Factors released from TGF beta2 Primed ES cells inhibit Apoptosis and Necrosis in the Cell Culture System and Improve Infarcted Heart Function Following Transplantation

Abstract

We reported that ES‐conditioned medium (CM) contains anti‐apoptotic factors that inhibit H2O2 induced apoptosis in H9c2 cells. However, whether ES‐CM prepared with the treatment of pro‐cardiac and anti‐apoptotic cytokine TGF beta 2 will demonstrate enhanced inhibition of apoptosis and necrosis in the cell culture is unknown. We provide evidence that TGFbeta 2 released factors enhanced inhibition of iodoacetic acid (IAA)‐induced apoptosis and necrosis in H9c2 cells. Conditioned medium (CM) from growing mouse ES cells treated with and without TGFbeta 2 was generated. Apoptosis and necrosis was induced after exposure to IAA in H9c2 cells for 1h followed by replacement with fresh cell culture or ES‐CMs. After 24 h, H9c2 cells treated with both ES‐CMs demonstrated significantly decreased apoptosis, as determined by TUNEL staining, apoptotic ELISA and caspase‐3 activity. Next, necrosis was also decreased as determined by flow cytometry and the release of lactate dehydrogenase (LDH). Importantly, TGF beta2‐ES‐CM significantly enhanced inhibition of apoptosis and necrosis compared with untreated ES‐CM. Furthermore, whether TGF beta 2 primed ES cells following transplantation in the infarcted mouse hearts will contribute to the improved cardiac function remains elusive. Using our standard myocardial infarction model, we transplanted with and without TGF beta2 primed ES cells (50,000 ES cells in 10‐20 μl of medium) in the infarcted heart. Echocardiography was performed at day 7 and 14. Cardiac function was significantly improved following TGF beta 2 primed ES‐cells transplantation compared with unprimed ES cells. In conclusions, we suggest that factors released from TGF beta2 primed ES cells enhances the inhibition of apoptosis and necrosis as well as improve cardiac function.