Novel Role of Reactive Oxygen Species–Activated trp Melastatin Channel-2 in Mediating Angiogenesis and Postischemic Neovascularization

Abstract

Objective— Transient receptor potential melastatin-2 (TRPM2) channel is a nonselective cation channel that mediates influx of Ca 2+ and Na + with relative permeability of P Ca: P Na ≈0.6 in response to cellular oxidative stress. As angiogenesis and ischemic neovascularization are both significantly dependent on oxidant signaling, here we investigated the possible role of vascular endothelial growth factor (VEGF)–induced reactive oxygen species production in activating TRPM2-dependent Ca 2+ signaling and in the mechanism of angiogenesis and ischemic neovascularization. Approach and Results— We observed that VEGF stimulation rapidly induced the association of TRPM2 and cellular Src kinase with vascular endothelial-cadherin forming a signalplex at vascular endothelial-cadherin junctions in endothelial cells. Using endothelial cells isolated from TRPM2 −/− mice or after small interfering RNA depletion of TRPM2, we demonstrated that TRPM2-activated Ca 2+ signaling was required for cellular Src kinase–induced phosphorylation of vascular endothelial-cadherin at Y658 and Y731, the crucial sites involved in vascular endothelial-cadherin internalization in response to VEGF. VEGF-induced reactive oxygen species generation activated TRPM2-induced Ca 2+ entry, whereas the reactive oxygen species–insensitive TRPM2 mutant (C1008→A) showed impaired Ca 2+ entry. Endothelial cells depleted of TRPM2 also displayed significantly perturbed migratory phenotype and impaired activation of cellular Src in response to VEGF. TRPM2 −/− mice reconstituted with wild-type myeloid cells demonstrated aberrant angiogenesis and neovascularization in the hindlimb ischemia model as compared with wild-type mice. Conclusions— VEGF-induced angiogenesis and postischemic neovascularization in mice required reactive oxygen species generation in endothelial cells and resultant TRPM2 activation. Thus, our findings provide novel insight into the role of TRPM2 in mechanism of angiogenesis and ischemic neovascularization.