Master of Science
Dr. Zia A. Khan
Infantile hemangiomas (IHs) are benign vascular neoplasms characterized by the differentiation of multipotential stem cells (hemSCs) into endothelial cells during the early proliferative phase, and later into adipocytes during spontaneous involution. Transforming growth factor-β (TGFβ) has been shown to be significantly elevated upon IH involution and this coincides with repression of a developmentally-regulated transcription factor T-box 2 (TBX2). These findings implicate both TGFβ and TBX2 in mediating hemSC differentiation during IH evolution. The aim of my study is to understand the role of TGFβ and TBX2 in hemSC differentiation. I performed immunofluorescence staining to localize TBX2 protein in sectioned IH tissues. I then characterized hemSCs isolated from IH patient specimens for TBX2 expression through mRNA analysis and cell staining. To determine the role of TBX2 in hemSC differentiation, I cultured the cells in mesenchymal, endothelial, neuroglial, and hematopoietic induction media following TBX2 knockdown. Next, I studied the potential downstream targets by conducting expression analysis for cell cycle regulators, pluripotency factors, and T-box family members. To investigate potential upstream regulators of TBX2, I cultured hemSCs with the addition of exogenous TGFβ, and an inhibitor of TGFβ signaling pathway. To confirm functional response of hemSCs to the treatments, expression of TGFβ-responsive genes collagen I and IV was assessed. My results show that TBX2 is expressed and primarily localized to the nuclei of hemSCs; the level of expression varied between cultures of different patient-derived hemangiomas. HemSCs showed morphological changes upon differentiation towards mesenchymal, endothelial, and neuroglial lineages. TBX2 knockdown caused upregulation of hematopoietic lineage markers CD34, CD45, and ckit. Pluripotency factor and cell cycle regulator p16 expression was also upregulated upon TBX2 knockdown. Lastly, my studies show that TGFβ represses TBX2 expression, which correlates with upregulated pluripotency factor expression. My studies suggest that TBX2 modulates the differentiation-competence of hemSCs. TBX2 may in turn be regulated by TGFβ during IH involution. Understanding the signaling mechanisms regulated by TBX2 may provide a novel target pathway to promote premature involution of IH as a therapeutic option for patients.
Richmond, Niamh, "Mechanisms regulating stem cell phenotype in infantile hemangioma" (2016). Electronic Thesis and Dissertation Repository. 3835.