TY - JOUR
T1 - VEGF-Induced Endothelial Podosomes via ROCK2-Dependent Thrombomodulin Expression Initiate Sprouting Angiogenesis
AU - Kuo, Cheng Hsiang
AU - Huang, Yi Hsun
AU - Chen, Po Ku
AU - Lee, Gang Hui
AU - Tang, Ming Jer
AU - Conway, Edward M.
AU - Shi, Guey Yueh
AU - Wu, Hua Lin
N1 - Funding Information:
We thank Miss Bi-Ing Chang’s assistance on performing Western blot assay, proximity ligation assay and analysis of experiment results, thank Miss Wei-Ting Chang’s support on performing experimental wound healing assay in mice, and thank Miss I-Chen Peng’s help on performing oxygen-induced retinopathy model in mice. We thank the staff of the imaging core at the First Core Labs, National Taiwan University College of Medicine and from the Core Research Laboratory, College of Medicine, National Cheng Kung University, for technical assistance. This work was supported in part by the Electron Microscopy Facility in National Yang-Ming University. We are grateful for administrative and laboratory support from the International Center for Wound Repair and Regeneration of National Cheng Kung University.
Publisher Copyright:
© 2021 American Heart Association, Inc.
PY - 2021/5/5
Y1 - 2021/5/5
N2 - Objective: VEGF (vascular endothelial growth factor) plays a critical role in physiological and pathological angiogenesis. Endothelial 3D podosomes (3DPs) are a type of F-actin-rich membrane microdomain, predominantly found in endothelial tip cells controlled by VEGF signaling during sprouting angiogenesis, such as occurs in retinal vasculature development. The molecular mechanisms governing 3DP formation have not been completely elucidated. Approach and Results: By using in vitro cell models and in vivo mouse models, we study the role of TM (thrombomodulin) in VEGF-induced endothelial 3DPs. Here, we report that VEGF can induce the expression of TM via ROCK2 (Rho-associated coiled-coil kinase 2). Furthermore, ROCK2 can catalyze the phosphorylated activation of ezrin to promote the association of the cytoplasmic domain of TM with F-actin in 3DPs and thereby promote the formation of 3DPs. We used endothelial cells transfected with different TM mutants as models to verify the role of TM domains in 3DPs and angiogenic activity. TM expression in endothelial cells augments angiogenic activity, a response that is dependent on the interaction of the cytoplasmic tail of TM with ezrin, and the integrity of the lectin-like domain of TM. Thus, as compared with wild-type counterparts, mice lacking the lectin-like domain of TM exhibit reduced neovascularization of granulation tissues during cutaneous wound healing and less retinal neovascularization in a model of oxygen-induced retinopathy. Conclusions: VEGF-ROCK2-ezrin-TM-F-actin axis promotes the formation of the lipid raft membrane-associated complex configuration, 3DP, which plays a critical role in mediating tube formation and cell migration of endothelial cells in sprouting angiogenesis.
AB - Objective: VEGF (vascular endothelial growth factor) plays a critical role in physiological and pathological angiogenesis. Endothelial 3D podosomes (3DPs) are a type of F-actin-rich membrane microdomain, predominantly found in endothelial tip cells controlled by VEGF signaling during sprouting angiogenesis, such as occurs in retinal vasculature development. The molecular mechanisms governing 3DP formation have not been completely elucidated. Approach and Results: By using in vitro cell models and in vivo mouse models, we study the role of TM (thrombomodulin) in VEGF-induced endothelial 3DPs. Here, we report that VEGF can induce the expression of TM via ROCK2 (Rho-associated coiled-coil kinase 2). Furthermore, ROCK2 can catalyze the phosphorylated activation of ezrin to promote the association of the cytoplasmic domain of TM with F-actin in 3DPs and thereby promote the formation of 3DPs. We used endothelial cells transfected with different TM mutants as models to verify the role of TM domains in 3DPs and angiogenic activity. TM expression in endothelial cells augments angiogenic activity, a response that is dependent on the interaction of the cytoplasmic tail of TM with ezrin, and the integrity of the lectin-like domain of TM. Thus, as compared with wild-type counterparts, mice lacking the lectin-like domain of TM exhibit reduced neovascularization of granulation tissues during cutaneous wound healing and less retinal neovascularization in a model of oxygen-induced retinopathy. Conclusions: VEGF-ROCK2-ezrin-TM-F-actin axis promotes the formation of the lipid raft membrane-associated complex configuration, 3DP, which plays a critical role in mediating tube formation and cell migration of endothelial cells in sprouting angiogenesis.
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U2 - 10.1161/ATVBAHA.121.315931
DO - 10.1161/ATVBAHA.121.315931
M3 - Article
C2 - 33730876
AN - SCOPUS:85104821042
SN - 1079-5642
VL - 41
SP - 1657
EP - 1671
JO - Arteriosclerosis, thrombosis, and vascular biology
JF - Arteriosclerosis, thrombosis, and vascular biology
IS - 5
ER -