VEGF-A promotes cardiac stem cell engraftment and myocardial repair in the infarcted heart Jun-Ming Tang a,b,c, ⁎, Bin Luo b , Jun-hui Xiao a , Yan-xia Lv b , Xiao-lin Li a , Jin-he Zhao a , Fei Zheng a , Lei Zhang a , Long Chen a , Jian-Ye Yang a , Lin-Yun Guo a , Lu Wang a , Yu-Wen Yan a , Ya-Mo Pan a , Jia-Ning Wang a , Dong-sheng Li a , Yu Wan c, ⁎⁎, Shi-You Chen d a Institute of Clinical Medicine and Department of Cardiology, Renmin Hospital, Hubei University of Medicine, Shiyan, Hubei 442000, China b Department of Physiology and Key Lab of human Embryonic Stem Cell of Hubei Province, Hubei University of Medicine, Hubei 442000, China c Center for Medical Research and Department of Physiology, School of Basic Medical Sciences, Wuhan University, Hubei 430071, China d Department of Physiology & Pharmacology, The University of Georgia, Athens, GA 30602, USA abstract article info Article history: Received 28 July 2014 Received in revised form 24 November 2014 Accepted 25 January 2015 Available online 27 January 2015 Keywords: VEGF-A Cardiac stem cells Myocardial infarction VCAM-1 PKCα Background: The objective of this study was to determine whether vascular endothelial growth factor (VEGF)-A subtypes improve cardiac stem cell (CSC) engraftment and promote CSC-mediated myocardial repair in the in- farcted heart. Methods: CSCs were treated with VEGF receptor (VEGFR) inhibitors, VCAM-1 antibody (VCAM-1-Ab), or PKC-α inhibitor followed by the treatment with VEGF-A. CSC adhesion assays were performed in vitro. In vivo, the PKH26-labeled and VCAM-1-Ab or PKC-α inhibitor pre-treated CSCs were treated with VEGF-A followed by im- plantation into infarcted rat hearts. The hearts were then collected for measuring CSC engraftment and evaluat- ing cardiac fibrosis and function 3 or 28 days after the CSC transplantation. Results: All three VEGF-A subtypes promoted CSC adhesion to extracellular matrix and endothelial cells. VEGF-A- mediated CSC adhesion required VEGFR and PKCα signaling. Importantly, VEGF-A induced VCAM-1, but not ICAM-1 expression in CSCs through PKCα signaling. In vivo, VEGF-A promoted the engraftment of CSCs in infarct- ed hearts, which was attenuated by PKCα inhibitor or VCAM-1-Ab. Moreover, VEGF-A-mediated CSC engraft- ment resulted in a reduction in infarct size and fibrosis. Functional studies showed that the transplantation of the VEGF-A-treated CSCs stimulated extensive angiomyogenesis in infarcted hearts as indicated by the expres- sion of cardiac troponin T and von Willebrand factor, leading to an improved performance of left ventricle. Block- ade of PKCα signaling or VCAM-1 significantly diminished the beneficial effects of CSCs treated with VEGF-A. Conclusion: VEGF-A promotes myocardial repair through, at least in part, enhancing the engraftment of CSCs me- diated by PKCα/VCAM-1 pathway. © 2015 Elsevier Ireland Ltd. All rights reserved. 1. Introduction Vascular endothelial growth factor (VEGF) and its receptor (VEGFR) are crucial regulators of the growth, development, and differentiation of heart and blood vessels. The therapeutic effect of VEGF is signified by the improvement of heart structure and function [1,2]. VEGF-A, as a key angiogenesis factor of the VEGF family, mainly exists in three subtypes, namely VEGF121, VEGF165, and VEGF189 (mouse VEGF less one amino acid, so the corresponding VEGF subtypes VEGF120, VEGF164 and VEGF188) [3]. VEGF121 and VEGF165 are secreted into the extracellular fluid, whereas the longer isoform binds to heparin sulfate proteoglycans at the cell surface. VEGF189 is almost entirely bound to the cell surface or extracellular matrix, and is considered active after its cleavage and release from its extracellular binding site [4]. VEGF exerts its biological function by binding to tyrosine kinase receptors 1 (Flt-1) and 2 (Flk/KDR), which are also located on the surface of stem cells [5]. The role of different VEGF-A subtypes in engaging cardiac stem cells (CSCs) during the cardiac repair process, however, remains to be determined. CSCs, shown as c-Kit +, Sca1 +, MDR + and other features, are found in heart tissues and are responsible for the self-renewal of myocardial tis- sue under physiological state and the repair of damaged myocardial tissue under pathological state [6]. Studies have shown that cardioprotective c- Kit+ cells regulate the myocardial balance of angiogenic cytokines [7]. Furthermore, c-Kit+ CSCs, which can better give rise to heart tissue, offer a greater potential solution than mesenchymal stem cells (MSC) [8–10]. However, a crucial problem is the extensive loss of the cells once transplanted. Regardless of cell type, many studies have shown that the International Journal of Cardiology 183 (2015) 221–231 ⁎ Correspondence to: J.-M. Tang, Institute of Clinical Medicine, Renmin Hospital, Hubei University of Medicine, Shiyan, Hubei 442000, China. ⁎⁎ Correspondence to: Y. Wan, Department of Physiology, School of Basic Medical Sciences, Wuhan University, Wuhan, Hubei 430071, China. E-mail addresses: tangjm416@163.com (J.-M. Tang), wanyu@whu.edu.cn (Y. Wan). http://dx.doi.org/10.1016/j.ijcard.2015.01.050 0167-5273/© 2015 Elsevier Ireland Ltd. All rights reserved. Contents lists available at ScienceDirect International Journal of Cardiology journal homepage: www.elsevier.com/locate/ijcard