Vascular disease, caused by the blockage of blood vessels, is the largest cause of mortality in the developed world. Autologous or synthetic vascular grafts are used in treating this disease. However, some patients either lack suitable autologous tissue or cannot receive synthetic grafts due to the small size of target vessels. Tissue-engineered blood vessels (TEBVs) grown using vascular smooth muscle cells (SMCs) isolated from primary tissue hold great potential as tools for surgical replacement of the affected vessels in these patients. However, the development of autologous TEBVs for clinical application using SMCs has been hampered by limited accessibility to patient vascular SMCs, rapid loss of SMC differentiation in cell culture and limited ability of primary SMCs to expand. Thus, it is of great interest to establish a human cell-based model that provides an abundant and renewable source of functional SMCs for the establishment of TEBVs. A renewable source of human cells can be generated by human induced pluripotent stem cells (hiPSCs), which resemble human embryonic stem cells (hESCs) and can be derived from a person's own somatic cells by forced gene expression. Both hiPSCs and hESCs can self-renew and differentiate into virtually every cell type in the human body including functional vascular SMCs, providing ideal cell sources for generating TEBVs to treat vascular diseases. We recently established hiPSC lines and derived unlimited amounts of highly homogeneous functional vascular SMCs from hiPSCs (hiPSC-SMCs) and hESCs (hESC-SMCs). As the potential reactivation of reprogramming transgenes in these iPSCs could ultimately affect their safety as therapy and utility in disease modeling, we will generate and validate transgene-free hiPSC lines by using Cre recombinase and then derive and characterize hiPSC-SMCs from these transgene-free hiPSC lines. TEBVs typically lack elastin (ELN), which is essential to mechanical properties of blood vessels, providing recoil and resistance to aneurysm and dilation. Since we have shown that inhibition of microRNA-29a (miR-29a) markedly increases the expression of ELN, crosslinking of ELN fiber and distensibility of TEBVs derived from primary SMCs, we will generate TEBVs using hiPSC-SMCs and hESC-SMCs in the presence of a miR-29a inhibitor and then determine the suture retention strength, burst pressure, collagen content, ELN content, and mechanical properties of TEBVs. To investigate the function of SMC-derived TEBVs in vivo we will implant TEBVs as aortic interpositional grafts in nude rats. We choose the rat model since other preferred large animals (dog or pig) might reject the human tissue even with immunosuppression due to a significant xenogenic response. Although a non-human primate model is not appropriate for a first in vivo study of such new hiPSC technology, it could be used in the future with immunosuppression as a follow-up model if the rodent model succeeds. We will test the hypothesis that TEBVs derived from hiPSC-SMCs and hESC-SMCs possess suitable properties for implantation in vitro and then remain mechanically stable in a rat aortic model in vivo.
This research will lead to the production of autologous, well-characterized vascular smooth muscle cells as an abundant renewable cell source for cell-based therapies and for disease mechanism study. Development of tissue-engineered blood vessels with markedly improved elasticity/compliance using smooth muscle cells derived from human induced pluripotent stem cells will enable us to take the next step toward developing autologous tissue engineered grafts for clinical intervention in vascular diseases.
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