lung
Thyroid
Xenopus laevis
BMC Developmental Biology
11
75
http://dx.doi.org/10.1186/1471-213X-11-75
Biochemistry, Biophysics, and Structural Biology
Biology
Obstetrics and Gynecology
Pediatrics
<p>BACKGROUND: The lung and thyroid are derived from the anterior endoderm. Retinoic acid and Fgf signalling are known to be essential for development of the lung in mouse but little is known on how the lung and thyroid are specified in Xenopus.</p>
<p>RESULTS: If either retinoic acid or Fgf signalling is inhibited, there is no differentiation of the lung as assayed by expression of sftpb. There is no change in expression of thyroid gland markers when retinoic acid signalling is blocked after gastrulation and when Fgf signalling is inhibited there is a short window of time where pax2 expression is inhibited but expression of other markers is unaffected. If exogenous retinoic acid is given to the embryo between embryonic stages 20 and 26, the presumptive thyroid expresses sftpb and sftpc, specific markers of lung differentiation and expression of key thyroid transcription factors is lost. When the presumptive thyroid is transplanted into the posterior embryo, it also expresses sftpb, although pax2 expression is not blocked.</p>
<p>CONCLUSIONS: After gastrulation, retinoic acid is required for lung but not thyroid differentiation in Xenopus while Fgf signalling is needed for lung but only for early expression of pax2 in the thyroid. Exposure to retinoic acid can cause the presumptive thyroid to switch to a lung developmental program.</p>
https://ir.lib.uwo.ca/biologypub/22
oai:ir.lib.uwo.ca:obsgynpub-1018
2016-07-14T15:29:31Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
1338415
How to make a blastocyst.
Watson, A J
Kidder, G M
Schultz, G A
Article
1992-10-01T07:00:00Z
Animals
Animals
Domestic
Blastocyst
DNA
Recombinant
Embryonic and Fetal Development
Female
Humans
Male
Mice
Reproductive Techniques
Sodium-Potassium-Exchanging ATPase
Biochemistry and cell biology = Biochimie et biologie cellulaire
70
10-11
849
855
Obstetrics and Gynecology
<p>Several of the new reproductive technologies have been cultivated from our current understanding of the genetic programming and cellular processes that are involved in the major morphogenetic events of mammalian preimplantation development. Research directed at characterizing the patterns of gene expression during early development has shown that the embryo is initially under maternal control and later superseded by new transcriptional activity provided by the activation of the embryonic genome. Several embryonic transcripts encoding: (i) growth factors, (ii) cell junctions, (iii) plasma membrane ion transporters, and (iv) cell adhesion molecules have been identified as contributing directly to the progression of the embryo through the preimplantation interval of development. In this brief review, we have outlined the patterns of expression and the integral roles that these gene families play in the morphogenetic events of compaction and cavitation. Research of this type has greatly facilitate our understanding of the control processes that underlie preimplantation development and represent but one area of this exciting and vigorous field of research.</p>
https://ir.lib.uwo.ca/obsgynpub/19
oai:ir.lib.uwo.ca:obsgynpub-1014
2016-07-14T13:26:24Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
2830159
Immunofluorescence assessment of the timing of appearance and cellular distribution of Na/K-ATPase during mouse embryogenesis.
Watson, A J
Kidder, G M
Article
1988-03-01T08:00:00Z
Animals
Blastomeres
Embryo
Mammalian
Fixatives
Fluorescent Antibody Technique
Mice
Morula
Polyethylene Glycols
Sodium-Potassium-Exchanging ATPase
Wheat Germ Agglutinins
Developmental biology
126
1
80
90
http://dx.doi.org/10.1016/0012-1606(88)90241-2
Obstetrics and Gynecology
<p>We have employed immunofluorescence with a rat kidney Na+/K+-ATPase polyclonal antibody to investigate the cellular distribution and timing of appearance of this enzyme during preimplantation development. The enzyme is first detected in the late morula within the cytoplasm of each blastomere. When cavitation begins this distribution changes dramatically to a ring encircling the blastocoel, restricted to the basolateral cell margins. Using this enzyme as a marker for cavitation, we examined its expression in embryos that had been treated with wheat germ agglutinin (WGA), which causes cleavage arrest and was reported to trigger premature compaction- and cavitation-like events in early cleavage stages (L. V. Johnson, 1986, Dev. Biol. 113, 1-9). Although WGA-treated 2-,4-, and 8-cell embryos quickly underwent compaction- and cavitation-like events, no Na+/K+-ATPase expression was observed. Thus the WGA effect does not likely involve acceleration of the developmental program for cavitation. Embryos arrested at the 8-cell stage but cultured overnight to Day 4, however, expressed the enzyme in the typical blastocyst pattern (around each fluid-filled cavity). We conclude that Na+/K+-ATPase expression is initiated or increases dramatically in the late morula and is independent of cytokinesis. The enzyme assumes a distribution during cavitation consistent with its presumed role in transtrophectodermal fluid transport.</p>
https://ir.lib.uwo.ca/obsgynpub/15
oai:ir.lib.uwo.ca:obsgynpub-1015
2016-07-14T13:37:07Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
2163294
Expression of Na,K-ATPase alpha and beta subunit genes during preimplantation development of the mouse.
Watson, A J
Pape, C
Emanuel, J R
Levenson, R
Kidder, G M
Article
1990-01-01T08:00:00Z
Animals
Blastocyst
Blotting
Northern
Densitometry
Female
Gene Expression Regulation
Enzymologic
Histones
Male
Mice
RNA
Messenger
Sodium-Potassium-Exchanging ATPase
Developmental genetics
11
1
41
48
http://onlinelibrary.wiley.com/doi/10.1002/dvg.1020110106/abstract
Obstetrics and Gynecology
<p>Na,K-ATPase is a plasma membrane enzyme that plays a critical role in eutherian blastocoel formation (cavitation) by pumping Na+ into the extracellular space enclosed by the trophectoderm. Previous experiments with the mouse had shown that the alpha (catalytic) subunit of the enzyme becomes detectable by immunocytochemistry in the late morula, just prior to the onset of cavitation. In the present study we have used cDNAs corresponding to three mRNA isoforms of the alpha subunit and a beta subunit to determine which genes are expressed during preimplantation development and to explore the timing of their expression. Of the three alpha subunit cDNAs tested by Northern blot hybridization with blastocyst RNA, only alpha 1 produced a hybridization signal, recognizing a single mRNA about 4 kb in length. This mRNA is relatively abundant in zygotes but barely detectable by the 2-cell stage and then accumulates steadily thereafter to reach its preimplantation maximum in blastocysts. The beta 1 cDNA detected mRNA of about 2.6-2.8 kb. This mRNA is present in zygotes but could not be detected in 2-, 4-, or 8-cell stages; it is present at a low level in late morulae and is abundant in blastocysts. The temporal profile of accumulation of beta 1 mRNA thus matches more closely than does alpha 1 the timing of appearance of the catalytic subunit. This suggests that the beta subunit may regulate production of the holoenzyme and hence the timing of cavitation.</p>
https://ir.lib.uwo.ca/obsgynpub/17
oai:ir.lib.uwo.ca:obsgynpub-1016
2016-07-14T13:41:22Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
2188651
Cell polarity and development of the first epithelium.
Wiley, L M
Kidder, G M
Watson, A J
Article
1990-02-01T08:00:00Z
Animals
Blastomeres
Cell Differentiation
Cell Membrane
Epithelial Cells
Epithelium
BioEssays : news and reviews in molecular, cellular and developmental biology
12
2
67
73
http://onlinelibrary.wiley.com/doi/10.1002/bies.950120204/abstract
Obstetrics and Gynecology
<p>In the 4 1/2 to 5 days between fertilization and implantation, the mouse conceptus must gain the abilities to implant and produce an embryo. Each of these is the sole developmental responsibility of one of two cell types forming the blastocyst, trophectoderm and inner cell mass (ICM), respectively. Trophectoderm is a polarized transporting epithelium while the ICM is an aggregate of non-epithelial pluripotent stem cells. These two cell types originate from the division of polar blastomeres when their cleavage furrows parallel their apical surfaces. Blastomeres polarize in response to asymmetric cell--cell contact, and understanding the mechanism of this induction is regarded as the key to understanding the origin of trophectoderm and ICM. Here we propose a model based on transcellular ion current loops for the induction of cell polarity during the development of the first epithelium, trophectoderm.</p>
https://ir.lib.uwo.ca/obsgynpub/18
oai:ir.lib.uwo.ca:obsgynpub-1017
2016-07-14T13:29:32Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
2167855
Differentiation of an epithelium: factors affecting the polarized distribution of Na+,K(+)-ATPase in mouse trophectoderm.
Watson, A J
Damsky, C H
Kidder, G M
Article
1990-09-01T07:00:00Z
Animals
Biomarkers
Blastocyst
Blastomeres
Cadherins
Cell Adhesion
Cell Differentiation
Cytochalasin B
Epithelial Cells
Fluorescent Antibody Technique
In Vitro Techniques
Mice
Microscopy
Electron
Sodium-Potassium-Exchanging ATPase
Up-Regulation
Developmental biology
141
1
104
114
http://dx.doi.org/10.1016/0012-1606(90)90105-R
Obstetrics and Gynecology
<p>Na+,K(+)-ATPase is a marker of the basolateral plasma membrane domain of polarized epithelial cells, including the mural trophectoderm of the mammalian blastocyst (Watson and Kidder (1988). Dev. Biol. 126, 80-90). We have used this marker to explore the factors governing the establishment and maintenance of apical/basolateral polarity during differentiation of trophectoderm. A polyclonal antiserum (anti-GP80) against human cell-CAM 120/80, a homolog of the mouse cell-cell adhesion protein, uvomorulin, was used to prevent cell flattening (compaction) and formation of the epithelial junctional complex. The majority of treated embryos failed to develop a blastocoel; instead their blastomeres developed fluid-filled cavities that expanded while untreated control embryos were cavitating. Immunocytochemistry revealed that the catalytic subunit of Na+,K(+)-ATPase was contained within the membranes lining these cavities, as well as within numerous punctate foci in the cytoplasm. The down-regulation of expression of the enzyme that normally occurs in the ICM and polar trophectoderm did not take place, since the immunoreactivity remained equally strong in all blastomeres. The enzyme could not be detected in plasma membranes. We conclude that uvomorulin-mediated cell adhesion is involved in spatially restricting the expression of the catalytic subunit and is a prerequisite for the insertion of enzyme-laden vesicles into plasma membranes, but not for expression of the catalytic subunit gene. When fully developed blastocysts were treated with cytochalasins to disrupt the epithelial junctional complex, the catalytic subunit shifted from the basolateral to the apical plasma membrane. This finding suggests a primary role for the apical plasma membrane in the process of polarization, and implies that tight junctions are a manifestation of polarity that serve to maintain the separation between apical and basolateral markers.</p>
https://ir.lib.uwo.ca/obsgynpub/16
oai:ir.lib.uwo.ca:obsgynpub-1024
2016-08-29T18:01:22Z
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:rwkex
23729591
Stress-inducible phosphoprotein 1 has unique cochaperone activity during development and regulates cellular response to ischemia via the prion protein.
Beraldo, Flavio H
Soares, Iaci N
Goncalves, Daniela F
Fan, Jue
Thomas, Anu A
Santos, Tiago G
Mohammad, Amro H
Roffé, Martin
Calder, Michele D
Nikolova, Simona
Hajj, Glaucia N
Guimaraes, Andre L
Massensini, Andre R
Welch, Ian
Betts, Dean H
Gros, Robert
Drangova, Maria
Watson, Andrew J
Bartha, Robert
Prado, Vania F
Martins, Vilma R
Prado, Marco A M
Article
2013-09-01T07:00:00Z
Animals
Blastocyst
Blotting
Western
Cells
Cultured
Embryo
Mammalian
Female
Heat-Shock Proteins
Homeodomain Proteins
In Vitro Techniques
Ischemia
Mice
Mice
Mutant Strains
Molecular Chaperones
Octamer Transcription Factor-3
Pregnancy
Prions
Transcription Factors
The FASEB journal : official publication of the Federation of American Societies for Experimental Biology
27
9
3594
3607
Obstetrics and Gynecology
<p>Stress-inducible phosphoprotein 1 (STI1) is part of the chaperone machinery, but it also functions as an extracellular ligand for the prion protein. However, the physiological relevance of these STI1 activities in vivo is unknown. Here, we show that in the absence of embryonic STI1, several Hsp90 client proteins are decreased by 50%, although Hsp90 levels are unaffected. Mutant STI1 mice showed increased caspase-3 activation and 50% impairment in cellular proliferation. Moreover, placental disruption and lack of cellular viability were linked to embryonic death by E10.5 in STI1-mutant mice. Rescue of embryonic lethality in these mutants, by transgenic expression of the STI1 gene, supported a unique role for STI1 during embryonic development. The response of STI1 haploinsufficient mice to cellular stress seemed compromised, and mutant mice showed increased vulnerability to ischemic insult. At the cellular level, ischemia increased the secretion of STI1 from wild-type astrocytes by 3-fold, whereas STI1 haploinsufficient mice secreted half as much STI1. Interesting, extracellular STI1 prevented ischemia-mediated neuronal death in a prion protein-dependent way. Our study reveals essential roles for intracellular and extracellular STI1 in cellular resilience.</p>
https://ir.lib.uwo.ca/obsgynpub/22
oai:ir.lib.uwo.ca:obsgynpub-1021
2016-08-29T17:57:01Z
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:rwkex
26081282
Simulated diabetic ketoacidosis therapy in vitro elicits brain cell swelling via sodium-hydrogen exchange and anion transport.
Rose, Keeley L
Watson, Andrew J
Drysdale, Thomas A
Cepinskas, Gediminas
Chan, Melissa
Rupar, C Anthony
Fraser, Douglas D
Article
2015-08-15T07:00:00Z
Alloxan
Animals
Anions
Brain
Brain Edema
Diabetes Mellitus
Experimental
Diabetes Mellitus
Type 1
Diabetic Ketoacidosis
Fluid Therapy
Insulin
Ion Transport
Mice
Organ Culture Techniques
Osmolar Concentration
Sodium-Hydrogen Antiporter
Streptozocin
American journal of physiology. Endocrinology and metabolism
309
4
370
379
Obstetrics and Gynecology
<p>A common complication of type 1 diabetes mellitus is diabetic ketoacidosis (DKA), a state of severe insulin deficiency. A potentially harmful consequence of DKA therapy in children is cerebral edema (DKA-CE); however, the mechanisms of therapy-induced DKA-CE are unknown. Our aims were to identify the DKA treatment factors and membrane mechanisms that might contribute specifically to brain cell swelling. To this end, DKA was induced in juvenile mice with the administration of the pancreatic toxins streptozocin and alloxan. Brain slices were prepared and exposed to DKA-like conditions in vitro. Cell volume changes were imaged in response to simulated DKA therapy. Our experiments showed that cell swelling was elicited with isolated DKA treatment components, including alkalinization, insulin/alkalinization, and rapid reductions in osmolality. Methyl-isobutyl-amiloride, a nonselective inhibitor of sodium-hydrogen exchangers (NHEs), reduced cell swelling in brain slices elicited with simulated DKA therapy (in vitro) and decreased brain water content in juvenile DKA mice administered insulin and rehydration therapy (in vivo). Specific pharmacological inhibition of the NHE1 isoform with cariporide also inhibited cell swelling, but only in the presence of the anion transport (AT) inhibitor 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid. DKA did not alter brain NHE1 isoform expression, suggesting that the cell swelling attributed to the NHE1 was activity dependent. In conclusion, our data raise the possibility that brain cell swelling can be elicited by DKA treatment factors and that it is mediated by NHEs and/or coactivation of NHE1 and AT.</p>
https://ir.lib.uwo.ca/obsgynpub/25
oai:ir.lib.uwo.ca:obsgynpub-1019
2016-08-31T13:54:57Z
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:rwkex
27430916
Effects of American Ginseng on Preimplantation Development and Pregnancy in Mice.
Belanger, Danyka
Calder, Michele D
Gianetto-Berruti, Alessandra
Lui, Edmund M
Watson, Andrew J
Feyles, Valter
Article
2016-07-19T07:00:00Z
2017-08-01T07:00:00Z
The American journal of Chinese medicine
44
5
981
995
Obstetrics and Gynecology
<p>In North America, a high proportion of pregnant women use herbal medications including North American ginseng. This medicinal plant contains high amounts of triterpene saponins (ginsenosides), which are the main bioactive compounds. It is important to assess ginseng's impact on all reproductive functions to ensure the safety of pregnant women and fetuses. In this study, we defined the concentration-responsive effects of North American alcoholic and aqueous ginseng extracts on preimplantation development in vitro and on pregnancy and post-partum development in the mouse. Two-cell mouse embryos were cultured with 5 different concentrations of whole ginseng root extracts, or ginsenosides Rb1, Rg1 and Re alone, a combinatorial ginsenoside solution and a crude polysaccharide fraction solution. Embryonic development and recovery from each treatment was assessed. To investigate the in vivo effects of ginseng extracts, female mice were gavaged with 50[Formula: see text]mg/kg/day, 500[Formula: see text]mg/kg/day or 2000[Formula: see text]mg/kg/day of either extract (treatment) or water (sham) for 2 weeks prior to mating and throughout gestation. Gestation period, litter size, pup growth and pup sex ratio were evaluated. Oral ginseng consumption did not significantly affect fertility or pregnancy in the mouse. High doses of ginseng (2000[Formula: see text]mg/kg/day) decreased maternal weight gain. Direct treatment of preimplantation embryos in vitro demonstrated that ALC and AQ extract treatment reduced development in a concentration responsive manner, while only ALC extract effects were largely reversible. Treatments with individual or combinatorial ginsenosides, or the polysaccharide fraction solution alone did not impair preimplantation development, in vitro. In conclusion, maternal oral consumption of ginseng has little negative impact on pregnancy in the mouse, however, direct exposure to ginseng extract during mouse preimplantation development in vitro is detrimental.</p>
https://ir.lib.uwo.ca/obsgynpub/36
oai:ir.lib.uwo.ca:obsgynpub-1020
2016-08-29T17:53:57Z
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:rwkex
27385725
P66Shc, a key regulator of metabolism and mitochondrial ROS production, is dysregulated by mouse embryo culture.
Edwards, Nicole A
Watson, Andrew J
Betts, Dean H
Article
2016-07-06T07:00:00Z
Molecular human reproduction
Obstetrics and Gynecology
<p>STUDY QUESTION: Do high oxygen tension and high glucose concentrations dysregulate p66Shc (Src homologous-collagen homologue adaptor protein) expression during mouse preimplantation embryo culture?</p>
<p>SUMMARY ANSWER: Compared with mouse blastocysts in vivo, P66Shc mRNA and protein levels in blastocysts maintained in vitro increased under high oxygen tension (21%), but not high glucose concentration.</p>
<p>WHAT IS KNOWN ALREADY: Growth in culture adversely impacts preimplantation embryo development and alters the expression levels of the oxidative stress adaptor protein p66Shc, but it is not known if p66Shc expression is linked to metabolic changes observed in cultured embryos.</p>
<p>STUDY DESIGN, SAMPLES/MATERIALS, METHODS: We used a standard wild-type CD1 mouse model of preimplantation embryo development and embryo culture with different atmospheric oxygen tension and glucose media concentrations. Changes to p66Shc expression in mouse blastocysts were measured using quantitative RT-PCR, immunoblotting and immunofluorescence followed by confocal microscopy. Changes to oxidative phosphorylation metabolism were measured by total ATP content and superoxide production. Statistical analyses were performed on a minimum of three experimental replicates using Students' t-test or one-way ANOVA.</p>
<p>MAIN RESULTS AND THE ROLE OF CHANCE: P66Shc is basally expressed during in vivo mouse preimplantation development. Within in vivo blastocysts, p66Shc is primarily localized to the cell periphery of the trophectoderm. Blastocysts cultured under atmospheric oxygen levels have significantly increased p66Shc mRNA transcript and protein abundances compared to in vivo controls (P < 0.05). However, the ratio of phosphorylated serine 36 (S36) p66Shc to total p66Shc decreased in culture regardless of O2 atmosphere used, supporting a shift in the mitochondrial fraction of p66Shc. Total p66Shc localized to the cell periphery of the blastocyst trophectoderm and phosphorylated S36 p66Shc displayed nuclear and cytoplasmic immunoreactivity, suggesting distinct compartmentalization of phosphorylated S36 p66Shc and the remaining p66Shc fraction. Glucose concentration in the culture medium did not significantly change p66Shc mRNA or protein abundance or its localization. Blastocysts cultured under low or high oxygen conditions exhibited significantly decreased cellular ATP and increased superoxide production compared to in vivo derived embryos (P < 0.05).</p>
<p>LIMITATIONS/REASONS FOR CAUTION: This study associates embryonic p66Shc expression levels with metabolic abnormalities but does not directly implicate p66Shc in metabolic changes. Additionally, we used one formulation of embryo culture medium that differs from that used in other mouse model studies and from clinical media used to support human blastocyst development. Our findings may, therefore, be limited to this media, or may be a species-specific phenomenon.</p>
<p>WIDER IMPLICATIONS OF THE FINDINGS: This is the first study to show distinct immunolocalization of p66Shc to the trophectoderm of mouse blastocysts and that its levels are abnormally increased in embryos exposed to culture conditions. Changes in p66Shc expression and/or localization could possibly serve as a molecular marker of embryo viability for clinical applications. The outcomes provide insight into the potential metabolic role of p66Shc. Metabolic anomalies are induced even under the current optimal culture conditions, which could negatively impact trophectoderm and placental development.</p>
<p>LARGE SCALE DATA: Not applicable.</p>
<p>STUDY FUNDING AND COMPETING INTERESTS: Canadian Institutes of Health Research (CIHR) operating funds, Ontario Graduate Scholarship (OGS). There are no competing interests.</p>
https://ir.lib.uwo.ca/obsgynpub/59
oai:ir.lib.uwo.ca:obsgynpub-1022
2016-08-29T17:58:44Z
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:rwkex
24877624
Implantation failure in female Kiss1-/- mice is independent of their hypogonadic state and can be partially rescued by leukemia inhibitory factor.
Calder, Michele
Chan, Yee-Ming
Raj, Renju
Pampillo, Macarena
Elbert, Adrienne
Noonan, Michelle
Gillio-Meina, Carolina
Caligioni, Claudia
Bérubé, Nathalie G
Bhattacharya, Moshmi
Watson, Andrew J
Seminara, Stephanie B
Babwah, Andy V
Article
2014-08-01T07:00:00Z
Animals
Embryo Implantation
Estradiol
Female
Gonadotropins
Kisspeptins
Leukemia Inhibitory Factor
Male
Mice
Mice
Knockout
Mice
Transgenic
Pregnancy
Pregnancy
Animal
Progesterone
Superovulation
Uterus
Endocrinology
155
8
3065
3078
Obstetrics and Gynecology
<p>The hypothalamic kisspeptin signaling system is a major positive regulator of the reproductive neuroendocrine axis, and loss of Kiss1 in the mouse results in infertility, a condition generally attributed to its hypogonadotropic hypogonadism. We demonstrate that in Kiss1(-/-) female mice, acute replacement of gonadotropins and estradiol restores ovulation, mating, and fertilization; however, these mice are still unable to achieve pregnancy because embryos fail to implant. Progesterone treatment did not overcome this defect. Kiss1(+/-) embryos transferred to a wild-type female mouse can successfully implant, demonstrating the defect is due to maternal factors. Kisspeptin and its receptor are expressed in the mouse uterus, and we suggest that it is the absence of uterine kisspeptin signaling that underlies the implantation failure. This absence, however, does not prevent the closure of the uterine implantation chamber, proper alignment of the embryo, and the ability of the uterus to undergo decidualization. Instead, the loss of Kiss1 expression specifically disrupts embryo attachment to the uterus. We observed that on the day of implantation, leukemia inhibitory factor (Lif), a cytokine that is absolutely required for implantation in mice, is weakly expressed in Kiss1(-/-) uterine glands and that the administration of exogenous Lif to hormone-primed Kiss1(-/-) female mice is sufficient to partially rescue implantation. Taken together, our study reveals that uterine kisspeptin signaling regulates glandular Lif levels, thereby identifying a novel and critical role for kisspeptin in regulating embryo implantation in the mouse. This study provides compelling reasons to explore this role in other species, particularly livestock and humans.</p>
https://ir.lib.uwo.ca/obsgynpub/24
oai:ir.lib.uwo.ca:obsgynpub-1023
2016-08-29T18:00:14Z
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:rwkex
23946536
Endogenous folate accumulation in oocytes and preimplantation embryos and its epigenetic implications.
Mann, Mellissa R W
Watson, Andrew J
Article
2013-09-01T07:00:00Z
Animals
Blastocyst
Cumulus Cells
Female
Folate Receptor 1
Folate Receptor 2
Folic Acid
Oocytes
Reduced Folate Carrier Protein
Biology of reproduction
89
3
62
62
Obstetrics and Gynecology
https://ir.lib.uwo.ca/obsgynpub/23
oai:ir.lib.uwo.ca:obsgynpub-1028
2016-08-29T18:07:27Z
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:rwkex
21901128
Ouabain stimulates a Na+/K+-ATPase-mediated SFK-activated signalling pathway that regulates tight junction function in the mouse blastocyst.
Giannatselis, Holly
Calder, Michele
Watson, Andrew J
Article
2011-01-01T08:00:00Z
Animals
Blastocyst
Blotting
Western
Cells
Cultured
Embryo
Mammalian
Female
Fluorescent Antibody Technique
Indirect
Indoles
Male
Mice
Microscopy
Confocal
Ouabain
Phosphorylation
Pregnancy
Reverse Transcriptase Polymerase Chain Reaction
Signal Transduction
Sodium-Potassium-Exchanging ATPase
Sulfonamides
Tight Junctions
src-Family Kinases
PLoS One
6
8
23704
23704
Obstetrics and Gynecology
<p>The Na(+)/K(+)-ATPase plays a pivotal role during preimplantation development; it establishes a trans-epithelial ionic gradient that facilitates the formation of the fluid-filled blastocyst cavity, crucial for implantation and successful pregnancy. The Na(+)/K(+)-ATPase is also implicated in regulating tight junctions and cardiotonic steroid (CTS)-induced signal transduction via SRC. We investigated the expression of SRC family kinase (SFK) members, Src and Yes, during preimplantation development and determined whether SFK activity is required for blastocyst formation. Embryos were collected following super-ovulation of CD1 or MF1 female mice. RT-PCR was used to detect SFK mRNAs encoding Src and Yes throughout preimplantation development. SRC and YES protein were localized throughout preimplantation development. Treatment of mouse morulae with the SFK inhibitors PP2 and SU6656 for 18 hours resulted in a reversible blockade of progression to the blastocyst stage. Blastocysts treated with 10(-3) M ouabain for 2 or 10 minutes and immediately immunostained for phosphorylation at SRC tyr418 displayed reduced phosphorylation while in contrast blastocysts treated with 10(-4) M displayed increased tyr418 fluorescence. SFK inhibition increased and SFK activation reduced trophectoderm tight junction permeability in blastocysts. The results demonstrate that SFKs are expressed during preimplantation development and that SFK activity is required for blastocyst formation and is an important mediator of trophectoderm tight junction permeability.</p>
https://ir.lib.uwo.ca/obsgynpub/28
oai:ir.lib.uwo.ca:obsgynpub-1026
2016-08-29T18:04:55Z
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:rwkex
22190704
Outer space and oocyte developmental competence.
Watson, Andrew J
Article
2012-03-01T08:00:00Z
Animals
Apoptosis
Female
Oocytes
Rad51 Recombinase
Radiation
Ionizing
Biology of reproduction
86
3
75
75
Obstetrics and Gynecology
https://ir.lib.uwo.ca/obsgynpub/26
oai:ir.lib.uwo.ca:obsgynpub-1025
2016-08-29T18:02:30Z
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:rwkex
23593143
p38 MAPK regulates cavitation and tight junction function in the mouse blastocyst.
Bell, Christine E
Watson, Andrew J
Article
2013-01-01T08:00:00Z
Animals
Apoptosis
Aquaporin 3
Blastocyst
Female
Gene Expression Regulation
Developmental
Imidazoles
Mice
Permeability
Pregnancy
Protein Transport
Pyrimidines
RNA
Messenger
Signal Transduction
Tight Junctions
Zonula Occludens-1 Protein
p38 Mitogen-Activated Protein Kinases
PLoS One
8
4
59528
59528
Obstetrics and Gynecology
<p>UNLABELLED: Blastocyst formation is essential for implantation and maintenance of pregnancy and is dependent on the expression and coordinated function of a series of proteins involved in establishing and maintaining the trans-trophectoderm ion gradient that enables blastocyst expansion. These consist of Na/K-ATPase, adherens junctions, tight junctions (TJ) and aquaporins (AQP). While their role in supporting blastocyst formation is established, the intracellular signaling pathways that coordinate their function is unclear. The p38 MAPK pathway plays a role in regulating these proteins in other cell types and is required for embryo development at the 8-16 cell stage, but its role has not been investigated in the blastocyst.</p>
<p>HYPOTHESIS: p38 MAPK regulates blastocyst formation by regulating blastocyst formation gene expression and function.</p>
<p>METHODS: Embryos were cultured from the early blastocyst stage for 12 h or 24 h in the presence of a potent and specific p38 MAPK inhibitor, SB 220025. Blastocyst expansion, hatching, gene family expression and localization, TJ function and apoptosis levels were analyzed.</p>
<p>RESULTS: Inhibition of the p38 MAPK pathway reduced blastocyst expansion and hatching, increased tight junction permeability, affected TJP1 localization, reduced Aqp3 expression, and induced a significant increase in apoptosis.</p>
<p>CONCLUSION: The p38 MAPK pathway coordinates the overall events that regulate blastocyst formation.</p>
https://ir.lib.uwo.ca/obsgynpub/21
oai:ir.lib.uwo.ca:obsgynpub-1027
2016-08-29T18:06:08Z
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:rwkex
22155729
Embryo collection induces transient activation of XBP1 arm of the ER stress response while embryo vitrification does not.
Abraham, Tamara
Pin, Christopher L
Watson, Andrew J
Article
2012-05-01T07:00:00Z
Animals
Apoptosis
Cryopreservation
DNA-Binding Proteins
Embryo Culture Techniques
Embryo
Mammalian
Embryonic Development
Endoplasmic Reticulum
Endoplasmic Reticulum Stress
Female
In Situ Nick-End Labeling
Mice
Mice
Inbred Strains
Molecular Chaperones
RNA Splicing
RNA
Messenger
Reproductive Techniques
Assisted
Transcription Factors
Unfolded Protein Response
Molecular human reproduction
18
5
229
242
Obstetrics and Gynecology
<p>Embryo cryopreservation has become a standard procedure in the practice of assisted reproduction. While routinely performed in IVF labs, the effects of embryo vitrification on the molecular mechanisms governing preimplantation development remain largely unknown. The endoplasmic reticulum stress (ER stress) response is an evolutionary conserved mechanism that cells employ to manage ER stress. ER stress can be defined as an imbalance between protein synthesis and secretion within the ER. The primary focus of this study was to investigate whether standard embryo manipulations, including embryo collection, culture and vitrification, result in activation of the ER stress pathway in vitro and to determine whether the embryo utilizes the unfolded protein response as an adaptive response. Our results indicate that the major ER stress pathway constituents are present at all stages of preimplantation development and that the activation of ER stress pathways can be induced at the 8-cell, morula and blastocyst stages. Additionally, we have demonstrated that the IRE1α arm of the ER Stress pathway is activated in freshly collected embryos but contrastingly, this ER Stress arm is not activated following embryo vitrification. It is important to understand the possible stresses that Assisted Reproductive Technologies place on the embryo and the mechanisms the embryo employs to adapt to these stresses. This study indicates that among the adaptive pathways available, cultured mammalian embryos can employ the ER stress pathway. Assisted reproduction techniques should be aware that their activities may induce the ER stress pathway in their patients' early embryos.</p>
https://ir.lib.uwo.ca/obsgynpub/27
oai:ir.lib.uwo.ca:obsgynpub-1029
2016-08-29T18:08:35Z
publication:rwkex_researcharticles
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
publication:rwkex
21846809
Culture medium, gas atmosphere and MAPK inhibition affect regulation of RNA-binding protein targets during mouse preimplantation development.
Calder, Michele D
Watson, Patricia H
Watson, Andrew J
Article
2011-11-01T07:00:00Z
Animals
Atmospheric Pressure
Cells
Cultured
Culture Media
Embryo Culture Techniques
Embryo
Mammalian
Embryonic Development
Extracellular Signal-Regulated MAP Kinases
Female
Gases
Gene Expression Regulation
Male
Mice
Pregnancy
Protein Kinase Inhibitors
RNA-Binding Proteins
Reproduction (Cambridge, England)
142
5
689
698
Obstetrics and Gynecology
<p>During oogenesis, mammalian oocytes accumulate maternal mRNAs that support the embryo until embryonic genome activation. RNA-binding proteins (RBP) may regulate the stability and turnover of maternal and embryonic mRNAs. We hypothesised that varying embryo culture conditions, such as culture medium, oxygen tension and MAPK inhibition, affects regulation of RBPs and their targets during preimplantation development. STAU1, ELAVL1, KHSRP and ZFP36 proteins and mRNAs were detected throughout mouse preimplantation development, whereas Elavl2 mRNA decreased after the two-cell stage. Potential target mRNAs of RBP regulation, Gclc, Slc2a1 and Slc7a1 were detected during mouse preimplantation development. Gclc mRNA was significantly elevated in embryos cultured in Whitten's medium compared with embryos cultured in KSOMaa, and Gclc mRNA was elevated under high-oxygen conditions. Inhibition of the p38 MAPK pathway reduced Slc7a1 mRNA expression while inhibition of ERK increased Slc2a1 mRNA expression. The half-lives of the potential RBP mRNA targets are not regulated in parallel; Slc2a1 mRNA displayed the longest half-life. Our results indicate that mRNAs and proteins encoding five RBPs are present during preimplantation development and more importantly, demonstrate that expression of RBP target mRNAs are regulated by culture medium, gas atmosphere and MAPK pathways.</p>
https://ir.lib.uwo.ca/obsgynpub/29
oai:ir.lib.uwo.ca:obsgynpub-1030
2016-08-29T18:11:01Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
20046880
SNAI1 and SNAI2 are asymmetrically expressed at the 2-cell stage and become segregated to the TE in the mouse blastocyst.
Bell, Christine E
Watson, Andrew J
Article
2009-01-01T08:00:00Z
Animals
Blastocyst
Blastomeres
Ectoderm
Embryonic Development
Fluorescent Antibody Technique
Gene Expression Regulation
Developmental
Immune Sera
Mice
NIH 3T3 Cells
Protein Transport
Reproducibility of Results
Reverse Transcriptase Polymerase Chain Reaction
Transcription Factors
Zygote
PLoS One
4
12
8530
8530
Obstetrics and Gynecology
<p>SNAI1 and SNAI2 are transcription factors that initiate Epithelial-to-Mesenchymal cell transitions throughout development and in cancer metastasis. Here we show novel expression of SNAI1 and SNAI2 throughout mouse preimplantation development revealing asymmetrical localization of both SNAI1 and SNAI2 in individual blastomeres beginning at the 2-cell stage through to the 8-cell stage where SNAI1 and SNAI2 are then only detected in outer cells and not inner cells of the blastocyst. This study implicates SNAI1 and SNAI2 in the lineage segregation of the trophectoderm and inner cell mass, and provides new insight into these oncogenes.</p>
https://ir.lib.uwo.ca/obsgynpub/30
oai:ir.lib.uwo.ca:obsgynpub-1031
2016-08-29T18:11:56Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
19846464
Oocyte peptides as paracrine tools for ovarian stimulation and oocyte maturation.
Mottershead, David G
Watson, Andrew J
Article
2009-12-01T08:00:00Z
Animals
Bone Morphogenetic Protein 15
Cell Line
Female
Growth Differentiation Factor 9
Humans
Mice
Oocytes
Ovulation Induction
Paracrine Communication
Peptides
Smad Proteins
Molecular human reproduction
15
12
789
794
Obstetrics and Gynecology
<p>Recent studies report the production and isolation of a stable bioactive recombinant human bone morphogenetic protein 15 (rhBMP15) that is appropriately processed in HEK-293 cells and activates the SMAD 1/5/8 pathway in mouse granulosa cell cultures. Further, the purified rhBMP15 induces the expression of genes associated with cumulus expansion. Thanks to recent research, we have a greater understanding of the importance of the dialogue that occurs between the oocyte and the granulosa cell layer with regard to regulating folliculogenesis and the acquisition of oocyte developmental competence and maturation. BMP15 is one of the critical components of these intra-follicular communication pathways. The production of recombinant human BMP15 is important for understanding the biochemistry of this specific pathway and for also fully understanding its functional contributions to mediating oocyte development. The production of a stable recombinant human BMP15 is also important for use in experiments aimed at optimizing ovarian stimulation protocols and in vitro oocyte maturation methods. This is required to improve oocyte and embryonic developmental competence and increase our ability to effectively use in vitro methods for animal production and the treatment of human infertility.</p>
https://ir.lib.uwo.ca/obsgynpub/31
oai:ir.lib.uwo.ca:obsgynpub-1032
2016-08-29T18:13:10Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
19258345
Mitogen-activated protein kinase (MAPK) pathways mediate embryonic responses to culture medium osmolarity by regulating Aquaporin 3 and 9 expression and localization, as well as embryonic apoptosis.
Bell, Christine E
Larivière, Nathalie M K
Watson, Patricia H
Watson, Andrew J
Article
2009-06-01T07:00:00Z
Animals
Apoptosis
Aquaporin 3
Aquaporins
Blastocyst
Culture Media
Embryo Culture Techniques
Enzyme Inhibitors
Female
Glycerol
MAP Kinase Signaling System
Male
Mice
Mitogen-Activated Protein Kinase 11
Mitogen-Activated Protein Kinase 14
Mitogen-Activated Protein Kinase 8
Osmolar Concentration
Pregnancy
RNA
Messenger
Sucrose
Water-Electrolyte Balance
Human reproduction (Oxford, England)
24
6
1373
1386
Obstetrics and Gynecology
<p>BACKGROUND: In order to advance the development of culture conditions and increase the potential for supporting normal preimplantation embryo development in vitro, it is critical to define the mechanisms that early embryos utilize to survive in culture. We investigated the mechanisms that embryos employ in response to culture medium osmolarity. We hypothesized that mitogen-activated protein kinase (MAPK) pathways mediate responses to hyperosmotic stress by regulating Aquaporin (AQP) 3 and 9 expression as well as embryonic apoptosis.</p>
<p>METHODS: Real-time reverse transcription and polymerase chain reaction and whole-mount immunofluorescence were used to determine the relative mRNA levels and protein localization patterns of AQP 3 and 9 after hyperosmotic medium treatment.</p>
<p>RESULTS: At 6 and 24 h, a significant increase in Aqp 3 and 9 mRNA was observed in the sucrose hyperosmotic treatment compared with standard medium and glycerol controls. Blockade of MAPK14/11 negated the increase in Aqp 3 and 9 mRNA levels, whereas culture in a MAPK8 blocker did not. Hyperosmotic sucrose treatment significantly increased embryonic apoptosis which was negated in the presence of MAPK8 blocker, but not MAPK14/11 blocker.</p>
<p>CONCLUSIONS: MAPK14/11 activation is a component of the rapid adaptive stress response mechanism that includes the effects of AQP mRNA expression and protein localization, whereas the MAPK8 pathway is a regulator of apoptosis.</p>
https://ir.lib.uwo.ca/obsgynpub/32
oai:ir.lib.uwo.ca:obsgynpub-1035
2016-08-29T18:16:39Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
18239045
Preimplantation embryo programming: transcription, epigenetics, and culture environment.
Duranthon, Veronique
Watson, Andrew J
Lonergan, Patrick
Article
2008-02-01T08:00:00Z
Animals
Blastocyst
Embryo Culture Techniques
Embryonic Induction
Epigenesis
Genetic
Female
Gene Expression Regulation
Developmental
Humans
Mammals
Mice
Models
Animal
Nuclear Transfer Techniques
Pregnancy
Transcription
Genetic
Reproduction (Cambridge, England)
135
2
141
150
Obstetrics and Gynecology
<p>Preimplantation development directs the formation of an implantation- or attachment-competent embryo so that metabolic interactions with the uterus can occur, pregnancy can be initiated, and fetal development can be sustained. The preimplantation embryo exhibits a form of autonomous development fueled by products provided by the oocyte and also from activation of the embryo's genome. Despite this autonomy, the preimplantation embryo is highly influenced by factors in the external environment and in extreme situations, such as those presented by embryo culture or nuclear transfer, the ability of the embryo to adapt to the changing environmental conditions or chromatin to become reprogrammed can exceed its own adaptive capacity, resulting in aberrant embryonic development. Nuclear transfer or embryo culture-induced influences not only affect implantation and establishment of pregnancy but also can extend to fetal and postnatal development and affect susceptibility to disease in later life. It is therefore critical to define the basic program controlling preimplantation development, and also to utilize nuclear transfer and embryo culture models so that we may design healthier environments for preimplantation embryos to thrive in and also minimize the potential for negative consequences during pregnancy and post-gestational life. In addition, it is necessary to couple gene expression analysis with the investigation of gene function so that effects on gene expression can be fully understood. The purpose of this short review is to highlight our knowledge of the mechanisms controlling preimplantation development and report how those mechanisms may be influenced by nuclear transfer and embryo culture.</p>
https://ir.lib.uwo.ca/obsgynpub/34
oai:ir.lib.uwo.ca:obsgynpub-1036
2016-08-29T18:18:36Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
17322120
Oocyte cytoplasmic maturation: a key mediator of oocyte and embryo developmental competence.
Watson, A J
Article
2007-03-01T08:00:00Z
Animals
Cytoplasm
Embryo Transfer
Embryonic Development
Female
Humans
Oocytes
Oogenesis
Reproductive Techniques
Transcription
Genetic
Journal of animal science
85
13 Suppl
1
3
Obstetrics and Gynecology
<p>Efforts have intensified to successfully mature and inseminate oocytes in vitro and then culture ensuing embryos to transferable stages from a large number of mammalian species. Success varies, but generally even for the most successful species it is only possible to obtain a maximum of a 40 to 50% development of zygotes to the blastocyst stage. Reduced oocyte developmental competence is suggested as a primary reason for the reduced potential of in vitro-produced embryos. The vast majority of in vitro-matured oocytes are meiotically competent; however, many do not attain an optimal oocyte diameter before insemination. Variations in oocyte in vitro maturation media can influence embryo development, blastocyst cell number, and apoptosis. In addition, studies have indicated that cytoplasmic donation from so-called competent to incompetent oocytes can improve developmental outcomes. Oocyte cytoplasmic maturation includes those events that instill upon the oocyte a capacity to complete nuclear maturation, insemination, early embryogenesis and thus provide a foundation for implantation, initiation of pregnancy, and normal fetal development. Although we can define oocyte cytoplasmic maturation, we are only now beginning to understand the molecular steps that underlie this process. In general terms, oocyte cytoplasmic maturation involves the accumulation of mRNA, proteins, substrates, and nutrients that are required to achieve the oocyte developmental competence that fosters embryonic developmental competence. Collectively we are beginning to specify oocyte cytoplasmic maturation, and eventually a coherent understanding of this critical event in oocyte biology will emerge.</p>
https://ir.lib.uwo.ca/obsgynpub/35
oai:ir.lib.uwo.ca:obsgynpub-1040
2016-08-29T18:22:39Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
16356488
Na+/K+ -ATPase regulates tight junction formation and function during mouse preimplantation development.
Violette, Michelle I
Madan, Pavneesh
Watson, Andrew J
Article
2006-01-15T08:00:00Z
Animals
Blastocyst
Cell Count
Dose-Response Relationship
Drug
Embryonic Development
Enzyme Inhibitors
Female
Male
Membrane Proteins
Mice
Microscopy
Confocal
Microscopy
Fluorescence
Occludin
Ouabain
Phosphoproteins
Sodium-Potassium-Exchanging ATPase
Tight Junctions
Time Factors
Zonula Occludens-1 Protein
Developmental biology
289
2
406
419
Obstetrics and Gynecology
<p>Research applied to the early embryo is required to effectively treat human infertility and to understand the primary mechanisms controlling development to the blastocyst stage. The present study investigated whether the Na(+)/K(+)-ATPase regulates tight junction formation and function during blastocyst formation. To investigate this hypothesis, three experimental series were conducted. The first experiments defined the optimal dose and treatment time intervals for ouabain (a potent and specific inhibitor of the Na(+)/K(+)-ATPase) treatment. The results demonstrated that mouse embryos maintained a normal development to the blastocyst stage following a 6-h ouabain treatment. The second experiments investigated the effects of ouabain treatment on the distribution of ZO-1 and occludin (tight junction associated proteins). Ouabain treatment (up to 6 h) or culture in K(+)-free medium (up to 6 h) resulted in the appearance of a discontinuous ZO-1 protein distribution and a loss of occludin immunofluorescence. The third set of experiments examined the influence of ouabain treatment on tight junction function. Ouabain treatment or culture in K(+)-free medium affected tight junction permeability as indicated by an increase in the proportion of treated embryos accumulating both 4 kDa and 40 kDa fluorescein isothiocyanate (FITC)-dextran into their blastocyst cavities. The results indicate that the Na(+)/K(+)-ATPase is a potent regulator of tight junction formation and function during mouse preimplantation development.</p>
https://ir.lib.uwo.ca/obsgynpub/39
oai:ir.lib.uwo.ca:obsgynpub-1045
2016-08-29T18:28:43Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
15831278
Effect of serum and cumulus cell expansion on marker gene transcripts in bovine cumulus-oocyte complexes during maturation in vitro.
Calder, Michele D
Caveney, Anita N
Sirard, Marc-Andre
Watson, Andrew J
Article
2005-04-01T08:00:00Z
Animals
Blood Proteins
Cattle
Cyclooxygenase 2
Female
Fertilization in Vitro
Gene Expression
Genetic Markers
Gonadotropins
In Vitro Techniques
Oocytes
Prostaglandin-Endoperoxide Synthases
RNA
Messenger
Receptors
FSH
Receptors
LH
Receptors
Prostaglandin E
Receptors
Prostaglandin E
EP2 Subtype
Receptors
Prostaglandin E
EP3 Subtype
Transcription
Genetic
Fertility and sterility
83 Suppl 1
1077
1085
Obstetrics and Gynecology
<p>OBJECTIVE: To determine the distribution of transcripts encoding the FSH receptor (FSHr), LH receptor (LHr), connexin 43 (Cx43), cyclooxygenase-2 (COX-2), and prostaglandin E(2) receptors 2 and 3 (EP2 and EP3) within bovine cumulus-oocyte complexes (COCs) and denuded oocytes and investigate the influence of gonadotropins, serum, and cumulus cell expansion on the abundance of transcripts encoding these genes.</p>
<p>DESIGN: Prospective controlled animal study.</p>
<p>SETTING: University research laboratory.</p>
<p>PATIENT(S): Animal models for human studies.</p>
<p>INTERVENTION(S): Cumulus-oocyte complexes were treated in culture with serum and gonadotropin-supplemented media to examine the effects to mRNA transcript levels.</p>
<p>MAIN OUTCOME MEASURE(S): Variation in mRNA transcript levels.</p>
<p>RESULT(S): Luteinizing hormone receptor, FSHr, and EP3 mRNAs were detected in intact COCs and not in cumulus cell-denuded oocytes, whereas Cx43, COX-2, and EP2 mRNAs were found in both COCs and oocytes. The relative abundance of marker gene mRNAs did not vary in media containing no additives or FSH alone, independent of whether the media induced cumulus cell expansion. However, the presence of serum in maturation media significantly decreased expression of all mRNAs except LHr.</p>
<p>CONCLUSION(S): The relative abundance of COC mRNAs is altered by serum in the maturation medium, which may signify long-term consequences for embryonic development.</p>
https://ir.lib.uwo.ca/obsgynpub/42
oai:ir.lib.uwo.ca:obsgynpub-1041
2016-08-31T18:34:56Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
16218831
Reprogramming of fibroblast nuclei after transfer into bovine oocytes.
De Sousa, P A
Winger, Q
Hill, J R
Jones, K
Watson, A J
Westhusin, M E
Article
1999-01-01T08:00:00Z
Animals
Blastocyst
Cattle
Cell Nucleus
Cloning
Organism
Female
Fibroblasts
Gene Expression
Gene Expression Profiling
Nuclear Transfer Techniques
Oocytes
RNA
Messenger
Cloning
1
1
63
69
Obstetrics and Gynecology
<p>Recent landmark achievements in animal cloning have demonstrated that the events of cell differentiation can, in principle, be reversed. This reversal necessarily requires large-scale genetic reprogramming, of which little is known. In the present study we characterized the extent to which blastocyst stage-specific mRNA expression would be conserved in bovine embryos produced by nuclear transfer (NT) using fetal fibroblasts as nuclei donors (FF NT). The mRNA pool of FF NT embryos was compared with that of NT embryos reconstructed from embryonic blastomeres (Emb NT), with embryos produced under in vivo or in vitro conditions, and finally with fibroblast cells. Embryo/cell-specific mRNA pools were contrasted using differential display methodology. Random oligonucleotide primer pair combinations were used to subfractionate mRNA populations and represent individual mRNAs as copy DNA (cDNA) bands ranging in size from 100 to 800 base pairs. Regardless of whether bovine blastocysts developed in vivo or in vitro, or were derived after nuclear transplantation with embryonic blastomeres or fetal fibroblasts, their mRNA profile was highly conserved and distinct from that of fetal fibroblast cells. There was approximately 95% conservation in cDNA banding patterns between FF NT, Emb NT, and in vivo derived blastocysts, when compared with in vitro derived blastocysts. In contrast, the cDNA banding in fibroblasts was only 67% conserved with in vitro derived blastocysts (p < 0.0001), indicating that dramatic changes in gene transcription are induced by nuclear transplantation. After nuclear transplantation, gene expression in fetal fibroblasts is reprogrammed so to mimic that of preimplantation embryo development. Future characterization of these changes will be invaluable for the identification of suitable cell types to serve as nuclear donors for embryo reconstruction and provide information that can be used to improve the efficiency of cloning animals by nuclear transplantation.</p>
https://ir.lib.uwo.ca/obsgynpub/44
oai:ir.lib.uwo.ca:obsgynpub-1044
2016-08-31T18:39:38Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
15850458
p38 mitogen-activated protein kinase (MAPK) first regulates filamentous actin at the 8-16-cell stage during preimplantation development.
Paliga, Andrew J M
Natale, David R
Watson, Andrew J
Article
2005-08-01T07:00:00Z
Actins
Animals
Blastocyst
Cadherins
Cell Adhesion Molecules
Cytoskeletal Proteins
Dose-Response Relationship
Drug
Embryonic Development
Enzyme Inhibitors
Female
Flavonoids
Fluorescent Antibody Technique
Indirect
Heat-Shock Proteins
Imidazoles
Intracellular Signaling Peptides and Proteins
Mice
Mitogen-Activated Protein Kinase 3
Morula
Pregnancy
Protein-Serine-Threonine Kinases
Pyridines
Pyrimidines
Signal Transduction
Time Factors
alpha Catenin
p38 Mitogen-Activated Protein Kinases
Biology of the cell / under the auspices of the European Cell Biology Organization
97
8
629
640
Obstetrics and Gynecology
<p>BACKGROUND INFORMATION: The MAPK (mitogen-activated protein kinase) superfamily of proteins consists of four separate signalling cascades: the c-Jun N-terminal kinase or stress-activated protein kinases (JNK/SAPK); the ERKs (extracellular-signal-regulated kinases); the ERK5 or big MAPK1; and the p38 MAPK group of protein kinases, all of which are highly conserved. To date, our studies have focused on defining the role of the p38 MAPK pathway during preimplantation development. p38 MAPK regulates actin filament formation through the downstream kinases MAPKAPK2/3 (MAPK-activated protein kinase 2/3) or MAPKAPK5 [PRAK (p38 regulated/activated kinase)] and subsequently through HSP25/27 (heat-shock protein 25/27). We recently reported that 2-cell-stage murine embryos treated with cytokine-suppressive anti-inflammatory drugs (CSAIDtrade mark; SB203580 and SB220025) display a reversible blockade of development at the 8-16-cell stage, indicating that p38 (MAPK) activity is required to complete murine preimplantation development. In the present study, we have investigated the stage-specific action and role of p38 MAPK in regulating filamentous actin during murine preimplantation development.</p>
<p>RESULTS: Treatment of 8-cell-stage embryos with SB203580 and SB220025 (CSAIDtrade mark) resulted in a blockade of preimplantation development, loss of rhodamine phalloidin fluorescence, MK-p (phosphorylated MAPKAPK2/3), HSP-p (phosphorylated HSP25/27) and a redistribution of alpha-catenin immunofluorescence by 12 h of treatment. In contrast, treatment of 2- and 4-cell-stage embryos with CSAIDtrade mark drugs resulted in a loss of MK-p and HSP-p, but did not result in a loss of rhodamine phalloidin fluorescence. All these effects of p38 MAPK inhibition were reversed upon removal of the inhibitor, and development resumed in a delayed but normal manner to the blastocyst stage. Treatment of 8-cell embryos with PD098059 (ERK pathway inhibitor) did not affect development or fluorescence of MK-p, HSP-p or rhodamine phalloidin.</p>
<p>CONCLUSION: Murine preimplantation development becomes dependent on p38 MAPK at the 8-16-cell stage, which corresponds to the stage when p38 MAPK first regulates filamentous actin during early development.</p>
https://ir.lib.uwo.ca/obsgynpub/45
oai:ir.lib.uwo.ca:obsgynpub-1034
2016-08-30T18:50:10Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
18405437
Bovine oocytes and early embryos express Staufen and ELAVL RNA-binding proteins.
Calder, M D
Madan, P
Watson, A J
Article
2008-05-01T07:00:00Z
Animals
Cattle
Embryo
Mammalian
Fertilization in Vitro
Oocytes
RNA
Messenger
RNA-Binding Proteins
Zygote (Cambridge, England)
16
2
161
168
Obstetrics and Gynecology
<p>RNA-binding proteins (RBP) influence RNA editing, localization, stability and translation and may contribute to oocyte developmental competence by regulating the stability and turnover of oogenetic mRNAs. The expression of Staufen 1 and 2 and ELAVL1, ELAVL2 RNA-binding proteins during cow early development was characterized. Cumulus-oocyte complexes were collected from slaughterhouse ovaries, matured, inseminated and subjected to embryo culture in vitro. Oocyte or preimplantation embryo pools were processed for RT-PCR and whole-mount immunofluorescence analysis of mRNA expression and protein distribution. STAU1 and STAU2 and ELAVL1 mRNAs and proteins were detected throughout cow preimplantation development from the germinal vesicle (GV) oocyte to the blastocyst stage. ELAVL2 mRNAs were detectable from the GV to the morula stage, whereas ELAVL2 protein was in all stages examined and localized to both cytoplasm and nuclei. The findings provide a foundation for investigating the role of RBPs during mammalian oocyte maturation and early embryogenesis.</p>
https://ir.lib.uwo.ca/obsgynpub/73
oai:ir.lib.uwo.ca:obsgynpub-1037
2016-08-30T19:16:46Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
17317668
Na/K-ATPase beta1 subunit expression is required for blastocyst formation and normal assembly of trophectoderm tight junction-associated proteins.
Madan, Pavneesh
Rose, Keeley
Watson, Andrew J
Article
2007-04-20T07:00:00Z
Animals
Blastocyst
Down-Regulation
Ectoderm
Embryo
Mammalian
Embryonic Development
Female
Fluorescent Antibody Technique
Indirect
Gene Expression Regulation
Developmental
Mice
Microscopy
Fluorescence
Peptides
Sodium-Potassium-Exchanging ATPase
Tight Junctions
The Journal of biological chemistry
282
16
12127
12134
Obstetrics and Gynecology
<p>Na/K-ATPase plays an important role in mediating blastocyst formation. Despite the expression of multiple Na/K-ATPase alpha and beta isoforms during mouse preimplantation development, only the alpha1 and beta1 isoforms have been localized to the basolateral membrane regions of the trophectoderm. The aim of the present study was to selectively down-regulate the Na/K-ATPase beta1 subunit employing microinjection of mouse 1 cell zygotes with small interfering RNA (siRNA) oligos. Experiments comprised of non-injected controls and two groups microinjected with either Stealthtrade mark Na/K-ATPase beta1 subunit oligos or nonspecific Stealthtrade mark siRNA as control. Development to the 2-, 4-, 8-, and 16-cell and morula stages did not vary between the three groups. However, only 2.3% of the embryos microinjected with Na/K-ATPase beta1 subunit siRNA oligos developed to the blastocyst stage as compared with 73% for control-injected and 91% for non-injected controls. Na/K-ATPase beta1 subunit down-regulation was validated by employing reverse transcription-PCR and whole-mount immunofluorescence methods to demonstrate that Na/K-ATPase beta1 subunit mRNAs and protein were not detectable in beta1 subunit siRNA-microinjected embryos. Aggregation chimera experiments between beta1 subunit siRNA-microinjected embryos and controls demonstrated that blockade of blastocyst formation was reversible. The distribution of Na/K-ATPase alpha1 and tight junction-associated proteins occludin and ZO-1 were compared among the three treatment groups. No differences in protein distribution were observed between control groups; however, all three polypeptides displayed an aberrant distribution in Na/K-ATPase beta1 subunit siRNA-microinjected embryos. Our results demonstrate that the beta1 subunit of the Na/K-ATPase is required for blastocyst formation and that this subunit is also required to maintain a normal Na/K-ATPase distribution and localization of tight junction-associated polypeptides during preimplantation development.</p>
<p>This research was originally published in The Journal of Biological Chemistry, 2007. Vol16:pp.12127-12134. © the American Society for Biochemistry and Molecular Biology.</p>
https://ir.lib.uwo.ca/obsgynpub/37
oai:ir.lib.uwo.ca:obsgynpub-1033
2016-08-29T18:14:29Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
19043080
Genomic RNA profiling and the programme controlling preimplantation mammalian development.
Bell, Christine E
Calder, Michele D
Watson, Andrew J
Article
2008-12-01T08:00:00Z
Animals
Blastocyst
Cell Lineage
Embryo Culture Techniques
Embryonic Development
Gene Expression Profiling
Gene Expression Regulation
Developmental
Humans
MicroRNAs
RNA
RNA Stability
Molecular human reproduction
14
12
691
701
Obstetrics and Gynecology
<p>Preimplantation development shifts from a maternal to embryonic programme rapidly after fertilization. Although the majority of oogenetic products are lost during the maternal to embryonic transition (MET), several do survive this interval to contribute directly to supporting preimplantation development. Embryonic genome activation (EGA) is characterized by the transient expression of several genes that are necessary for MET, and while EGA represents the first major wave of gene expression, a second mid-preimplantation wave of transcription that supports development to the blastocyst stage has been discovered. The application of genomic approaches has greatly assisted in the discovery of stage specific gene expression patterns and the challenge now is to largely define gene function and regulation during preimplantation development. The basic mechanisms controlling compaction, lineage specification and blastocyst formation are defined. The requirement for embryo culture has revealed plasticity in the developmental programme that may exceed the adaptive capacity of the embryo and has fostered important research directions aimed at alleviating culture-induced changes in embryonic programming. New levels of regulation are emerging and greater insight into the roles played by RNA-binding proteins and miRNAs is required. All of this research is relevant due to the necessity to produce healthy preimplantation embryos for embryo transfer, to ensure that assisted reproductive technologies are applied in the most efficient and safest way possible.</p>
https://ir.lib.uwo.ca/obsgynpub/33
oai:ir.lib.uwo.ca:obsgynpub-1039
2016-08-29T18:21:23Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
17214902
Mouse preimplantation embryo responses to culture medium osmolarity include increased expression of CCM2 and p38 MAPK activation.
Fong, Barry
Watson, Patricia H
Watson, Andrew J
Article
2007-01-01T08:00:00Z
Animals
Blastocyst
Cells
Cultured
Culture Media
Enzyme Activation
Female
Fluorescent Antibody Technique
Indirect
Gene Expression
Mice
Microfilament Proteins
Microscopy
Confocal
Osmolar Concentration
Phosphorylation
Pregnancy
Reverse Transcriptase Polymerase Chain Reaction
p38 Mitogen-Activated Protein Kinases
BMC developmental biology [electronic resource]
7
2
2
Obstetrics and Gynecology
<p>BACKGROUND: Mechanisms that confer an ability to respond positively to environmental osmolarity are fundamental to ensuring embryo survival during the preimplantation period. Activation of p38 mitogen-activated protein kinase (MAPK) occurs following exposure to hyperosmotic treatment. Recently, a novel scaffolding protein called Osmosensing Scaffold for MEKK3 (OSM) was linked to p38 MAPK activation in response to sorbitol-induced hypertonicity. The human ortholog of OSM is cerebral cavernous malformation 2 (CCM2). The present study was conducted to investigate whether CCM2 is expressed during mouse preimplantation development and to determine whether this scaffolding protein is associated with p38 MAPK activation following exposure of preimplantation embryos to hyperosmotic environments.</p>
<p>RESULTS: Our results indicate that Ccm2 along with upstream p38 MAPK pathway constituents (Map3k3, Map2k3, Map2k6, and Map2k4) are expressed throughout mouse preimplantation development. CCM2, MAP3K3 and the phosphorylated forms of MAP2K3/MAP2K6 and MAP2K4 were also detected throughout preimplantation development. Embryo culture in hyperosmotic media increased p38 MAPK activity in conjunction with elevated CCM2 levels.</p>
<p>CONCLUSION: These results define the expression of upstream activators of p38 MAPK during preimplantation development and indicate that embryo responses to hyperosmotic environments include elevation of CCM2 and activation of p38 MAPK.</p>
https://ir.lib.uwo.ca/obsgynpub/38
oai:ir.lib.uwo.ca:obsgynpub-1043
2016-08-29T18:26:53Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
15985630
Mitogen-activated protein kinase (MAPK) blockade of bovine preimplantation embryogenesis requires inhibition of both p38 and extracellular signal-regulated kinase (ERK) pathways.
Madan, Pavneesh
Calder, Michele D
Watson, Andrew J
Article
2005-07-01T07:00:00Z
Actins
Animals
Blastocyst
Butadienes
Cattle
Embryo Culture Techniques
Embryonic Development
Enzyme Inhibitors
Extracellular Signal-Regulated MAP Kinases
Fluorescent Antibody Technique
Indirect
HSP27 Heat-Shock Proteins
Heat-Shock Proteins
Imidazoles
Intracellular Signaling Peptides and Proteins
MAP Kinase Signaling System
Neoplasm Proteins
Nitriles
Phosphorylation
Protein Kinases
Protein-Serine-Threonine Kinases
Pyrimidines
Reverse Transcriptase Polymerase Chain Reaction
Signal Transduction
Time Factors
p38 Mitogen-Activated Protein Kinases
Reproduction (Cambridge, England)
130
1
41
51
Obstetrics and Gynecology
<p>Blastocyst formation, as a critical period during development, is an effective indicator of embryonic health and reproductive efficiency. Out of a number of mechanisms underlying blastocyst formation, highly conserved mitogen-activated protein kinase (MAPK) signaling has emerged as a major mechanism involved in regulating murine preimplantation embryo development. The objective of our study was to ascertain the role of MAPK signaling in regulating bovine development to the blastocyst stage. Using reverse transcriptase PCR and immunohistochemical staining procedures we have demonstrated that mRNA transcripts and polypeptides encoding p38 MAPK pathway constituents are detectable in preimplantation bovine embryos from the one-cell to the blastocyst stage. Further, the effects on bovine embryo development following inhibition of p38 alpha/beta and extracellular signal-regulated kinase (ERK) signaling by treatment with SB220025 and U0126, respectively, were investigated. Eight-cell bovine embryos (50 per group; three replicates) were placed into treatments consisting of synthetic oviductal fluid (SOF) medium: SOF + SB202474 (inactive analogue), SOF + SB220025, SOF + U0124 (inactive analogue), SOF + U0126, and SOF + SB220025 + U0126. Inhibition of p38 MAPK or ERK signaling individually did not affect development to the blastocyst stage. However, when both pathways were blocked simultaneously there was a significant reduction (P < 0.05) in blastocyst formation, cell number and immunofluorescence of phosphorylated downstream pathway constituents. We have determined that, in variance to what was observed during murine preimplantation development, bovine early embryos progress at normal frequencies to the blastocyst stage in the presence of p38 MAPK inhibitors.</p>
https://ir.lib.uwo.ca/obsgynpub/40
oai:ir.lib.uwo.ca:obsgynpub-1042
2016-08-29T18:25:17Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
16139691
Roles of Na,K-ATPase in early development and trophectoderm differentiation.
Kidder, Gerald M
Watson, Andrew J
Article
2005-09-01T07:00:00Z
Animals
Cell Differentiation
Humans
Isoenzymes
Protein Subunits
Sodium-Potassium-Exchanging ATPase
Trophoblasts
Seminars in nephrology
25
5
352
355
Obstetrics and Gynecology
<p>Before implantation into the uterine wall, the mammalian embryo undergoes a period of cell division, cell shape change, and cell differentiation leading to the formation of an outer epithelium, the trophectoderm. The trophectoderm is the part of the embryo that initiates uterine contact and, after transformation to become the trophoblast, uterine invasion. Similar to the kidney nephron, the trophectoderm is a transporting epithelium with distinct apical and basolateral membrane domains; its function is to facilitate transepithelial Na+ and fluid transport for blastocoel formation. That transport is driven by Na,K-adenosine triphosphatase (ATPase) localized in basolateral membranes of the trophectoderm. Preimplantation embryos express multiple alpha and beta subunit isoforms of Na,K-ATPase, potentially constituting multiple isozymes, but the basolaterally located alpha1beta1 isozyme appears to function uniquely to drive fluid transport. Embryos unable to express alpha1 subunits because of targeted deletion of the gene are able to form a blastocoel, but they fail to maintain their integrity and expire during the peri-implantation period. Preimplantation embryos also express the gamma subunit, a modulator of Na,K-ATPase activity, but targeted deletion of that gene did not reveal an essential developmental role. The preimplantation embryo offers a unique model for understanding the roles of Na,K-ATPase subunit isoforms in epithelial development and transepithelial transport.</p>
https://ir.lib.uwo.ca/obsgynpub/41
oai:ir.lib.uwo.ca:obsgynpub-1038
2016-08-29T18:20:28Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
17219434
PP2Cdelta (Ppm1d, WIP1), an endogenous inhibitor of p38 MAPK, is regulated along with Trp53 and Cdkn2a following p38 MAPK inhibition during mouse preimplantation development.
Hickson, Jenny A
Fong, Barry
Watson, Patricia H
Watson, Andrew J
Article
2007-07-01T07:00:00Z
Animals
Blastocyst
Cyclin-Dependent Kinase Inhibitor p16
Enzyme Inhibitors
Female
Gene Expression Regulation
Developmental
MAP Kinase Signaling System
Male
Mice
Mitogen-Activated Protein Kinase 11
Mitogen-Activated Protein Kinase 14
Phosphoprotein Phosphatases
RNA
Tumor Suppressor Protein p53
p38 Mitogen-Activated Protein Kinases
Molecular reproduction and development
74
7
821
834
Obstetrics and Gynecology
<p>Preimplantation embryos utilize mitogen-activated protein kinase signaling (MAPK) pathways to relay signals from the external environment to prepare appropriate responses and adaptations to a changing milieu. It is therefore important to investigate how MAPK pathways are regulated during preimplantation development. This study was conducted to investigate whether PP2Cdelta (Ppm1d, WIP1) is expressed during mouse preimplantation development and to determine the influences of p38 MAPK inhibition on expression of Trp53 (p53), Ppm1d, (WIP1), and Cdkn2a (p16) during mouse preimplantation development. Our results indicate that Trp53, Ppm1d, and Cdkn2a mRNAs and TRP53 and PP2Cdelta proteins are expressed throughout mouse preimplantation development. Treatment of 2-cell embryos with SB220025 (potent inhibitor of p38 MAPK alpha/beta/MAPK 14/11) significantly increased Trp53, Ppm1d and Cdkn2a and Mapk14 mRNA levels at 12 and 24 hr. Treatment of 8-cell embryos with SB220025 for 12 hr increased Trp53, Ppm1d, and Cdkn2a mRNA levels, but not Mapk14 mRNA levels. Treatment of 8-cell embryos for 24 hr increased Trp53, and Ppm1d mRNA levels, but decreased Cdkn2a and Mapk14 mRNA levels. Therefore, blockade of p38 MAPK activity is associated with embryo stage specific influences on Trp53, Ppm1d, Cdkn2a, and Mapk14 expression during mouse preimplantation development. These results define downstream targets of p38 MAPK during preimplantation development and indicate that the p38 MAPK pathway regulates Trp53, Ppm1d, and Cdkn2a expression. This study increases our understanding of the mechanisms controlling preimplantation development and of the interactions between preimplantation embryos and their culture environments.</p>
https://ir.lib.uwo.ca/obsgynpub/43
oai:ir.lib.uwo.ca:obsgynpub-1047
2016-08-29T18:30:52Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
15525537
RGS14 is a mitotic spindle protein essential from the first division of the mammalian zygote.
Martin-McCaffrey, Luke
Willard, Francis S
Oliveira-dos-Santos, Antonio J
Natale, David R C
Snow, Bryan E
Kimple, Randall J
Pajak, Agnieszka
Watson, Andrew J
Dagnino, Lina
Penninger, Josef M
Siderovski, David P
D'Souza, Sudhir J A
Article
2004-11-01T08:00:00Z
Animals
Antibodies
Monoclonal
Blastocyst
Cell Division
Fluorescein-5-isothiocyanate
Fluorescent Dyes
Gene Deletion
Genetic Vectors
HeLa Cells
Heterozygote
Humans
Hydrazines
Mammals
Mice
Mice
Knockout
Microscopy
Fluorescence
Molecular Sequence Data
RGS Proteins
RNA Interference
RNA
Small Interfering
Rats
Spindle Apparatus
Zygote
Developmental cell
7
5
763
769
Obstetrics and Gynecology
<p>Heterotrimeric G protein alpha subunits, RGS proteins, and GoLoco motif proteins have been recently implicated in the control of mitotic spindle dynamics in C. elegans and D. melanogaster. Here we show that "regulator of G protein signaling-14" (RGS14) is expressed by the mouse embryonic genome immediately prior to the first mitosis, where it colocalizes with the anastral mitotic apparatus of the mouse zygote. Loss of Rgs14 expression in the mouse zygote results in cytofragmentation and failure to progress to the 2-cell stage. RGS14 is found in all tissues and segregates to the nucleus in interphase and to the mitotic spindle and centrioles during mitosis. Alteration of RGS14 levels in exponentially proliferating cells leads to cell growth arrest. Our results indicate that RGS14 is one of the earliest essential product of the mammalian embryonic genome yet described and has a general role in mitosis.</p>
https://ir.lib.uwo.ca/obsgynpub/51
oai:ir.lib.uwo.ca:obsgynpub-1051
2016-09-01T20:02:30Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
14516655
Rac-1 and IQGAP are potential regulators of E-cadherin-catenin interactions during murine preimplantation development.
Natale, David R
Watson, Andrew J
Article
2002-12-01T08:00:00Z
Animals
Blastocyst
Cadherins
Cell Membrane
Embryonic Development
Mice
beta Catenin
Mechanisms of development
119 Suppl 1
21
26
Obstetrics and Gynecology
<p>Adherens junction formation is fundamental for compaction and trophectoderm differentiation during mammalian preimplantation development. We recently isolated an IQGAP-2 cDNA from a differential display-polymerase chain reaction screen of bovine preimplantation developmental stages. IQGAP-1 and -2 proteins mediate E-cadherin-based cell-to-cell adhesion through interactions with beta-catenin and the Rho GTPases, rac1 and cdc42. Our study demonstrates IQGAP-1,-2, rac-1 and cdc42 mRNAs are present throughout murine preimplantation development. IQGAP-1 and rac-1 protein distribution changes from predominantly plasma membrane associated to predominantly cytoplasmic as the embryo progresses through cleavage divisions and compaction to the blastocyst stage.</p>
https://ir.lib.uwo.ca/obsgynpub/52
oai:ir.lib.uwo.ca:obsgynpub-1050
2016-08-29T18:35:24Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
15031106
p38 MAPK signaling during murine preimplantation development.
Natale, David R
Paliga, Andrew J M
Beier, Frank
D'Souza, S J A
Watson, Andrew J
Article
2004-04-01T08:00:00Z
Animals
Base Sequence
Cloning
Molecular
DNA Primers
Embryonic Development
Enzyme Inhibitors
Female
Fluorescent Antibody Technique
Indirect
Imidazoles
Mice
Mitogen-Activated Protein Kinases
Pregnancy
Pyridines
Pyrimidines
Reverse Transcriptase Polymerase Chain Reaction
Signal Transduction
p38 Mitogen-Activated Protein Kinases
Developmental biology
268
1
76
88
Obstetrics and Gynecology
<p>Mitogen-activated protein kinase (MAPK) pathways mediate some important cellular processes and are likely to also regulate preimplantation development. The role of p38 MAP kinase signaling during murine preimplantation development was investigated in the present study. p38 MAPK, p38-regulated or -activated kinase (PRAK; MK5), map kinase-activated protein kinase 2 (MK2), and heat shock protein 25 (hsp25) mRNAs and proteins were detected throughout preimplantation development. Two-cell stage embryos cultured in the presence of SB220025 and SB203580 (specific inhibitors of p38 MAPK alpha/beta), progressed to the eight-cell stage with the same frequency as controls; however, treated embryos halted their development at the 8- to 16-cell stage. In addition, embryos treated with p38 MAPK inhibitors displayed a complete loss of MK2 and hsp25 phosphorylation and also a complete loss of filamentous actin as indicated by the absence of rhodamine-phalloidin staining. In these inhibitor-treated groups, the embryos were composed of a mixture of compacting and noncompacting cells, and the embryos were one to two cell divisions behind controls. Treated embryos remained viable as the developmental blockade was rescued by removing embryos from the drug treatment and placing them in drug-free medium until they progressed to the blastocyst stage. This study demonstrates that p38 MAPK activity is required to support development through the murine preimplantation interval.</p>
https://ir.lib.uwo.ca/obsgynpub/46
oai:ir.lib.uwo.ca:obsgynpub-1049
2016-08-29T18:34:06Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
15147760
Deletion of the Na/K-ATPase alpha1-subunit gene (Atp1a1) does not prevent cavitation of the preimplantation mouse embryo.
Barcroft, L C
Moseley, A E
Lingrel, J B
Watson, A J
Article
2004-05-01T07:00:00Z
Animals
Blastocyst
Cell Shape
Cell Size
Cells
Cultured
Female
Gene Deletion
Genotype
Homozygote
Immunohistochemistry
Male
Mice
Mice
Knockout
Protein Subunits
Sodium-Potassium-Exchanging ATPase
Time Factors
Mechanisms of development
121
5
417
426
Obstetrics and Gynecology
<p>Increases in Na/K-ATPase activity occur concurrently with the onset of cavitation and are associated with increases in Na(+)-pump subunit mRNA and protein expression. We have hypothesized that the alpha1-isozyme of the Na/K-ATPase is required to mediate blastocyst formation. We have tested this hypothesis by characterizing preimplantation development in mice with a targeted disruption of the Na/K-ATPase alpha1-subunit (Atp1a1) using embryos acquired from matings between Atp1a1 heterozygous mice. Mouse embryos homozygous for a null mutation in the Na/K-ATPase alpha1-subunit gene are able to undergo compaction and cavitation. These findings demonstrate that trophectoderm transport mechanisms are maintained in the absence of the predominant isozyme of the Na(+)-pump that has previously been localized to the basolateral membranes of mammalian trophectoderm cells. The presence of multiple isoforms of Na/K-ATPase alpha- and beta-subunits at the time of cavitation suggests that there may be a degree of genetic redundancy amongst isoforms of the catalytic alpha-subunit that allows blastocyst formation to progress in the absence of the alpha1-subunit.</p>
https://ir.lib.uwo.ca/obsgynpub/48
oai:ir.lib.uwo.ca:obsgynpub-1046
2016-09-26T16:41:43Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
15745631
Potential and limitations of bovine-specific arrays for the analysis of mRNA levels in early development: preliminary analysis using a bovine embryonic array.
Sirard, Marc-André
Dufort, Isabelle
Vallée, Maud
Massicotte, Lyne
Gravel, Catherine
Reghenas, Hélène
Watson, Andrew J
King, W Allan
Robert, Claude
Article
2005-01-01T08:00:00Z
Animals
Cattle
DNA
Complementary
Embryo Culture Techniques
Embryonic Development
Nucleic Acid Hybridization
Oligonucleotide Array Sequence Analysis
RNA
Messenger
Sequence Analysis
DNA
Reproduction, fertility, and development
17
1-2
47
57
http://www.publish.csiro.au/view/journals/dsp_journal_fulltext.cfm?nid=44&f=RD04113
Obstetrics and Gynecology
<p>New insights into the early development of large mammals are becoming available through the measurement of differential mRNA levels in oocytes and preimplantation embryos. These advances in knowledge are rapidly picking up in pace, mainly owing to the advantages brought by new molecular biology approaches being developed. The possibility of amplifying the starting material and therefore making measurements in single embryo units is now feasible. With these tools, the evaluation of variations in gene expression patterns during the preimplantation period or the impact of culture on mRNA levels is now possible. However, it is important to keep in mind that these methods still have limitations associated with sample preparation or the use of the appropriate controls. Even proper methods of analysis are very important to achieve the full benefit of the application of these tools. The present paper describes some of the potential, as well as limitations, of mRNA level analysis in early embryos, especially for microarray analysis. We have generated a bovine cDNA array (>2000 clones) that contains expressed sequence tags (ESTs) collected from various preimplantation development stages. Using this chip, we have initiated the characterisation of global mRNA level patterns of several key developmental stages from the immature oocyte to the blastocyst stage. As expected, the hybridisation results indicate very different expression profiles involving hundreds of genes when comparing oocyte and blastocyst samples to a reference mRNA sample made from a pool of ESTs from pooled somatic tissues. Although this array is still in its preliminary stage and the EST bank has not been processed to contain only unigenes, it is already a very useful tool for discovering candidate genes that may play important roles during early embryonic life.</p>
https://ir.lib.uwo.ca/obsgynpub/54
oai:ir.lib.uwo.ca:obsgynpub-1048
2016-10-21T16:27:19Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
15271481
Molecular regulation of blastocyst formation.
Watson, A J
Natale, D R
Barcroft, L C
Article
2004-07-01T07:00:00Z
Animals
Aquaporins
Blastocyst
Cell Adhesion
Embryonic Development
Female
Humans
Mitogen-Activated Protein Kinases
Pregnancy
Sodium-Potassium-Exchanging ATPase
ras GTPase-Activating Proteins
rho GTP-Binding Proteins
Animal reproduction science
82-83
583
592
Obstetrics and Gynecology
<p>Preimplantation development encompasses the interval from insemination until embryo implantation and thus includes the 'freeliving' period of oviduct and uterine development. Formation of the blastocyst is required for implantation and establishment of pregnancy, and is a principal determinant of embryo quality prior to embryo transfer. Development through this period is regulated by the expression of specific gene families that encode for cell polarity, cell junctional, cytoskeletal, ion transporter, and water channel gene products that direct the acquisition of cell polarity and differentiation of the outer cells of the early embryo. This results in the formation of the trophectoderm, which is the first epithelium of development. This review considers the roles of each of these gene families in trophectoderm differentiation and blastocyst formation. The principal hypothesis under investigation is that blastocyst formation is regulated by a Na/K-ATPase-generated trans-trophectoderm ion gradient that promotes the accumulation of water across the epithelium. This, combined with the formation of the tight junction seal controlling paracellular movement of water between adjacent trophectoderm cells, results in the formation of a fluid-filled blastocyst cavity and the expansion of the blastocyst. Results from recent experiments, however, have cast some doubt on the role of Na/K-ATPase in mediating these events and have defined water channels or Aquaporins (AQPs) as physiological mediators of fluid movement across the trophectoderm. In addition, studies have now implicated mitogen-activated protein kinase (MAPK) signaling as an important mediator of development to the blastocyst stage. Such studies define the physiology of blastocyst formation and serve to support the application of assisted reproductive technologies (ART) to both human and animal species.</p>
https://ir.lib.uwo.ca/obsgynpub/74
oai:ir.lib.uwo.ca:obsgynpub-1054
2016-08-29T18:40:54Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
12646061
Responsiveness of bovine cumulus-oocyte-complexes (COC) to porcine and recombinant human FSH, and the effect of COC quality on gonadotropin receptor and Cx43 marker gene mRNAs during maturation in vitro.
Calder, Michele D
Caveney, Anita N
Smith, Lawrence C
Watson, Andrew J
Article
2003-02-11T08:00:00Z
Animals
Cattle
Connexin 43
Culture Media
Serum-Free
Dose-Response Relationship
Drug
Estradiol
Female
Follicle Stimulating Hormone
Humans
Oocytes
Oogenesis
Ovarian Follicle
RNA
Messenger
Receptors
FSH
Receptors
LH
Recombinant Proteins
Swine
Reproductive biology and endocrinology [electronic resource] : RB&E
1
14
14
Obstetrics and Gynecology
<p>Substantially less development to the blastocyst stage occurs in vitro than in vivo and this may be due to deficiencies in oocyte competence. Although a large proportion of bovine oocytes undergo spontaneous nuclear maturation, less is known about requirements for proper cytoplasmic maturation. Commonly, supraphysiological concentrations of FSH and LH are added to maturation media to improve cumulus expansion, fertilization and embryonic development. Therefore, various concentrations of porcine FSH (pFSH) and recombinant human FSH (rhFSH) were investigated for their effect on bovine cumulus expansion in vitro. Expression of FSHr, LHr and Cx43 mRNAs was determined in cumulus-oocyte complexes to determine whether they would be useful markers of oocyte competence. In serum-free media, only 1000 ng/ml pFSH induced marked cumulus expansion, but the effect of 100 ng/ml pFSH was amplified in the presence of 10% serum. In contrast, cumulus expansion occurred with 1 ng/ml rhFSH in the absence of serum. FSHr mRNA was highest at 0-6 h of maturation, then abundance decreased. Similarly, Cx43 mRNA expression was highest from 0-6 h but decreased by 24 h of maturation. However, the relative abundance of LHr mRNA did not change from 6-24 h of maturation. Decreased levels of FSHr, LHr and Cx43 mRNAs were detected in COCs of poorer quality. In conclusion, expansion of bovine cumulus occurred at low doses of rhFSH in serum-free media. In summary, FSHr, LHr and Cx43 mRNA abundance reflects COC quality and FSHr and Cx43 mRNA expression changes during in vitro maturation; these genes may be useful markers of oocyte developmental competence.</p>
https://ir.lib.uwo.ca/obsgynpub/47
oai:ir.lib.uwo.ca:obsgynpub-1053
2016-08-29T18:39:47Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
12679107
Aquaporin proteins in murine trophectoderm mediate transepithelial water movements during cavitation.
Barcroft, Lisa C
Offenberg, Hanne
Thomsen, Preben
Watson, Andrew J
Article
2003-04-15T07:00:00Z
4-Chloromercuribenzenesulfonate
Animals
Aquaporin 3
Aquaporins
Blastocyst
Enzyme Inhibitors
Fluorescent Antibody Technique
Ion Channels
Mercury
Mice
Trophoblasts
Water
Developmental biology
256
2
342
354
Obstetrics and Gynecology
<p>Mammalian blastocyst formation is dependent on establishment of trophectoderm (TE) ion and fluid transport mechanisms. We have examined the expression and function of aquaporin (AQP) water channels during murine preimplantation development. AQP 3, 8, and 9 proteins demonstrated cell margin-associated staining starting at the 8-cell (AQP 9) or compacted morula (AQP 3 and 8) stages. In blastocysts, AQP 3 and 8 were detected in the basolateral membrane domains of the trophectoderm, while AQP3 was also observed in cell margins of all inner cell mass (ICM) cells. In contrast, AQP 9 was predominantly observed within the apical membrane domains of the TE. Murine blastocysts exposed to hyperosmotic culture media (1800 mOsm; 10% glycerol) demonstrated a rapid volume decrease followed by recovery to approximately 80% of initial volume over 5 min. Treatment of blastocysts with p-chloromercuriphenylsulfonic acid (pCMPS, > or =100 microM) for 5 min significantly impaired (P < 0.05) volume recovery, indicating the involvement of AQPs in fluid transport across the TE. Blastocysts exposure to an 1800-mOsm sucrose/KSOMaa solution did not demonstrate volume recovery as observed following treatment with glycerol containing medium, indicating glycerol permeability via AQPs 3 and 9. These findings support the hypothesis that aquaporins mediate trans-trophectodermal water movements during cavitation.</p>
https://ir.lib.uwo.ca/obsgynpub/50
oai:ir.lib.uwo.ca:obsgynpub-1055
2016-09-26T16:46:44Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
12412042
Targeting gene expression in the preimplantation mouse embryo using morpholino antisense oligonucleotides.
Siddall, Laura S
Barcroft, Lisa C
Watson, Andrew J
Article
2002-12-01T08:00:00Z
Animals
Blastocyst
Cytoskeletal Proteins
Dose-Response Relationship
Drug
Female
Fluorescent Antibody Technique
Gene Expression Regulation
Developmental
Lysophosphatidylcholines
Mice
Morpholines
Oligonucleotides
Antisense
Phosphatidylethanolamines
RNA
Messenger
alpha Catenin
Molecular reproduction and development
63
4
413
421
Obstetrics and Gynecology
<p>Morpholino antisense oligonucleotides act by blocking translation of their target gene products and are effective tools for down-regulating gene expression. The current study was conducted to define treatment conditions for the use of morpholino oligonucleotides (MOs) in mammalian preimplantation embryos, and to employ MOs to target genes and study gene function in the early embryo. For the first time, ethoxylated polyethylenimine (EPEI), Lipofectin or Lysolecithin delivery agents were employed in combination with a fluorescent control MO and an alpha-catenin specific MO, to down-regulate gene expression during murine preimplantation development. Experiments applied to both two- and eight-cell stage murine preimplantation embryos contrasted the efficacy of MO concentrations of 1, 2, 5, 10, and 20 microM and treatment delivery times of 3, 6, 24, and 48 hr. Continuous treatment of two-cell embryos with Lipofectin and 20 microM alpha-catenin MO for 48 hr resulted in a significant (P < 0.05) reduction in development to the blastocyst stage and was accompanied by a marked reduction in alpha-catenin protein. These results indicate that morpholino antisense oligonucleotides are effective tools for down-regulating gene expression during mammalian preimplantation development.</p>
https://ir.lib.uwo.ca/obsgynpub/55
oai:ir.lib.uwo.ca:obsgynpub-1056
2016-09-26T16:54:46Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
12239104
Ovarian stanniocalcin is structurally unique in mammals and its production and release are regulated through the luteinizing hormone receptor.
Paciga, Mark
Watson, Andrew J
DiMattia, Gabriel E
Wagner, Graham F
Article
2002-10-01T07:00:00Z
Adenylyl Cyclases
Animals
Cattle
Cells
Cultured
Cyclic AMP-Dependent Protein Kinases
Enzyme Activation
Enzyme Inhibitors
Female
Gene Expression
Glycoproteins
Hormones
Isoquinolines
Molecular Weight
Ovary
Rats
Rats
Sprague-Dawley
Receptors
LH
Sulfonamides
Theca Cells
Endocrinology
143
10
3925
3934
Obstetrics and Gynecology
<p>Stanniocalcin (STC) is a recently discovered mammalian hormone that is widely distributed in many tissues. In rodents the STC gene is most highly expressed in ovary, specifically in androgen-producing thecal and interstitial cells. In addition, ovarian levels of expression rise 15-fold over pregnancy. The objective of this study was to develop a primary culture system for ovarian thecal-interstitial cells (TICs) to identify factors governing STC production and release. We used highly purified primary cultures of rat and bovine TICs, the purity of which was routinely assessed with antigenic and enzymatic markers. The functionality of cells was assured by their responsiveness to LH in the form of progesterone release. We found that forskolin significantly increased STC gene expression and secretion by both rat and bovine TICs, an effect that was only replicated by human (h) chorionic gonadotropin (CG). Coincubation of TICs with hCG and phosphodiesterase inhibitors further increased STC secretion, whereas coincubation of TICs with hCG and protein kinase A inhibitors attenuated hCG-stimulated release. Intriguingly, ovarian STC proved to be substantially larger than the 50-kDa homodimer produced in most other tissues. These results indicate that ovarian STC is physically distinct, a feature that could explain its presence in serum during pregnancy and lactation.</p>
https://ir.lib.uwo.ca/obsgynpub/56
oai:ir.lib.uwo.ca:obsgynpub-1057
2016-08-29T18:43:52Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
12201812
The gamma-subunit of the Na-K-ATPase as a potential regulator of apical and basolateral Na+-pump isozymes during development of bovine pre-attachment embryos.
Barcroft, L C
Gill, S E
Watson, A J
Article
2002-09-01T07:00:00Z
Animals
Base Sequence
Blastocyst
Cattle
Embryonic Development
Female
Fluorescent Antibody Technique
Indirect
Gene Expression
Isoenzymes
Molecular Sequence Data
Pregnancy
RNA
Messenger
Sodium
Sodium-Potassium-Exchanging ATPase
Transcription
Genetic
Reproduction (Cambridge, England)
124
3
387
397
Obstetrics and Gynecology
<p>Expression and activity of the Na-K-ATPase within the basolateral membrane domains of the trophectoderm epithelium provide the driving force for accumulation of Na(+) and Cl(-) across the nascent epithelium, mediating fluid movement into the forming blastocoel. Within the trophectoderm of the bovine blastocyst, multiple isozymes of the Na-K-ATPase are expressed. Immunolocalization has demonstrated that the alpha1-isozyme localizes within the basolateral membrane, whereas the alpha 3-isozyme localizes to the apical cell margins. Gene-specific RT-PCR and wholemount indirect immunofluorescence confocal laser scanning microscopy were used to examine expression of the Na-K-ATPase gamma-subunit (a regulatory subunit of the Na-K-ATPase) throughout development of bovine preattachment embryos in vitro. Expression of mRNA transcripts for the gamma-subunit was detected throughout bovine pre-attachment development from the fertilized one-cell embryo to the blastocyst stage. A similar pattern of expression was also observed for gamma-subunit protein, and immunofluorescence was detected within the membranes of embryonic blastomeres at all stages of development. In contrast to the expression patterns observed for the alpha-subunits, gamma-subunit proteins were detected in both the basolateral and apical cell margins of the trophectoderm, and surrounding all cells of the inner cell mass. Co-localization studies demonstrated that gamma-subunit peptides are co-expressed with the alpha1-subunit in the basolateral domains of the trophectoderm. These results indicate a role for the gamma-subunit of the Na-K-ATPase in modulating Na(+)-pump activity in both apical and basolateral margins of the trophectoderm during formation and expansion of the bovine blastocyst, and adds a further level of complexity to Na(+)-pump regulation of cavitation.</p>
https://ir.lib.uwo.ca/obsgynpub/20
oai:ir.lib.uwo.ca:obsgynpub-1052
2016-08-29T18:38:47Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
14499643
A null mutation for tissue inhibitor of metalloproteinases-3 (Timp-3) impairs murine bronchiole branching morphogenesis.
Gill, Sean E
Pape, M Cynthia
Khokha, Rama
Watson, Andrew J
Leco, Kevin J
Article
2003-09-15T07:00:00Z
Animals
Bronchi
Female
Fibronectins
Male
Metalloendopeptidases
Mice
Mice
Inbred C57BL
Mice
Knockout
Mutation
Tissue Inhibitor of Metalloproteinase-3
Developmental biology
261
2
313
323
Obstetrics and Gynecology
<p>Tissue inhibitors of metalloproteinases (TIMPs) regulate extracellular matrix (ECM) degradation by matrix metalloproteinases (MMPs). We have examined the role of TIMP-3 on ECM homeostasis and bronchiole branching morphogenesis during murine embryogenesis. Employing an in vitro organ culture system, we found decreased bronchiolar branching in null lungs when compared with wild type (WT) counterparts after 2 days in culture. When a synthetic inhibitor of MMPs at low dose was added to the culture system, branching was augmented regardless of genotype. Gelatin and in situ zymography revealed that null lungs exhibited enhanced activation of MMPs throughout lung development. We analysed the impact of increased MMP activity on a number of ECM molecules by Western blot analysis, but found that only fibronectin abundance was consistently reduced in the null lungs throughout development. To confirm that our observed defect in culture was not simply a developmental delay in the null lung, we examined null and WT lungs from newborn pups. Here, we found not only a reduced number of bronchioles in the null, but also that the bronchiole tubes were dilated compared with controls and that alveologenesis was attenuated. We propose that the deletion of TIMP-3 disrupts the exquisite TIMP/MMP balance required for proper focal ECM proteolysis, which leads to correct bronchiole branching morphogenesis in the developing mouse lung.</p>
https://ir.lib.uwo.ca/obsgynpub/49
oai:ir.lib.uwo.ca:obsgynpub-1059
2016-08-29T18:47:17Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
11420233
Cyclooxygenase-2 and prostaglandin E(2)(PGE(2)) receptor messenger RNAs are affected by bovine oocyte maturation time and cumulus-oocyte complex quality, and PGE(2) induces moderate expansion of the bovine cumulus in vitro.
Calder, M D
Caveney, A N
Westhusin, M E
Watson, A J
Article
2001-07-01T07:00:00Z
Animals
Base Sequence
Cattle
Cyclooxygenase 2
Dinoprostone
Female
Isoenzymes
Molecular Sequence Data
Oocytes
Prostaglandin-Endoperoxide Synthases
RNA
Messenger
Receptors
Prostaglandin E
Reverse Transcriptase Polymerase Chain Reaction
Biology of reproduction
65
1
135
140
Obstetrics and Gynecology
<p>Expression of cyclooxygenase-2 (COX-2) and prostaglandin E(2) (PGE(2)) receptor 2 (EP2) are necessary for rodent cumulus expansion in vivo. Prostaglandin E(2) receptor 3 (EP3) has been detected in bovine preovulatory follicles and corpora lutea. The current experiments examined the effect of PGE(2) on bovine cumulus expansion in vitro and expression of COX-2, EP1, EP2, EP3, and EP4 mRNAs in bovine cumulus-oocyte complexes (COCs) at 0, 6, 12, 18, and 24 h time points during maturation in vitro. Concentrations of PGE(2) above 50 ng/ml resulted in moderate cumulus expansion of bovine COCs, but expansion did not occur in the absence of serum. COX-2 mRNA expression increased in bovine COCs at 6 h and 12 h of maturation, then decreased. EP2 mRNA was detectable by reverse transcription-polymerase chain reaction at all time points. EP3 mRNA expression increased in COCs from 0 to 6 h and remained at this higher level through the culture period. Very low levels of EP4 mRNA expression were detectable, but EP1 was not detected in bovine COCs. Because EP receptor mRNAs and COX-2 mRNA are expressed in bovine COCs, there exists the potential for a prostaglandin autocrine/paracrine regulatory pathway during oocyte maturation. Differential expression of the EP3 mRNA among varying COC classes indicates that this gene product may be a useful marker of oocyte competence. Although the PGE(2) pathway is involved in cumulus expansion, serum factors are required to mediate PGE(2)-induced expansion.</p>
https://ir.lib.uwo.ca/obsgynpub/53
oai:ir.lib.uwo.ca:obsgynpub-1058
2016-09-26T16:58:04Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
11690528
Sensitivity of bovine blastocyst gene expression patterns to culture environments assessed by differential display RT-PCR.
Natale, D R
De Sousa, P A
Westhusin, M E
Watson, A J
Article
2001-11-01T08:00:00Z
Animals
Blastocyst
Cattle
Cells
Cultured
Cloning
Molecular
Culture Media
DNA
Complementary
Embryonic Development
Embryonic and Fetal Development
Female
Gene Expression
Gene Expression Regulation
Developmental
Gestational Age
Mice
Pregnancy
RNA
Messenger
Reverse Transcriptase Polymerase Chain Reaction
Zygote
Reproduction (Cambridge, England)
122
5
687
693
Obstetrics and Gynecology
<p>The use of culture media to support the development of preimplantation embryos to the blastocyst stage is often associated with detrimental effects on normal development. These effects have been uncovered largely by investigating the phenotypic abnormalities displayed by fetuses and newborns derived from cultured preimplantation embryos. Research to understand the impact of culture on the embryonic developmental programme has focused on embryo metabolism, gene expression and genomic imprinting. We have used differential display RT-PCR to examine culture influences on global transcript pools in bovine embryos. Others have examined culture influences on candidate "marker genes" in cultured murine, ovine and bovine embryos. These studies have demonstrated that culture conditions influence the amount of marker gene transcripts and downregulate or induce the expression of novel genes during early development. Optimized defined culture media maintain embryonic gene expression patterns closely resembling those displayed by embryos derived in vivo. Preimplantation mammalian embryos display an impressive capacity to respond to the pressures that suboptimal culture environments place upon them. However, this plasticity operates within a defined range of tolerances. Continued research using molecular techniques will lead to increased understanding of developmental mechanisms causing culture-related phenotypic abnormalities in post-implantation embryos.</p>
https://ir.lib.uwo.ca/obsgynpub/57
oai:ir.lib.uwo.ca:obsgynpub-1061
2016-08-29T18:49:38Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
11333210
Regulation of blastocyst formation.
Watson, A J
Barcroft, L C
Article
2001-05-01T07:00:00Z
Animals
Blastocyst
Cell Adhesion
Cell Polarity
Embryo
Mammalian
Gene Expression Regulation
Developmental
Humans
Sodium-Potassium-Exchanging ATPase
Frontiers in bioscience : a journal and virtual library
6
708
730
Obstetrics and Gynecology
<p>Preimplantation or pre-attachment development encompasses the "free"-living period of mammalian embryogenesis, which directs development of the zygote through to the blastocyst stage. Blastocyst formation is essential for implantation, establishment of pregnancy and is a principal determinant of embryo quality prior to embryo transfer. Cavitation (blastocyst formation) is driven by the expression of specific sets of gene products that direct the acquisition of cell polarity within the trophectoderm, which is both the first epithelium of development and the outer cell layer encircling the inner cell mass of the blastocyst. Critical gene families controlling these events include: the E-cadherin-catenin cell adhesion family, the tight junction gene family, the Na/K-ATPase gene family and perhaps the aquaporin gene family. This review will update the roles of each of these gene families in trophectoderm differentiation and blastocyst formation. The current principal hypothesis under investigation is that blastocyst formation is mediated by a trans-trophectoderm ion gradient(s) established, in part, by Na/K-ATPase, which drives the movement of water through aquaporins (AQPs) across the epithelium into the extracellular space of the blastocyst to form the fluid-filled blastocoel. The trophectoderm tight junctional permeability seal regulates the leakage of blastocoel fluid, and also assists in the maintenance of a polarized Na/K-ATPase distribution to the basolateral plasma membrane domain of the mural trophectoderm. The cell-to-cell adhesion provided by the E-cadherin-catenin gene families is required for the establishment of the tight junction seal and the maintenance of the polarized Na/K-ATPase distribution. Blastocyst formation is therefore directly linked with trophectoderm cell differentiation, which arises through fundamental cell biological processes that are associated with the establishment of cell polarity.</p>
https://ir.lib.uwo.ca/obsgynpub/58
oai:ir.lib.uwo.ca:obsgynpub-1064
2016-10-19T14:41:18Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
10911395
Genetic reprogramming of lactate dehydrogenase, citrate synthase, and phosphofructokinase mRNA in bovine nuclear transfer embryos produced using bovine fibroblast cell nuclei.
Winger, Q A
Hill, J R
Shin, T
Watson, A J
Kraemer, D C
Westhusin, M E
Article
2000-08-01T07:00:00Z
Animals
Blastocyst
Cattle
Cell Line
Cells
Cultured
Citrate (si)-Synthase
Clone Cells
DNA Primers
Embryo Transfer
Female
Fibroblasts
L-Lactate Dehydrogenase
Oocytes
Phosphofructokinase-1
RNA
Messenger
Reverse Transcriptase Polymerase Chain Reaction
Molecular reproduction and development
56
4
458
464
Obstetrics and Gynecology
<p>Adult animal cloning has progressed to allow the production of offspring cloned from adult cells, however many cloned calves die prenatally or shortly after birth. This study examined the expression of three important metabolic enzymes, lactate dehydrogenase (LDH), citrate synthase, and phosphofructokinase (PFK), to determine if their detection in nuclear transfer (NT) embryos mimics that determined for in vitro produced embryos. A day 40 nuclear transfer produced fetus derived from an adult cell line was collected and fetal fibroblast cultures were established and maintained. Reconstructed NT embryos were then produced from this cell line, and RT-PCR was used to evaluate mRNA reprogramming. All three mRNAs encoding these enzymes were detected in the regenerated fetal fibroblast cell line. Detection patterns were first determined for IVF produced embryos (1-cell, 2-cell, 6-8 cell, morula, and blastocyst stages) to compare with their detection in NT embryos. PFK has three subunits: PFK-L, PFK-M, and PFK-P. PFK-L and PFK-P were not detected in bovine oocytes. PFK subunits were not detected in 6-8 cell embryos but were detected in blastocysts. Results from NT embryo RT-PCR demonstrated that PFK was not detected in 8-cell NT embryos but was detected in NT blastocysts indicating that proper nuclear reprogramming had occurred. Citrate synthase was detected in oocytes and throughout development to the blastocyst stage in both bovine IVF and NT embryos. LDH-A and LDH-B were detected in bovine oocytes and in all stages of IVF and NT embryos examined up to the blastocyst stage. A third subunit, LDH-C was not detected at the blastocyst stage in IVF or NT embryos but was detected in all earlier stages and in mature oocytes. In addition, LDH-C mRNA was detected in gonad isolated from the NT and an in vivo produced control fetus. These results indicate that the three metabolic enzymes maintain normal expression patterns and therefore must be properly reprogrammed following nuclear transfer.</p>
https://ir.lib.uwo.ca/obsgynpub/63
oai:ir.lib.uwo.ca:obsgynpub-1065
2016-08-29T19:02:44Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
10837135
Differential involvement of Na(+),K(+)-ATPase isozymes in preimplantation development of the mouse.
MacPhee, D J
Jones, D H
Barr, K J
Betts, D H
Watson, A J
Kidder, G M
Article
2000-06-15T07:00:00Z
Animals
Base Sequence
Blastocyst
Female
Gene Expression Regulation
Developmental
Isoenzymes
Macromolecular Substances
Mice
Molecular Sequence Data
RNA
Messenger
Reverse Transcriptase Polymerase Chain Reaction
Sequence Alignment
Sequence Homology
Nucleic Acid
Sodium-Potassium-Exchanging ATPase
Transcription
Genetic
Developmental biology
222
2
486
498
Obstetrics and Gynecology
<p>Na(+),K(+)-ATPase plays an essential role in mammalian blastocoel formation (cavitation) by driving trans-epithelial sodium transport. Previously, the alpha1 and beta1 subunit isoforms of this enzyme were identified in preimplantation mouse embryos and were assumed to be responsible for this function. Here we show that mRNAs encoding an additional alpha subunit isoform (alpha3) and the remaining two beta subunit isoforms are also present in preimplantation embryos. Whereas alpha3 mRNA accumulates between the four-cell and the blastocyst stages and thus results from embryonic transcription, the same could not be demonstrated for beta2 and beta3 mRNAs. Immunoblot analyses confirmed that these subunits are present in cavitating embryos. Using confocal immunofluorescence microscopy we found that alpha1 and beta1 subunits are concentrated in the basolateral membranes of the trophectoderm while being equally distributed in plasma membranes of the inner cell mass. In contrast, alpha3, beta2, and beta3 subunits were not detected in plasma membranes. Our current assessment, therefore, is that as many as six isozymes of Na(+),K(+)-ATPase could be involved in preimplantation development although it is primarily the alpha1beta1 isozyme that is responsible for blastocoel formation. Our findings imply that the regulation of sodium transport within the preimplantation mouse embryo is more complex than had been appreciated.</p>
https://ir.lib.uwo.ca/obsgynpub/64
oai:ir.lib.uwo.ca:obsgynpub-1060
2016-10-19T14:26:02Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
11414495
Amino acid concentrations in fluids from the bovine oviduct and uterus and in KSOM-based culture media.
Elhassan, Y M
Wu, G
Leanez, A C
Tasca, R J
Watson, A J
Westhusin, M E
Article
2001-06-01T07:00:00Z
Amino Acids
Animals
Cattle
Chromatography
High Pressure Liquid
Culture Media
Female
Oviducts
Uterus
Theriogenology
55
9
1907
1918
Obstetrics and Gynecology
<p>Amino acids in bovine oviductal and uterine fluids were measured and compared with those in modified simplex optimized medium (KSOM) supplemented with either fetal calf serum or Minimum Essential Medium amino acids in addition to bovine serum albumin, fetal calf serum or polyvinyl alcohol. Concentrations of cysteine, threonine, tryptophan, alanine, aspartate, glycine, glutamate, proline, beta-alanine, and citrulline were higher in oviductal fluids than in KSOM-based culture media. Nonessential and essential amino acids were present in ratios of 5:1 and 2:1 in oviductal and uterine fluids, respectively. Concentrations of alanine (3.7 mM), glycine (14.1 mM) and glutamate (5.5 mM) were high in oviductal fluids, comprising 73% of the free amino acid pool. Of the amino acids measured in uterine fluids, alanine (3.1 mM), glycine (12.0 mM), glutamate (4.2 mM), and serine (2.7 mM) were highest in concentration, and the first three comprised 43% of the free amino acid pool. In conclusion, amino acid concentrations in the bovine reproductive tract were substantially higher than those in embryo culture media. Certain amino acids, particularly alanine, glutamate, glycine and taurine, are present in strikingly high concentrations in both oviductal and uterine fluids, suggesting that they might play important roles in early embryo development. The particular pattern of amino acid concentrations may be an important factor to be considered for the improvement of embryo culture media.</p>
https://ir.lib.uwo.ca/obsgynpub/60
oai:ir.lib.uwo.ca:obsgynpub-1062
2016-10-19T14:29:53Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
11303903
Expression of PTHrP and PTHR (PTH/PTHrP-r) mRNAs and polypeptides in bovine ovary and stimulation of bovine blastocyst development in vitro following PTHrP treatment during oocyte maturation.
Watson, P H
Westhusin, M E
Watson, A J
Article
2001-03-01T08:00:00Z
Animals
Blastocyst
Cattle
Dose-Response Relationship
Drug
Female
Fertilization in Vitro
Immunohistochemistry
In Situ Hybridization
In Vitro Techniques
Ovary
Parathyroid Hormone
Parathyroid Hormone-Related Protein
Proteins
RNA
Messenger
Receptor
Parathyroid Hormone
Type 1
Receptors
Parathyroid Hormone
Anatomy and embryology
203
3
175
184
Obstetrics and Gynecology
<p>Parathyroid hormone related protein (PTHrP) and its receptor have well-established roles in the development and regulation of many tissues, including bone and mammary gland. The objectives of this study were: (1) to characterize the distribution of mRNAs encoding parathyroid hormone (PTH)-related protein (PTHrP) and receptor (PTHR) in bovine ovary; (2) to characterize the distribution of PTHrP and PTHR polypeptides in bovine ovary; (3) to examine the influences of PTHrP (1-141) treatment during bovine oocyte maturation in vitro on blastocyst development. mRNAs encoding PTHrP and PTHR were detected by in situ hybridization methods in oocytes, and granulosa cells in all follicles from primordial to large antral. PTHrP and PTHR polypeptides displayed distinct distribution patterns with PTHrP polypeptides primarily confined to oocytes from primordial to large antral follicles. PTHrP polypeptides were detectable but at a reduced level in ovarian stroma and in granulosa and thecal layers. PTHR polypeptides were detected in oocytes of all follicular stages but were predominantly found in ovarian stroma, granulosa and theca follicular layers. Supplementation of serum-free cSOFMaa oocyte maturation medium with PTHrP (1-141) resulted in a concentration-dependent increase in development to the blastocyst stage in vitro. The results suggest that granulosa cells may be a primary site of PTHrP production and release. Oocytes from all follicular stages stained strongly for PTHrP polypeptides and PTHrP enhanced development to the blastocyst stage in vitro.</p>
https://ir.lib.uwo.ca/obsgynpub/61
oai:ir.lib.uwo.ca:obsgynpub-1063
2016-10-19T14:34:34Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
11066060
mRNAs encoding aquaporins are present during murine preimplantation development.
Offenberg, H
Barcroft, L C
Caveney, A
Viuff, D
Thomsen, P D
Watson, A J
Article
2000-12-01T08:00:00Z
Animals
Aquaporin 6
Aquaporins
Base Sequence
DNA
Complementary
Embryonic Development
Embryonic and Fetal Development
Female
Male
Mice
Mice
Inbred C57BL
Molecular Sequence Data
Pregnancy
RNA
Messenger
Sequence Analysis
DNA
Molecular reproduction and development
57
4
323
330
Obstetrics and Gynecology
<p>The present study was conducted to investigate the mechanisms underlying fluid movement across the trophectoderm during blastocyst formation by determining whether aquaporins (AQPs) are expressed during early mammalian development. AQPs belong to a family of major intrinsic membrane proteins and function as molecular water channels that allow water to flow rapidly across plasma membranes in the direction of osmotic gradients. Ten different AQPs have been identified to date. Murine preimplantation stage embryos were flushed from the oviducts and uteri of superovulated CD1 mice. Reverse transcription-polymerase chain reaction (RT-PCR) methods employing primer sets designed to amplify conserved sequences of AQPs (1-9) were applied to murine embryo cDNA samples. PCR reactions were conducted for up to 40 cycles involving denaturation of DNA hybrids at 95 degrees C, primer annealing at 52-60 degrees C and extension at 72 degrees C. PCR products were separated on 2% agarose gels and were stained with ethidium bromide. AQP PCR product identity was confirmed by sequence analysis. mRNAs encoding AQPs 1, 3, 5, 6, 7, and 9 were detected in murine embryos from the one-cell stage up to the blastocyst stage. AQP 8 mRNAs were not detected in early cleavage stages but were present in morula and blastocyst stage embryos. The results were confirmed in experimental replicates applied to separate embryo pools of each embryo stage. These results demonstrate that transcripts encoding seven AQP gene products are detectable during murine preimplantation development. These findings predict that AQPs may function as conduits for trophectoderm fluid transport during blastocyst formation.</p>
https://ir.lib.uwo.ca/obsgynpub/62
oai:ir.lib.uwo.ca:obsgynpub-1067
2016-10-20T16:17:03Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
10732125
Temporal patterns of embryonic gene expression and their dependence on oogenetic factors.
De Sousa, P A
Caveney, A
Westhusin, M E
Watson, A J
Article
1998-01-01T08:00:00Z
Animals
Blastocyst
Cattle
Embryonic and Fetal Development
Female
Gene Expression Regulation
Developmental
Genomic Imprinting
Oogenesis
Transcriptional Activation
Zygote
Theriogenology
49
1
115
128
Obstetrics and Gynecology
<p>Successful development of a fertilized egg beyond early cleavage divisions requires the de novo initiation and subsequent regulation of embryonic transcription. The egg provides the specialized environment within which the newly formed zygotic nucleus initiates its developmental program and as a result plays an obligatory role in its regulation. Although the precise timing of the onset of embryonic transcription in mammals varies during early cleavage divisions, several common elements exist. In the present essay we review the current literature on the timing and control of embryonic gene expression in mammals, and discuss recent findings from our laboratory on gene expression patterns in bovine embryos and their relation to other species, and zygotic gene activation (ZGA). Lastly, we discuss the putative role of maternally inherited factors in conferring developmental competence to the blastocyst stage, and a method to identify such factors present in oocytes as mRNA.</p>
https://ir.lib.uwo.ca/obsgynpub/68
oai:ir.lib.uwo.ca:obsgynpub-1066
2016-08-29T19:03:47Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
10793627
Effects of superovulated heifer diet type and quantity on relative mRNA abundances and pyruvate metabolism in recovered embryos.
Wrenzycki, C
De Sousa, P
Overström, E W
Duby, R T
Herrmann, D
Watson, A J
Niemann, H
O'Callaghan, D
Boland, M P
Article
2000-01-01T08:00:00Z
Analysis of Variance
Animals
Cattle
Diet
Embryo
Mammalian
Female
Gestational Age
Globins
Pregnancy
Pyruvic Acid
RNA
Messenger
Reverse Transcriptase Polymerase Chain Reaction
Sodium-Potassium-Exchanging ATPase
Superovulation
Superoxide Dismutase
Journal of reproduction and fertility
118
1
69
78
Obstetrics and Gynecology
<p>This study investigated the effects of quantity and type of diet fed to superovulated donor heifers on molecular and metabolic indices of embryonic development. These effects included the relative abundances of mRNAs for the alpha 1 subunit of Na/K-ATPase and the antioxidant enzyme Cu/Zn-SOD, as well as pyruvate utilization in bovine morulae and blastocysts developed in vivo. Heifers were fed a daily ration of either grass silage and a citrus-beet pulp-based concentrate or grass silage and a barley-based concentrate for 116 days, both at 3 kg per day or ad libitum. In embryos derived from heifers fed the pulp-based diets, the relative abundances of the transcripts were not affected by either day of collection or quantity of diet. In embryos derived from heifers fed the barley-based diets, the relative abundances of the Na/K-ATPase transcripts were also not changed by these main effects, while the relative abundances of the Cu/Zn-SOD transcripts were affected by day of collection and by the quantity of diet. Pyruvate metabolism was affected by day of collection, and was significantly increased in day 8 embryos compared with day 7 and day 6 embryos. Diet quantity did not affect pyruvate utilization, whereas diet type did increase pyruvate metabolism in the barley group when compared with the pulp group. The results of this study show for the first time that molecular and metabolic variations may exist in embryos derived in vivo and developed in donor heifers on nutritional regimens differing in type and quantity. Differences in embryos collected on different developmental days may be attributed to varying cell numbers. Alterations in the relative abundances of the Cu/Zn-SOD transcripts and pyruvate metabolism caused by the quantity of diet fed to the donor animal were likely to have been due to alterations in metabolic end products that accumulate in reproductive tract fluids, whereas differences in embryonic metabolism caused by type of diet are related to the composition of the diet. These findings characterize embryos produced in vivo at the molecular level, indicating that the molecular markers used in the present study can differentiate between populations of embryos produced under different nutritional regimens and determine conditions conductive to the production of good quality embryos.</p>
https://ir.lib.uwo.ca/obsgynpub/65
oai:ir.lib.uwo.ca:obsgynpub-1070
2016-08-29T19:10:52Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
10642573
Impact of bovine oocyte maturation media on oocyte transcript levels, blastocyst development, cell number, and apoptosis.
Watson, A J
De Sousa, P
Caveney, A
Barcroft, L C
Natale, D
Urquhart, J
Westhusin, M E
Article
2000-02-01T08:00:00Z
Amino Acids
Animals
Apoptosis
Blastocyst
Cattle
Cell Count
Coloring Agents
Culture Media
Serum-Free
DNA
Embryonic and Fetal Development
Epidermal Growth Factor
Female
Fertilization in Vitro
Genetic Markers
Image Processing
Computer-Assisted
In Situ Nick-End Labeling
In Vitro Techniques
Microscopy
Video
Oocytes
Propidium
RNA
Messenger
Reverse Transcriptase Polymerase Chain Reaction
Biology of reproduction
62
2
355
364
Obstetrics and Gynecology
<p>The objectives were 1) to investigate the effects of oocyte maturation in serum-free and amino acid-supplemented defined media on oocyte transcript levels, blastocyst cell number, and apoptosis; 2) to investigate the influence of oocyte maturation culture atmosphere on blastocyst development, total cell number, and apoptosis; and 3) to examine the influence of epidermal growth factor (EGF) during oocyte maturation on blastocyst cell number and apoptosis. The results demonstrate that blastocysts derived from in vitro maturation, fertilization, and embryo culture protocols undergo apoptosis but that apoptotic levels are not greatly influenced by the oocyte maturation environment. Amino acid supplementation of oocyte maturation media was associated with enhanced developmental frequencies, increased blastocyst cell number, and elevated oocyte maternal mRNA levels. Oocyte maturation with supplemented synthetic oviduct fluid medium (cSOFMaa) resulted in blastocyst cell numbers comparable to those observed with Tissue Culture Medium 199 + newborn calf serum. Blastocyst development was reduced following oocyte maturation under a 5% CO(2), 7% O(2), 88% N(2) culture atmosphere. EGF supplementation of oocyte maturation medium resulted in a concentration-dependent increase in blastocyst development but did not influence blastocyst total cell number or apoptosis. Our findings indicate that cSOFMaa medium is an effective base medium for bovine oocyte maturation.</p>
https://ir.lib.uwo.ca/obsgynpub/67
oai:ir.lib.uwo.ca:obsgynpub-1068
2016-10-20T15:37:00Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
10729067
Gene expression regulating blastocyst formation.
Watson, A J
Westhusin, M E
De Sousa, P A
Betts, D H
Barcroft, L C
Article
1999-01-01T08:00:00Z
Animals
Blastocyst
Cattle
Cell Adhesion
Gene Expression Regulation
Developmental
Oogenesis
Theriogenology
51
1
117
133
Obstetrics and Gynecology
<p>Development of embryos to the blastocyst stage is a critical event in the early lives of all eutherian mammalian species. Blastocyst formation is essential for implantation and is the principal morphological determinant of embryo quality prior to embryo transfer. The physiological events and roles of specific gene families that regulate blastocyst formation are subjects of intense research Recent findings have demonstrated that bovine embryos express multiple members of the Na/K-ATPase ion transporter gene family. Two members of this family have been co-localized to bovine trophectoderm, but each becomes largely confined to opposing cell membrane margins. Bovine blastocysts display a greater sensitivity to ouabain (potent inhibitor of the Na/K-ATPase) than murine blastocysts, and enzyme activity (ouabain sensitive 86Rb+ uptake) undergoes a 9-fold increase from the bovine morula to the blastocyst stage. Disruption of Na/K-ATPase gene expression by antisense oligodeoxynucleotide inhibition abolishes blastocyst formation. These results have implicated the Na/K-ATPase as a key regulator of bovine blastocyst formation and have provided insights necessary for the production of healthy bovine embryos by the application of in vitro maturation, in vitro fertilization and in vitro culture methods.</p>
https://ir.lib.uwo.ca/obsgynpub/69
oai:ir.lib.uwo.ca:obsgynpub-1069
2016-08-29T19:09:28Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
10692863
IGF paracrine and autocrine interactions between conceptus and oviduct.
Watson, A J
Westhusin, M E
Winger, Q A
Article
1999-01-01T08:00:00Z
Animals
Autocrine Communication
Cattle
Embryo
Mammalian
Fallopian Tubes
Female
Insulin-Like Growth Factor Binding Proteins
Insulin-Like Growth Factor I
Insulin-Like Growth Factor II
Paracrine Communication
Pregnancy
Receptors
Somatomedin
Ruminants
Sheep
Somatomedins
Journal of reproduction and fertility. Supplement
54
303
315
Obstetrics and Gynecology
<p>Development in vitro is influenced by embryo density, serum, somatic cell co-culture and the production of 'embryotrophic' paracrine and autocrine factors. Research in our laboratory has focussed principally on the insulin-like growth factor (IGF) family. We have demonstrated that pre-attachment bovine and ovine embryos express mRNAs encoding a number of growth factor ligand and receptor genes including all members of the IGF ligand and receptor family throughout this developmental interval. In addition, early embryos express mRNAs encoding IGF-binding proteins (IGFBPs) 2-5 from the one-cell to the blastocyst stage and IGFBP5 mRNA at the blastocyst stage. Cultured bovine blastocysts release up to 35 pg per embryo in 24 h, whereas release of IGF-I was below detectable values. Analysis extended to bovine oviductal cultures has also demonstrated that mRNAs encoding these IGF family members are present throughout an 8 day culture period. Transcripts encoding IGFBPs 2-6 were also present. Release of both IGFs was recorded over an 8 day culture period. IGF-II release was significantly greater than that observed for IGF-I. Therefore, the IGFs are present throughout the maternal environment during early embryo development. The oocyte, within the follicle, is held in an environment high in IGFs and IGFBPs. The zygote, after fertilization, is maintained in an IGF-rich environment while free-living in the oviduct and the uterus. This review is focused on the IGF family and IGFBPs and their roles in enhancing development up to the blastocyst stage.</p>
https://ir.lib.uwo.ca/obsgynpub/66
oai:ir.lib.uwo.ca:obsgynpub-1071
2016-10-20T16:51:04Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
10618654
Assessment by differential display-RT-PCR of mRNA transcript transitions and alpha-amanitin sensitivity during bovine preattachment development.
Natale, D R
Kidder, G M
Westhusin, M E
Watson, A J
Article
2000-02-01T08:00:00Z
Amanitins
Animals
Blastocyst
Cattle
Embryo
Mammalian
Embryonic Development
Enzyme Inhibitors
Female
Fertilization in Vitro
Gene Expression Regulation
Developmental
Oocytes
Pregnancy
RNA Polymerase II
RNA
Messenger
Reverse Transcriptase Polymerase Chain Reaction
Uridine
Molecular reproduction and development
55
2
152
163
Obstetrics and Gynecology
<p>The objectives of this study were to compare patterns of mRNA expression, investigate the onset of transcription, and isolate stage-specific and alpha-amanitin-sensitive mRNAs during early bovine development by differential-display-reverse transcription-polymerase chain reaction (DD-RT-PCR). Embryos representing a preattachment developmental series from the 1-cell to the expanded/hatched blastocyst stage were cultured in synthetic oviduct fluid medium + citrate and amino acids (cSOFMaa) with and without alpha-amanitin (100 microg/mL) for 4 and 12 hr. mRNA profiles were displayed by DD-RT-PCR using 5' primers A and N. Total conserved cDNA banding patterns varied according to embryo stage with cDNA band numbers declining during early cleavage stages compared to oocyte values and then increasing in total number from the 6-8-cell stage through to the blastocyst stage. A cDNA banding pattern was established at the 8-16-cell stage that was largely unchanged through to the blastocyst stage. These findings with respect to cDNA banding patterns were conserved between oligo primer sets and experimental replicates. alpha-Amanitin sensitivity was first detected at the 2-5-cell stage but became predominant following the 6-8-cell stage of development to eventually affect the appearance of up to 40% of all cDNA bands by the blastocyst stage. A 12 hr alpha-amanitin treatment was required to effectively block (3)H-uridine incorporation into mRNA in blastocyst stage embryos. Several stage-specific and alpha-amanitin-sensitive cDNAs were isolated and they will be a focus for future studies. In conclusion, DD-RT-PCR is an effective tool for contrasting gene expression patterns and isolating uncharacterized mRNA transcripts during bovine early development. Mol. Reprod. Dev. 55:152-163, 2000.</p>
https://ir.lib.uwo.ca/obsgynpub/70
oai:ir.lib.uwo.ca:obsgynpub-1072
2016-10-20T16:55:00Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
10602269
Characterization of a bovine cDNA encoding citrate synthase, and presence of citrate synthase mRNA during bovine pre-attachment development.
Winger, Q A
Hill, J R
Watson, A J
Westhusin, M E
Article
2000-01-01T08:00:00Z
Amino Acid Sequence
Animals
Base Sequence
Blastocyst
Cattle
Citrate (si)-Synthase
DNA
Complementary
Embryo
Mammalian
Gene Expression
Humans
Molecular Sequence Data
RNA
Messenger
Reverse Transcriptase Polymerase Chain Reaction
Sequence Analysis
DNA
Sequence Homology
Amino Acid
Sequence Homology
Nucleic Acid
Swine
Molecular reproduction and development
55
1
14
19
Obstetrics and Gynecology
<p>Citrate synthase is a key regulatory metabolic enzyme that catalyzes the first step in the tricarboxylic acid (TCA) cycle, the synthesis of citrate from acetyl coenzyme A and oxaloacetate. Aerobic metabolism via the TCA cycle is high in bovine embryos at the 4-cell stage then decreases until the compact morula stage before increasing at the expanded blastocyst stage. This study characterizes the presence of citrate synthase mRNA in bovine pre-attachment embryos to determine if a variation in mRNA transcript expression patterns is associated with previous reports of the patterns of TCA cycle activity. The reverse transcription-polymerase chain reaction (RT-PCR) method was used to detect citrate synthase mRNA from the 1-cell to blastocyst stage of bovine embryo development, and in embryos cultured under either an atmosphere of 5% CO(2) in air or 5% CO(2)/5% O(2)/90%N(2). The nucleotide sequence encoding citrate synthase was determined from bovine heart cDNA by the rapid amplification of cDNA ends (RACE) technique. This 1455-bp nucleotide fragment contained an open reading frame that encoded a deduced protein of 466 amino acids. The bovine nucleotide sequence was 92.1% and 93.8% identical to the human and porcine coding sequence, respectively. The amino acid sequence predicted from the bovine sequence is 95.1% identical to the human sequence and 96.3% identical to the porcine sequence. The porcine sequence contains a stop codon that results in a peptide truncated by 2 amino acids. The detection of citrate synthase transcripts from the 1-cell to blastocyst stage demonstrates that the decrease in TCA cycle activity observed following the 4-cell stage is not associated with an absence of citrate synthase mRNA.</p>
https://ir.lib.uwo.ca/obsgynpub/71
oai:ir.lib.uwo.ca:obsgynpub-1073
2016-08-29T19:38:44Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
10070362
Trophectoderm differentiation in the bovine embryo: characterization of a polarized epithelium.
Barcroft, L C
Hay-Schmidt, A
Caveney, A
Gilfoyle, E
Overstrom, E W
Hyttel, P
Watson, A J
Article
1998-11-01T08:00:00Z
Animals
Base Sequence
Blastocyst
Cadherins
Cattle
Cell Differentiation
Cytoskeletal Proteins
Embryonic and Fetal Development
Epithelial Cells
Humans
Immunohistochemistry
Membrane Proteins
Mice
Microscopy
Fluorescence
Molecular Sequence Data
Phosphoproteins
RNA
Messenger
Reverse Transcriptase Polymerase Chain Reaction
Sequence Homology
Nucleic Acid
Trans-Activators
Zonula Occludens-1 Protein
beta Catenin
Journal of reproduction and fertility
114
2
327
339
Obstetrics and Gynecology
<p>Blastocytst formation is dependent on the differentiation of a transporting epithelium, the trophectoderm, which is coordinated by the embryonic expression and cell adhesive properties of E-cadherin. The trophectoderm shares differentiative characteristics with all epithelial tissues, including E-cadherin-mediated cell adhesion, tight junction formation, and polarized distribution of intramembrane proteins, including the Na-K ATPase. The present study was conducted to characterize the mRNA expression and distribution of polypeptides encoding E-cadherin, beta-catenin, and the tight junction associated protein, zonula occludens protein 1, in pre-attachment bovine embryos, in vitro. Immunocytochemistry and gene specific reverse transcription--polymerase chain reaction methods were used. Transcripts for E-cadherin and beta-catenin were detected in embryos of all stages throughout pre-attachment development. Immunocytochemistry revealed E-cadherin and beta-catenin polypeptides evenly distributed around the cell margins of one-cell zygotes and cleavage stage embryos. In the morula, detection of these proteins diminished in the free apical surface of outer blastomeres. E-cadherin and beta-catenin became restricted to the basolateral membranes of trophectoderm cells of the blastocyst, while maintaining apolar distributions in the inner cell mass. Zonula occludens protein 1 immunoreactivity was undetectable until the morula stage and first appeared as punctate points between the outer cells. In the blastocyst, zonula occludens protein 1 was localized as a continuous ring at the apical points of trophectoderm cell contact and was undetectable in the inner cell mass. These results illustrate that the gene products encoding E-cadherin, beta-catenin and zonula occludens protein 1 are expressed and maintain cellular distribution patterns consistent with their predicted roles in mediating trophectoderm differentiation in in vitro produced bovine embryos.</p>
https://ir.lib.uwo.ca/obsgynpub/72
oai:ir.lib.uwo.ca:obsgynpub-1075
2016-08-29T19:41:10Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
9746750
Transient expression of a translation initiation factor is conservatively associated with embryonic gene activation in murine and bovine embryos.
De Sousa, P A
Watson, A J
Schultz, R M
Article
1998-10-01T07:00:00Z
Amanitins
Animals
Blotting
Northern
Cattle
Conserved Sequence
DNA
DNA Fragmentation
Embryo
Mammalian
Embryonic Development
Eukaryotic Initiation Factor-1
Eukaryotic Initiation Factor-5
Female
Fertilization in Vitro
Gene Expression Regulation
Gene Expression Regulation
Developmental
Mice
Nucleic Acid Synthesis Inhibitors
Peptide Initiation Factors
Pregnancy
RNA
Messenger
Reverse Transcriptase Polymerase Chain Reaction
Transcriptional Activation
Biology of reproduction
59
4
969
977
Obstetrics and Gynecology
<p>In the present study the abundance of mRNAs for eukaryotic translation initiation factors eIF-1A (formerly known as eIF-4C), -2alpha, -4A, -4E, and -5 was examined in in vivo-derived mouse embryos throughout preimplantation development using a semiquantitative reverse transcription-polymerase chain reaction assay. Although the mRNA profile for each gene is unique, only mRNA for eIF-1A transiently increases during embryonic gene activation (EGA) at the 2-cell stage, and this was confirmed by an independent hybridization-based assay. In in vitro-developed bovine embryos, mRNA for eIF-1A was transiently detected at the 8-cell stage, when the major activation of the genome occurs in this species. As in the mouse, detection in 8-cell bovine embryos was sensitive to the transcriptional inhibitor alpha-amanitin. It was also observed at the same time relative to cleavage in embryos cultured in defined medium under a reduced oxygen environment, and in medium supplemented with serum and somatic cells in 5% CO2 in air. Neither the chronology of early cleavage divisions nor the yield of bovine blastocysts differed in these culture media. Our results suggest that transient expression of eIF-1A in the mouse and cow is a conserved pattern of gene expression associated with EGA in mammals.</p>
https://ir.lib.uwo.ca/obsgynpub/76
oai:ir.lib.uwo.ca:obsgynpub-1074
2016-10-21T16:33:02Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
9795343
Effects of colony stimulating factor-1 on human extravillous trophoblast growth and invasion.
Hamilton, G S
Lysiak, J J
Watson, A J
Lala, P K
Article
1998-10-01T07:00:00Z
Analysis of Variance
Antibodies
Blotting
Northern
Cell Division
Collagenases
Female
Humans
Macrophage Colony-Stimulating Factor
Matrix Metalloproteinase 9
Microscopy
Fluorescence
Pregnancy
Pregnancy Trimester
First
RNA
Messenger
Receptor
Macrophage Colony-Stimulating Factor
Reverse Transcriptase Polymerase Chain Reaction
Stimulation
Chemical
Tissue Inhibitor of Metalloproteinase-1
Tissue Inhibitor of Metalloproteinase-2
Trophoblasts
The Journal of endocrinology
159
1
69
77
Obstetrics and Gynecology
<p>Colony stimulating factor (CSF)-1 has been localized in a variety of tissues and shown to influence proliferation and differentiation of numerous cell types. Messenger RNA and protein products of CSF-1 and its receptor (c-fms) have been identified in the human placenta and decidua. We examined whether CSF-1 and c-fms mRNA and protein are expressed by normal human first trimester invasive extravillous trophoblast (EVT) cells propagated in culture and whether CSF-1 influences proliferation and/or invasion of these cells. CSF-1 mRNA and protein expression was determined by RT-PCR and immunofluorescence microscopy. Proliferation was assessed by the cellular uptake of tritiated thymidine and invasion was evaluated by Matrigel invasion assay as well as Northern blot analysis of mRNA expression for invasion-associated enzymes and their inhibitors. Results revealed that normal invasive EVT cells in culture express both CSF-1 and c-fms mRNA and protein. Under serum-free conditions, exogenous CSF-1 greatly stimulated the proliferation of these cells. CSF-1 neutralizing and c-fms receptor blocking antibody (Ab) each abolished the growth stimulatory effects of CSF-1, indicating that CSF-1 and c-fms interaction was responsible for these effects. In fact, c-fms Ab alone reduced proliferation to below background levels. While exogenous CSF-1 failed to influence EVT cell invasiveness, Northern blot analysis of mRNA indicated a slight upregulation of the invasion-associated enzyme 72 kDa type IV collagenase as well as its natural inhibitor tissue inhibitor of metalloprotease (TIMP)-1, so that the balance between the two remained unaltered. These findings suggest that CSF-1 may represent an autocrine (and possibly paracrine) growth stimulatory factor for the invasive trophoblast cells in situ with no net effect on their invasiveness.</p>
https://ir.lib.uwo.ca/obsgynpub/75
oai:ir.lib.uwo.ca:obsgynpub-1076
2016-10-21T16:42:05Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
9712325
Oogenetic and zygotic gene expression directing early bovine embryogenesis: a review.
De Sousa, P A
Watson, A J
Schultz, G A
Bilodeau-Goeseels, S
Article
1998-09-01T07:00:00Z
Animals
Cattle
Cell Adhesion
Embryonic and Fetal Development
Gene Expression Regulation
Developmental
Growth Substances
Humans
Oogenesis
RNA
Small Nuclear
Sodium-Potassium-Exchanging ATPase
Tight Junctions
Zygote
Molecular reproduction and development
51
1
112
121
Obstetrics and Gynecology
https://ir.lib.uwo.ca/obsgynpub/77
oai:ir.lib.uwo.ca:obsgynpub-1077
2016-08-29T19:45:05Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
9578620
Na/K-ATPase-mediated 86Rb+ uptake and asymmetrical trophectoderm localization of alpha1 and alpha3 Na/K-ATPase isoforms during bovine preattachment development.
Betts, D H
Barcroft, L C
Watson, A J
Article
1998-05-01T07:00:00Z
Animals
Blastocyst
Brain
Cattle
Cells
Cultured
Culture Techniques
Cytochalasin D
Ectoderm
Embryo
Mammalian
Embryonic Development
Embryonic and Fetal Development
Enzyme Inhibitors
Epithelial Cells
Female
Isoenzymes
Kidney
Microscopy
Fluorescence
Ouabain
Pregnancy
Rubidium
Sodium-Potassium-Exchanging ATPase
Developmental biology
197
1
77
92
Obstetrics and Gynecology
<p>This study evaluated Na/K-ATPase alpha 1- and alpha 3-subunit isoform polypeptide expression and localization during bovine preattachment development. Na/K-ATPase cation transport activity from the one-cell to blastocyst stage was also determined by measuring ouabain-sensitive 86Rb+ uptake. Both alpha1- and alpha 3-subunit polypeptides were detected by immunofluorescence to encircle the entire cell margins of each blastomere of inseminated zygotes, cleavage stage embryos, and morulae. Immunofluorescent localization of alpha1-subunit polypeptide in bovine blastocysts revealed an alpha1 immunofluorescence signal confined to the basolateral membrane margins of the trophectoderm and encircling the cell periphery of each inner cell mass (ICM) cell. In contrast, alpha 3-subunit polypeptide immunofluorescence was localized primarily to the apical cell surfaces of the trophectoderm with a reduced signal present in basolateral trophectoderm regions. There was no apparent alpha 3-subunit signal in the ICM. Analysis of 86Rb+ transport in vitro demonstrated ouabain-sensitive activity throughout development from the one-cell to the six- to eight-cell stage of bovine development. 86Rb+ uptake by morulae (day 6 postinsemination) did not vary significantly from uptake detected in cleavage stage embryos; however, a significant increase was measured at the blastocyst stage (P < 0.05). Treatment of embryos with cytochalasin D (5 micrograms/ml) did not influence 86Rb+ uptake in cleavage stage embryos. Cytochalasin D treatment however was associated with a significant rise in ion transport in morulae and blastocysts (13.49 and 61.57 fmol/embryo/min, respectively) compared to untreated controls (2.65 and 22.83 fmol/embryo/min, respectively). Our results, for the first time, demonstrate that multiple Na/K-ATPase alpha-subunit isoforms are distributed throughout the first week of mammalian development and raise the possibility that multiple isozymes of the Na/K-ATPase contribute to blastocyst formation.</p>
https://ir.lib.uwo.ca/obsgynpub/78
oai:ir.lib.uwo.ca:obsgynpub-1078
2016-10-21T16:46:33Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
9444655
Analysis of variation in relative mRNA abundance for specific gene transcripts in single bovine oocytes and early embryos.
De Sousa, P A
Westhusin, M E
Watson, A J
Article
1998-02-01T08:00:00Z
Actins
Animals
Blastocyst
Cattle
Embryo
Mammalian
Genetic Variation
Globins
Oocytes
Polymerase Chain Reaction
RNA
Messenger
RNA-Directed DNA Polymerase
Rabbits
Sodium-Potassium-Exchanging ATPase
Transcription
Genetic
Molecular reproduction and development
49
2
119
130
Obstetrics and Gynecology
<p>Variation in the abundance of a specific gene transcript was assessed in single bovine oocytes and in vitro-derived blastocysts. Transcripts encoding the Na+,K(+)-ATPase alpha 1 subunit were detected by reverse-transcription polymerase chain reaction (RT-PCR) and quantified relative to an exogenously supplied rabbit alpha-globin mRNA using laser-induced fluorescence capillary electrophoresis (LIF-CE). The precision of this relative abundance (RA) calculation was predicted and shown to resolve 2-fold differences in transcript abundance between individual blastocysts and predicted in oocytes to resolve 3-fold differences. The RA of the alpha 1 subunit transcript differed by 2- to 3-fold among blastocysts, and 3- to 6-fold among oocytes. Comparison of a general population of oocytes with blastocysts revealed little overlap in RA values between the two groups, with a 8- to 14-fold increase in the mean RA for each group with development observed in two successive experiments (P < or = 0.05). In contrast, oocytes selected for their developmental competence on the basis of morphologic criteria exhibited only a 1.6- to 1.7-fold developmental increase when the assay was performed on cDNA generated from either embryo pools (n = 6 versus 6) or individuals (n = 7 versus 7), respectively. These results provide the first characterization of the degree of heterogeneity in the abundance of a specific mRNA transcript among individual mammalian oocytes and preimplantation embryos and demonstrate that transcript relative abundance can be correlated with bovine oocyte morphology.</p>
https://ir.lib.uwo.ca/obsgynpub/79
oai:ir.lib.uwo.ca:obsgynpub-1079
2016-08-29T19:53:13Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
9166693
Bovine oviductal and embryonic insulin-like growth factor binding proteins: possible regulators of "embryotrophic" insulin-like growth factor circuits.
Winger, Q A
de los Rios, P
Han, V K
Armstrong, D T
Hill, D J
Watson, A J
Article
1997-06-01T07:00:00Z
Animals
Base Sequence
Cattle
Culture Media
Conditioned
Culture Techniques
DNA Primers
Embryo
Mammalian
Fallopian Tubes
Female
Gene Expression
Insulin-Like Growth Factor Binding Proteins
Insulin-Like Growth Factor I
Insulin-Like Growth Factor II
RNA
Messenger
Receptor
IGF Type 1
Receptor
IGF Type 2
Biology of reproduction
56
6
1415
1423
Obstetrics and Gynecology
<p>Bovine oviductal monolayer and vesicle primary cultures express insulin-like growth factor (IGF)-I and -II mRNAs and polypeptides. Early bovine embryos also express IGF-I, IGF-II, IGF-I receptor, IGF-II receptor, and insulin receptor mRNAs. This study reports the expression of IGF binding protein (IGFBP) mRNAs and polypeptides in bovine oviduct primary cultures and IGFBP mRNAs in preattachment embryos. Release of immunoreactive IGF-I and IGF-II by oviduct cultures and bovine blastocysts was also determined. IGFBP-2, -3, -4, and -5 transcripts were observed in oviduct primary cultures throughout an 8-day interval. IGFBP-1 and -6 mRNAs were consistently not detected in the oviduct. Messenger RNAs encoding IGFBPs -2, -3, and -4 were detected throughout bovine preattachment development, while transcripts encoding IGFBP-5 were detected only in blastocysts. IGFBP-1 and -6 transcripts were not detected in early embryos. Ligand blot analysis with 125I-labeled IGF-II revealed the presence of four prominent polypeptide bands of approximate molecular masses 24, 31, and 36 kDa, and a broad band extending from 46 to 53 kDa, in conditioned media samples prepared from oviduct primary cultures. Western immunoblot analysis confirmed the identity of the 24-kDa, 31-kDa, and 36-kDa species as IGFBP-4, -5, and -2, respectively. Levels of the release of IGF-II from oviductal vesicle cultures were significantly greater than levels observed for monolayer cultures (p < 0.005). No significant difference in the levels of IGF-I release between monolayer and vesicle cultures was observed. Pools of 10 blastocysts released on average 36.2 +/- 3.9 pg of IGF-II per embryo, while the release of embryonic IGF-I was below the levels of detection for our assay. The results suggest that maternally derived IGF may be regulated by IGFBPs to support bovine preattachment development.</p>
https://ir.lib.uwo.ca/obsgynpub/80
oai:ir.lib.uwo.ca:obsgynpub-1080
2016-10-21T16:50:32Z
publication:obsgyn
publication:obsgynpub
publication:pmid
publication:faculties
9021743
Ouabain sensitivity and expression of Na/K-ATPase alpha- and beta-subunit isoform genes during bovine early development.
Betts, D H
MacPhee, D J
Kidder, G M
Watson, A J
Article
1997-02-01T08:00:00Z
Animals
Base Sequence
Blastomeres
Cattle
Culture Techniques
DNA
Complementary
Enzyme Inhibitors
Female
Isoenzymes
Male
Mice
Molecular Sequence Data
Ouabain
RNA
Messenger
Rats
Sequence Analysis
DNA
Sequence Homology
Nucleic Acid
Sodium-Potassium-Exchanging ATPase
Molecular reproduction and development
46
2
114
126
Obstetrics and Gynecology
<p>The fluid movements that arise during blastocyst formation (cavitation) are, at least in part, driven by the Na/K-ATPase. In this study, the reverse transcriptase-polymerase chain reaction (RT-PCR) was used to survey bovine pre-attachment embryos for transcripts encoding known isoforms of the Na/K-ATPase alpha- and beta-subunits, including isoforms not previously detected during the first week of mammalian development. Transcripts encoding the Na-K-ATPase alpha 1, alpha 2, alpha 3 and beta 2 isoforms were detected throughout bovine preattachment development. This is the first indication that alpha 2, alpha 3 and beta 2 mRNAs are expressed during this early developmental interval. As in the mouse, beta 1-subunit transcripts were not detected until the morula stage and were also present in blastocysts. Thus, in two mammalian species an increase in abundance of beta 1 isoform transcripts in the morula stage is coincident with the onset of cavitation. Transcripts encoding the recently characterized alpha 4 isoform were not detected. The sensitivity of bovine blastocysts to ouabain (a potent inhibitor of Na/K-ATPase) was determined by assessing the ability of bovine blastocysts to recover in ouabain supplemental culture medium following cytochalasin-induced blastocyst collapse. Re-expansion of bovine blastocysts was inhibited in all ouabain concentrations down to 10(-9) M. Mouse blastocysts, in contrast, were sensitive to ouabain at or above 10(-3)M. These results have established that transcripts encoding multiple isoforms of both the alpha and beta subunits of the Na/K-ATPase are expressed throughout early bovine development and that bovine blastocysts display a greater sensitivity to ouabain than murine blastocysts. Future analysis will determine the possible individual and collective roles of these isoforms during blastocyst formation.</p>
https://ir.lib.uwo.ca/obsgynpub/81
860630/simple-dublin-core/100//