Yun Liu1, Min Gao2, Ming-Ming Ma2, Yong-Bo Tang2, Jia-Guo Zhou2, Guan-Lei Wang2, Yan-Hua Du3, Yong-Yuan Guan4. 1. Department of Pharmacology, Cardiac & Cerebral Vascular Research Center, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, 510089, China; Department of Pharmacolgy, Guangzhou Medical University, Guangzhou, 510182, China. 2. Department of Pharmacology, Cardiac & Cerebral Vascular Research Center, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, 510089, China. 3. Department of Pharmacology, Cardiac & Cerebral Vascular Research Center, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, 510089, China. Electronic address: duyanhua@mail.sysu.edu.cn. 4. Department of Pharmacology, Cardiac & Cerebral Vascular Research Center, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, 510089, China. Electronic address: guanyy@mail.sysu.edu.cn.
Abstract
BACKGROUND: Apoptosis plays a central role in maintaining the normal cell number and tissue homeostasis. Endophilins are a family of evolutionarily conserved proteins that have the critical role in endocytosis. Here, we determined whether endophilin A2 (EndoII) contributes to hydrogen peroxide (H2O2)-induced apoptosis in rat basilar artery smooth muscle cells (BASMCs) and the underlying mechanisms. METHODS AND RESULTS: By using small interference RNA (siRNA) and EndoII overexpression strategy, we found that EndoII siRNA knockdown reduced cell viability and promoted H2O2-induced cell apoptosis, evidenced by loss of mitochondrial membrane potential, release of cytochrome c, and activation of caspase-9, 3 and poly (ADP-ribose) polymerase (PARP). In contrast, EndoII overexpression showed opposite effects and inhibited H2O2-induced BASMCs apoptosis. Further studies revealed that there was a direct interaction between EndoII and Bax. Upon H2O2-induced apoptosis, the association of EndoII with Bax were significantly decreased, while the interaction of Bax/tBid were increased, accompanied by a translocation of Bax from cytosol to mitochondria. Knockdown of EndoII did not affect the expression of Bax, but further promoted the binding of Bax with tBid and favored the accumulation of Bax to mitochondria as well as Bax activation; whereas EndoII overexpression produced the opposite effects. In addition, EndoII siRNA aggravated, but EndoII overexpression alleviated, the reduction of Bcl-2 expression in H2O2-treated cells. CONCLUSIONS: These data suggested a role of EndoII in protecting BASMCs apoptosis induced by H2O2, possibly by inhibiting the addressing of Bax to mitochondria. Targeting on EndoII may be a new strategy to treat apoptosis-associated diseases.
BACKGROUND: Apoptosis plays a central role in maintaining the normal cell number and tissue homeostasis. Endophilins are a family of evolutionarily conserved proteins that have the critical role in endocytosis. Here, we determined whether endophilin A2 (EndoII) contributes to hydrogen peroxide (H2O2)-induced apoptosis in rat basilar artery smooth muscle cells (BASMCs) and the underlying mechanisms. METHODS AND RESULTS: By using small interference RNA (siRNA) and EndoII overexpression strategy, we found that EndoII siRNA knockdown reduced cell viability and promoted H2O2-induced cell apoptosis, evidenced by loss of mitochondrial membrane potential, release of cytochrome c, and activation of caspase-9, 3 and poly (ADP-ribose) polymerase (PARP). In contrast, EndoII overexpression showed opposite effects and inhibited H2O2-induced BASMCs apoptosis. Further studies revealed that there was a direct interaction between EndoII and Bax. Upon H2O2-induced apoptosis, the association of EndoII with Bax were significantly decreased, while the interaction of Bax/tBid were increased, accompanied by a translocation of Bax from cytosol to mitochondria. Knockdown of EndoII did not affect the expression of Bax, but further promoted the binding of Bax with tBid and favored the accumulation of Bax to mitochondria as well as Bax activation; whereas EndoII overexpression produced the opposite effects. In addition, EndoII siRNA aggravated, but EndoII overexpression alleviated, the reduction of Bcl-2 expression in H2O2-treated cells. CONCLUSIONS: These data suggested a role of EndoII in protecting BASMCs apoptosis induced by H2O2, possibly by inhibiting the addressing of Bax to mitochondria. Targeting on EndoII may be a new strategy to treat apoptosis-associated diseases.