Vascular tissue engineering is a rapidly accelerating area. Recently, several groups have shown the feasibility of culturing vessels using cells derived from elderly humans with vascular disease. These vessels may one day be implantable for arterial bypass. However, success in culturing vessels has depended strongly on the age of the donor. Because of limited lifespan, cells from older donors (i.e. over 60 years of age) display very poor growth properties and often fail to form confluent vascular tissues. We have recently shown that lifespan extension critical for success in culturing arteries from vascular cells from older donors. Retrovirally-mediated overexpression of the human telomerase enzyme reverse transcriptase (hTERT) extends cell lifespan and enables the culture of arteries for patients older than 70 years. To be successful, conventional hTERT needs to be expressed over a long time period to lengthen telomeres and extend lifespan. However, significant concerns exist regarding use of retroviruses for hTERT delivery, in light of recent cancerous complications in retrovirally-treated patients with severe combined immunodeficiency. In addition, constitutive over-expression of hTERT could itself pre-dispose cells to malignant transformation. Hence, while lifespan extension of differentiated cells will likely continue to be critical for engineering many types of tissues, sustained retroviral delivery of hTERT is problematic. Recently, a mutant of hTERT fused with the poll DNA binding protein has been described. This mutant binds to cellular DNA and extends telomeres very rapidly, thereby providing lifespan extension of cells in a short time period. Because of its rapid action, pot1-hTERT may be delivered via more transient, nonretroviral approaches - either adenoviral, or by direct nucleofection. We anticipate that transient delivery of pot1-hTERT will affect cellular lifespan extension, without concomitant risks associated with retroviruses and sustained hTERT expression. In this application, we will study the biology of transient pot1-hTERT expression in elderly vascular cells, as a paradigm for cellular lifespan extension and tissue engineering in the elderly. We will determine effects of transient hTERT-pot1 on lifespan, phenotype, cell-cycle regulators, and on engineered vessel formation. We anticipate that pot1-hTERT may have general applicability as a powerful tool to enable the engineering of multiple types of tissues for older humans. ? ?

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Exploratory/Developmental Grants (R21)
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Vascular Cell and Molecular Biology Study Section (VCMB)
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Lundberg, Martha
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Yale University
Schools of Medicine
New Haven
United States
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