Keyang Zhu1, Qiang Wu1, Cheng Ni1, Peng Zhang1, Zhiwei Zhong1, Yan Wu1, Yingchao Wang1, Yinchuan Xu1, Minjian Kong1, Haifeng Cheng1, Zhihua Tao1, Qian Yang1, He Liang1, Yun Jiang1, Qingju Li1, Jing Zhao1, Jijun Huang1, Fengjiang Zhang1, Qi Chen1, Yi Li1, Jinghai Chen1, Wei Zhu1, Hong Yu1, Jianyi Zhang1, Huang-Tian Yang2, Xinyang Hu2, Jian'an Wang2. 1. From the Department of Cardiology (K.Z., C.N., Z.Z., Y. Wu, Y. Wang, Y.X., Q.Y., Q.L., J. Zhao, J.C., W.Z., H.Y., X.H., J.W.), Department of Cardiovascular Surgery (M.K., H.C.), Department of Laboratory Medicine (Z.T.), and Department of Anesthesiology (F.Z., Q.C., Y.L.), Second Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, PR China; Key Laboratory of Stem Cell Biology and Laboratory of Molecular Cardiology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences (CAS) Shanghai, University of CAS, Beijing, PR China (Q.W., P.Z., H.L., Y.J., J.H., H.-T.Y.); Cardiovascular Key Laboratory of Zhejiang Province, Hangzhou, PR China (K.Z., C.N., Z.Z., Y. Wu, Y. Wang, Y.X., Q.L., J. Zhao, J.C., W.Z., H.Y., X.H., Y.W.); and Department of Biomedical Engineering, University of Alabama at Birmingham (J. Zhang). 2. From the Department of Cardiology (K.Z., C.N., Z.Z., Y. Wu, Y. Wang, Y.X., Q.Y., Q.L., J. Zhao, J.C., W.Z., H.Y., X.H., J.W.), Department of Cardiovascular Surgery (M.K., H.C.), Department of Laboratory Medicine (Z.T.), and Department of Anesthesiology (F.Z., Q.C., Y.L.), Second Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, PR China; Key Laboratory of Stem Cell Biology and Laboratory of Molecular Cardiology, Institute of Health Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences (CAS) Shanghai, University of CAS, Beijing, PR China (Q.W., P.Z., H.L., Y.J., J.H., H.-T.Y.); Cardiovascular Key Laboratory of Zhejiang Province, Hangzhou, PR China (K.Z., C.N., Z.Z., Y. Wu, Y. Wang, Y.X., Q.L., J. Zhao, J.C., W.Z., H.Y., X.H., Y.W.); and Department of Biomedical Engineering, University of Alabama at Birmingham (J. Zhang). wangjianan111@zju.edu.cn hxy0507@zju.edu.cn htyang@sibs.ac.cn.
Abstract
RATIONALE: Human pluripotent stem cell-derived cardiovascular progenitor cells (hPSC-CVPCs) should be thoroughly investigated in large animal studies before testing in clinical trials. OBJECTIVE: The main of this study is to clarify whether hPSC-CVPCs can engraft for long time in the heart of primates after myocardial infarction (MI) and compare the effectiveness and safety of immunosuppression with cyclosporine alone or multiple-drug regimen (MDR) containing cyclosporine, methylprednisolone, and basiliximab in cynomolgus monkeys that had received intramyocardial injections of 1×107 EGFP (enhanced green fluorescent protein)-expressing hPSC-CVPCs after MI. A third group of animals received the immunosuppression MDR but without cell therapy after MI (MI+MDR group). METHODS AND RESULTS: Measurements of EGFP gene levels and EGFP immunofluorescence staining indicated that the hPSC-CVPC engraftment rate was greater in the MI+MDR+CVPC group than that in the MI+cyclosporine+CVPC group. However, even in the MI+MDR+CVPC group, no transplanted cells could be detected at 140 days after transplantation. Concomitantly, immunofluorescent analysis of CD3, CD4, and CD8 expression indicated that T-lymphocyte infiltration in the CVPC-transplanted hearts was less in the MDR-treated animals than in the cyclosporine-alone-treated animals. The recovery of left ventricular function on day 28 post-MI in the MI+MDR+CVPC group was better than that in the MI+MDR group. Apoptotic cardiac cells were also less common in the MI+MDR+CVPC group than in the MI+MDR group, although both immunosuppression regimens were associated with transient hepatic dysfunction. CONCLUSIONS: This is the largest study of hPSCs in nonhuman primates in cardiovascular field to date (n=32). Compared with cyclosporine alone, MDR attenuates immune rejection and improves survival of hPSC-CVPCs in primates; this is associated with less apoptosis of native cardiac cells and better recovery of left ventricular function at 28 days. However, even with MDR, transplanted hPSC-CVPCs do not engraft and do not survive at 140 days after transplantation, thereby excluding remuscularization as a mechanism for the functional effect.
RATIONALE: Human pluripotent stem cell-derived cardiovascular progenitor cells (hPSC-CVPCs) should be thoroughly investigated in large animal studies before testing in clinical trials. OBJECTIVE: The main of this study is to clarify whether hPSC-CVPCs can engraft for long time in the heart of primates after myocardial infarction (MI) and compare the effectiveness and safety of immunosuppression with cyclosporine alone or multiple-drug regimen (MDR) containing cyclosporine, methylprednisolone, and basiliximab in cynomolgus monkeys that had received intramyocardial injections of 1×107 EGFP (enhanced green fluorescent protein)-expressing hPSC-CVPCs after MI. A third group of animals received the immunosuppression MDR but without cell therapy after MI (MI+MDR group). METHODS AND RESULTS: Measurements of EGFP gene levels and EGFP immunofluorescence staining indicated that the hPSC-CVPC engraftment rate was greater in the MI+MDR+CVPC group than that in the MI+cyclosporine+CVPC group. However, even in the MI+MDR+CVPC group, no transplanted cells could be detected at 140 days after transplantation. Concomitantly, immunofluorescent analysis of CD3, CD4, and CD8 expression indicated that T-lymphocyte infiltration in the CVPC-transplanted hearts was less in the MDR-treated animals than in the cyclosporine-alone-treated animals. The recovery of left ventricular function on day 28 post-MI in the MI+MDR+CVPC group was better than that in the MI+MDR group. Apoptotic cardiac cells were also less common in the MI+MDR+CVPC group than in the MI+MDR group, although both immunosuppression regimens were associated with transient hepatic dysfunction. CONCLUSIONS: This is the largest study of hPSCs in nonhuman primates in cardiovascular field to date (n=32). Compared with cyclosporine alone, MDR attenuates immune rejection and improves survival of hPSC-CVPCs in primates; this is associated with less apoptosis of native cardiac cells and better recovery of left ventricular function at 28 days. However, even with MDR, transplanted hPSC-CVPCs do not engraft and do not survive at 140 days after transplantation, thereby excluding remuscularization as a mechanism for the functional effect.