Abstract 14016: Adenoviral Gene Delivery Reprograms Cardiac Fibroblasts into Cardiomyocyte-like Cells with in vitro Contractile Activity: Implications for Clinical Cardiac Regeneration

Background: A newly demonstrated capability to directly convert fibroblasts into cardiomyocyte-like cells (induced cardiomyocytes [iCMs]) has great potential for the in situ regeneration of functional myocardial tissue from cardiac scar tissue and treatment of cardiovascular disease. Whereas such transdifferentiation has previously been demonstrated using retrovirus and lentivirus vectors, the clinical utility of this cardiac regeneration strategy would be facilitated by the use of a non-integrating, short-term expression viral vector such as adenovirus. In the present study, we sought to determine whether rat cardiac fibroblasts could be converted to iCMs by adenoviral expression of Gata4 (G), Mef2c (M), and Tbx5 (T) and compare the efficiencies of two viral reprogramming strategies. Methods and Results: Human adenovirus serotype 5 (Ad5) or lentivirus expressing rat GMT or corresponding null viruses were transduced into cultures of rat primary cardiac fibroblasts. After 48 hours viruses were removed and the cells were allowed to transdifferentiate under normal culture conditions. Transdifferentiation was determined 2 weeks post-transduction by FACS and immunofluorescence (IF) analysis for the activation of endogenous cardiac troponin T (cTnT) and other cardiac markers, as well as detection of in vitro contractile activity. Consistent with IF expression patterns, FACS analysis demonstrated that the Ad encoding for GMT induced 7% of adult cardiac fibroblasts to express cTnT, compared with 6% cells treated with lentivirus encoding GMT, and 0.3% of cells treated with a null virus. Importantly, only AdGMT-treated cells demonstrated in vitro contractile activity, occurring 14 days after GMT administration. Conclusion: Adenoviral vectors encoding GMT effectively reprogram cardiac fibroblasts into cardiomyocyte-like cells possessing in vitro contractile activity. These findings represent an important milestone in the utilization of a clinically relevant, short-term expression (adenovirus) vector to generate induced cardiomyocytes.