The potential of marrow stromal cells for ex vivo gene therapy of cardiovascular diseases.

摘要

Marrow stromal cells (MSCs), also known as mesenchymal stem cells, are multipotent adult stem cells from bone marrow. MSCs can be easily isolated, readily ex vivo expanded, and efficiently gene engineered. MSCs do not elicit immune rejection and can survive for a long period of time in vivo after autologous transplantation. These properties make MSCs attractive vehicles for ex vivo gene therapy of various diseases. Endothelial nitric oxide synthase (eNOS) and calcitonin gene-related peptide (CGRP) are important molecules in the cardiovascular system. Nitric oxide (NO) or CGRP deficiency has been suggested to be involved in cardiovascular diseases. Therefore, the enhancement of NO or CGRP delivery is a promising approach for treatment of cardiovascular diseases. An attractive method of increasing NO or CGRP delivery is the use of gene transfer technology to locally increase eNOS or CGRP gene expression. However, in vivo administration of nonviral vectors or viral vectors usually leads to low levels of gene transfer, and as the transgene is randomly expressed in almost all cell types, adverse effects can occur.; To develop an improved therapy using adult stem cells, we hypothesize that MSCs can be used for ex vivo gene therapy of cardiovascular diseases. Using adenoviral vectors containing eNOS, CGRP, and reporter gene ntlacZ, we demonstrate that transduction efficiency of adenoviral-mediated gene transfer into ex vivo expanded rat MSCs (rMSCs) is dose-dependent, that transgene expression persists for over 21 days in culture, that rMSCs retain multipotentiality after adenoviral transduction and express transgene after cell differentiation, and that transduction at MOI 300 does not alter the proliferation and viability of rMSCs. We demonstrate that high level eNOS expression and high level CGRP secretion by rMSCs can be achieved after adenoviral-mediated gene transfer. Using a rat model of age-related erectile dysfunction, we demonstrate that intracavernosal injection of rMSCs expressing eNOS can improve erectile function. These findings suggest that this novel adult stem cell-based ex vivo gene therapy may represent a new form of therapy for cardiovascular diseases. In appendix, we demonstrate that human MSCs can be differentiated into early progenitors of neural cells by conditions that increase intracellular cyclic AMP.