Yingnan Bai1, Rongle Liu1, Zhixing Li1, Yaqi Zhang1, Xiaoyan Wang1, Jian Wu1,2, Zhuoquan Li3, Sanli Qian1, Bingyu Li1, Zhenzhong Zhang1, Abdel Hamid Fathy3, Donato Cappetta4, Jingmin Zhou1, Yunzeng Zou1,2, Juying Qian1, Junbo Ge1. 1. Department of Cardiology, Shanghai Institute of Cardiovascular Diseases, Zhongshan Hospital, Fudan University, Shanghai 200032, China. 2. Institute of Biomedical Science, Fudan University, Shanghai 200032, China. 3. The Institute for Translational Nanomedicine, Shanghai East Hospital, The Institute for Biomedical Engineering & Nano Science, Tongji University School of Medicine, Shanghai 200092, China. 4. Department of Experimental Medicine, Section of Pharmacology, University of Campania Luigi Vanvitelli, Napoli, Italy.
Abstract
BACKGROUND: The regulation of angiogenesis in the treatment of cardiovascular diseases has been widely studied and the vascular endothelial growth factor (VEGF) families and VEGF receptor (VEGFR) have been proven to be one of the key regulators. The VEGFR endocytosis has been recently proved to be involved in the regulation of angiogenesis. Our previous study showed that the upregulation of VEGFR endocytosis enhanced angiogenesis in vitro. In this research, we utilized mice with induced hindlimb ischemia, as a model to investigate the role of VEGFR endocytosis in the regulation of angiogenesis in vivo. Our goal was to observe the effect of revascularization with different degrees of VEGFR endocytosis after injecting atypical protein kinase C inhibitor (αPKCi) and dynasore, which could respectively promote and inhibit the VEGFR endocytosis. METHODS: We induced the hindlimb ischemia in adult male mice by ligating the hindlimb artery. By directly injecting the ischemic muscles with endothelial progenitor cells (EPCs) alone or EPCs + αPKCi/EPCs + dynasore or control medium (sham group), we divided the mice into four groups and detected lower limb blood flow using a laser Doppler blood perfusion imager. We also measured the immunohistochemistry (IHC) of markers for angiogenesis, such as CD31 and alpha smooth muscle actin (α-SMA) in the ischemic hindlimb tissues. RESULTS: We demonstrated VEGFR endocytosis played an important role in the angiogenesis of the ischemic hindlimb model in vivo. By using atypical PKC inhibitor that increase the VEGFR endocytosis, the angiogenesis in the mice model was promoted. Treatment with EPCs + αPKCi showed greater effects on blood perfusion recovery and increased the α-SMA-positive vessels. CONCLUSIONS: The regulation of VEGFR endocytosis represents a valuable method of improving angiogenesis and thus revascularization in ischemic disease model.
BACKGROUND: The regulation of angiogenesis in the treatment of cardiovascular diseases has been widely studied and the vascular endothelial growth factor (VEGF) families and VEGF receptor (VEGFR) have been proven to be one of the key regulators. The VEGFR endocytosis has been recently proved to be involved in the regulation of angiogenesis. Our previous study showed that the upregulation of VEGFR endocytosis enhanced angiogenesis in vitro. In this research, we utilized mice with induced hindlimb ischemia, as a model to investigate the role of VEGFR endocytosis in the regulation of angiogenesis in vivo. Our goal was to observe the effect of revascularization with different degrees of VEGFR endocytosis after injecting atypical protein kinase C inhibitor (αPKCi) and dynasore, which could respectively promote and inhibit the VEGFR endocytosis. METHODS: We induced the hindlimb ischemia in adult male mice by ligating the hindlimb artery. By directly injecting the ischemic muscles with endothelial progenitor cells (EPCs) alone or EPCs + αPKCi/EPCs + dynasore or control medium (sham group), we divided the mice into four groups and detected lower limb blood flow using a laser Doppler blood perfusion imager. We also measured the immunohistochemistry (IHC) of markers for angiogenesis, such as CD31 and alpha smooth muscle actin (α-SMA) in the ischemic hindlimb tissues. RESULTS: We demonstrated VEGFR endocytosis played an important role in the angiogenesis of the ischemic hindlimb model in vivo. By using atypical PKC inhibitor that increase the VEGFR endocytosis, the angiogenesis in the mice model was promoted. Treatment with EPCs + αPKCi showed greater effects on blood perfusion recovery and increased the α-SMA-positive vessels. CONCLUSIONS: The regulation of VEGFR endocytosis represents a valuable method of improving angiogenesis and thus revascularization in ischemic disease model.
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