Stimulation of adult resident cardiac progenitor cells by durable myocardial expression of thymosin beta 4 with ultrasound-targeted microbubble delivery

It has been proposed that thymosin beta 4 (TB4)-protein delivery stimulates differentiation of resident adult WT1-positive cardiac progenitor cells, but with very low efficiency. We determined whether gene therapy with human TB4 stimulates proliferation of resident adult cardiac progenitor cells in normal rat heart. Ultrasound-targeted microbubble destruction (UTMD) was used to deliver the human TB4 gene under a piggybac transposon plasmid to normal rat heart. The rat hearts were assayed by quantitative reverse transcription-PCR and immunohistology with a confocal microscope at 1, 2, 3,4 and 12 weeks after UTMD. Exogenous TB4 stimulation resulted in the presence of WT1-positive cardiac progenitor cells from epicardium to endocardium. TB4 stimulated angiogenesis and arteriogenesis. One month after TB4 gene therapy by UTMD, the percentage of NKX2.5-positive cardiomyocytes was 5.5 +/- 1.0% and NKX2.5 mRNA was 24-fold higher than in the control groups (P < 0.001). Similar results were found for ISL-1, BrDu, Ki-67, PHH3 and aurora B (P < 0.001). Cardiac-specific delivery of exogenous human TB4 gene efficiently stimulates proliferation and differentiation of resident WT1-positive adult cardiac progenitor cells into three intact cardiac cell lineages vascular endothelial cells, coronary artery smooth muscle cells and cardiac muscle cells in normal adult rat heart. Gene Therapy (2013) 20, 225-233; doi:10.1038/gt.2012.89; published online 15 November 2012