Ischemic heart disease often results in myocardial infarction and is the leading cause of mortality and morbidity worldwide. Repopulating myocardium with contractile cells is a purpose of regenerative medicine. The most investigated strategy is implantation of stem cells. Mesenchymal stem cells (MSCs) are multipotent stromal cells that can differentiate into a variety of cell types. Tbx-20 is a transcription factor which is essential for heart development and function. Any mutations in this gene can result in congenital heart disease. The aim of current study is constructing lentiviral vectors bearing TBX- 20, to differentiate MSCs to cardiomyocytes. To do so, specific sequences of TBX-20 was cloned into lentiviral vector. Lentiviruses were made through cotransfection of 293T cells with main construct and helper plasmids. Transfection efficiency was checked by fluorescent microscopy and then lentiviruses were harvested and titrated. Transduction of MSCs was carried out for 16 h before fresh growth medium was added to the cells. Cardiogenesis was checked by real-time PCR and immunostaining after 14 days of transduction. Analytical digestion and nucleotide sequence analysis showed that the lentiviral construct did not have any mutations. Transfection efficiency of 293T cells was about 80%. Then, concentrated viruses were successfully harvested and titrated. Real-time PCR results showed the over expression of ACTC1, TNNI3 and ACTN genes, all as cardiogenic differentiation markers. Immunostaining results showed over expression of α-actinin and cardiac muscle troponin T in MSCs. In this study we successfully showed that Tbx-20 transcription factor has cardiogenic effects and can differentiate MSCs to cardiomyocytes.

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