Doctor of Philosophy
Chemical and Biochemical Engineering
University of Western Ontario
Vascular tissue engineering (VTE) is an emerging alternative therapeutic intervention strategy to treat diseases such as atherosclerosis. While the ultimate goal of VTE is designing tissues to serve as graft substitutes, they can also serve as powerful tools to study tissue and disease development and drug discovery.
In this work, engineered vascular tissues from fibrin gel, mouse embryonic multipotent progenitor cell line (10T1/2 cells), and rat embryonic thoracic artery smooth muscle cells (A-10 cells) were used as models to study the Notch signaling pathway and vascular calcification. The 10T1/2 cells were successfully differentiated into vascular smooth muscle cells with TGFβ1 treatment and compacted the tubular gel significantly owing to the contractile cytoskeletal stress fibers. Notch signaling studies in engineered vascular tissues from A-10 cells demonstrated cis-inhibition while 10T1/2 cells activated Notch and its downstream targets Hes-1 and the smooth muscle α-actin genes.
The results from the calcification studies showed that vascular tissues fabricated from both progenitor and differentiated 10T1/2 cells calcified in response to high inorganic phosphate concentrations and expressed the osteopontin protein. Treatment of the tissues with a model therapeutic agent, Vitamin K, led to the reduction of calcium deposits and osteopontin expression, suggesting its potential protective role. In addition, vitamin K treated engineered tissues resulted in the restoration of smooth muscle cells contractile markers. The effect of elastin degradation on calcification was simulated using exogenous elastin and showed that while elastin alone did not impact the undifferentiated tissues, it led to an increase in osteogenic markers in the differentiated counterparts.
This work also investigated the role of endothelial cell vimentin in the regulation of Notch signaling and neovascularization in coculture tissue models. The preliminary results showed that vimentin might enhance the Notch signaling strength since the inhibition of vimentin using a chemical inhibitor or siRNA did not completely inhibit the signal. Notwithstanding this, vimentin appeared to be essential for new micro-vasculature formation.
The data collectively presented in this thesis demonstrated the potential of engineered vascular tissues as a novel tool to study cell signaling, vascular calcification, and therapeutic discovery.
Summary for Lay Audience
The lack of organ donors has brought about the need for engineering tissues from the lab bench. Whether patients require a blood vessel, bone, a patch of skin, liver, or even a heart, tissue engineering provides a practical alternative to the lack of donated tissues and organs. This research focused on engineering vascular tissues (blood vessels). While the main purpose of tissue engineering is to replace damaged or diseased organs, the use of engineered vascular tissues to study cell communication and diseases that affect the human vasculature is a target application. The clotting protein fibrinogen, which is naturally found in the blood of mammals, was used to entrap precursor cells and to form a tube-shaped tissue that resembles blood vessels.
Communication between cells is called signaling, and it is a defined “dialogue” that carries instructions from one cell to another in a process called signaling pathways. In vascular tissues, a vital Notch signaling pathway occurs between endothelial cells and smooth muscle cells and dictates vital functions such as cell division and survival, among other functions. In this research, the Notch signaling was studied by comparing two different cell types responses to endothelial cells in the engineered tissues. It was found that while one type of cell was responsive, the other type was not. This has implications for the use of certain cell types that are capable of communicating the proper signal for the proper development of engineered tissues. Furthermore, by using the engineered tissues as models, a common problem that occurs in arteries, vascular calcification was studied. The results in this work showed that engineered tissues were capable of calcification just like natural tissues and that there is a potential of using vitamin K to reduce the negative effects of calcification. This will help us study diseases and discover treatments for vascular diseases.
In conclusion, engineered vascular tissues have the potential to provide insight into human vasculature physiology and pathology. In this research, it has been shown that engineered vascular tissues provide excellent platforms to study cell signaling, vascular diseases, and potential drug discovery.
Dayekh, Khalil, "Fibrin-Based Engineered Vascular Tissues as Platforms for Cellular Studies and Disease Modeling" (2021). Electronic Thesis and Dissertation Repository. 7695.
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