M.Sc Thesis

M.Sc StudentSela Yogev
SubjectInducing Differentiation of Human Embryonic Stem Cells to
Endothelial Cells by Genetic Manipulation
DepartmentDepartment of Medicine
Supervisor PROFESSOR EMERITUS Joseph Itskovitz
Full Thesis textFull thesis text - English Version


Introduction: Ischemic diseases are one the major mortality causes in the world, and so far lacks efficient treatment. Transplantation of endothelial cells and endothelial progenitor cells had been suggested as a treatment to ischemic diseases and other vascular abnormalities. However, cell transplantation therapy requires sufficient number of cells which are difficult to obtain using other cell types as they show limited proliferation capacity. Human embryonic stem cells (hESC) are pluripotent cells capable of differentiation to virtually all cell lineages including endothelial cells.  Endothelial cell and vascular smooth muscle cells arise in early gastrulation of embryonic development in a process called vasculogenesis in which blood vessels are being formed. VEGF plays a key role in the vasculogenesis process and is known to promote vascular cell proliferation, survival and migration and might also be involved in their differentiation.  Aim: Since only a small fraction of hESC spontaneously differentiate to endothelial cells or their progenitors, it is important to develop methods to better direct hESC differentiation to the vascular lineages. Thus we chose to study the effect of VEGF on differentiating hESC.  Methods:  hESC have been transduced with a VEGF overexpression retroviral vector and cultured in α-MEM medium that promotes mesodermal differentiation. Results: Retroviral vectors are poorly expressed in undifferentiated hESC, probably due to gene silencing. However, overexpression of VEGF in the small fraction of differentiating cells of the undifferentiated population, following antibiotic selection, results in a great number of vascular cells containing both endothelial cells and vascular smooth muscle cells overexpressing VEGF that appear to be functional IN VITRO. Conclusion: Residual differentiated cells but not undifferentiated ESC can be genetically manipulated using MMLV-based retroviral vector, and those show increased vascular phenotype and growth under VEGF and Angiopoietin-1 overexpression regiment, suggestive of selective pressure towards cell types that express their receptors.