An efficient method to generate cardiac tissue from other tissues has great therapeutic potential for patients suffering from cardiovascular disease. Pluripotent stem cells, such as embryonic stem (ES) cells, and so-called induced pluripotent stem (iPS) cells can be differentiated into multiple cell types, including cardiac myocytes. However, therapeutic use of these cells has several important risks, including cancer as well as loss of differentiated cell identity and function or "drift." Transdifferentiation, the generation of cell types from other differentiated cell types, is an attractive alternative because it poses little risk of cancer and may give rise to cells whose identity is locked in. However, the direct reprogramming of adult fibroblasts into cardiac myocytes is inefficient and the transcription factors that drive this process are unknown. To circumvent these hurdles we hypothesized that fetal derived cells may be more amenable to reprogramming into cardiac myocytes with defined transcription factors. To this end, fetal derived fibroblasts and mesenchymal cells were transduced with transcription factors that have been shown to play a role in myocyte specification and maintenance, including GATA4, Mef2c, Tbx5 and Nkx2.5. To test this hypothesis, the transcription factors were cloned into lentiviral vectors, which were used to infect fibroblasts and mesenchymal cells. Infected cells were then cultured in different media. To assess the efficiency of the reprogramming, RNA was extracted from infected and uninfected cells, and quantitative RT-PCR was used to evaluate the expression levels of the transcription factors and known cardiac genes. We expected to observe higher levels of the transcription factors and cardiac markers in the infected cells relative to the uninfected cells. The results showed that the infected cells expressed higher levels of the transcription factors and a few cardiac markers relative to the uninfected cells. However, due to the small increase in the levels of only a few cardiac markers, the reprogramming was concluded to be inefficient. To make the reprogramming more efficient, other as yet unrecognized transcription factors possibly in the GATA family of transcription factors and culture conditions are currently being considered.


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