Zhao Hengli, Zhang Kaiyuan, Tang Rongrui, Meng Hui, Zou Yongjie, Wu Pengfei, Hu Rong, Liu Xin, Feng Hua, Chen Yujie
Department of Neurosurgery, Southwest Hospital, Third Military Medical University, Chongqing, China.
Department of Neurosurgery, Daping Hospital, Third Military Medical University, Chongqing, China.
Front Mol Neurosci. 2018 Mar 27;11:97. doi: 10.3389/fnmol.2018.00097. eCollection 2018.
Blood-brain barrier (BBB) disruption and subsequent brain edema play important roles in the secondary neuronal death and neurological dysfunction that are observed following intracerebral hemorrhage (ICH). In previous studies, transient receptor potential vanilloid 4 (TRPV4), a calcium-permeable mechanosensitive channel, was shown to induce cytotoxicity in many types of cells and to play a role in orchestrating barrier functions. In the present study, we explored the role of TRPV4 in ICH-induced brain injury, specifically investigating its effect on BBB disruption. Autologous arterial blood was injected into the basal ganglia of rats to mimic ICH. Adult male Sprague Dawley rats were randomly assigned to sham and experimental groups for studies on the time course of TRPV4 expression after ICH. The selective TRPV4 antagonist HC-067047 and TRPV4 siRNA were administered to evaluate the effects of TRPV4 inhibition. GSK1016790A, a TRPV4 agonist, was administered to naive rats to verify the involvement of TRPV4-induced BBB disruption. A PKC inhibitor, dihydrochloride (H7), and a selective RhoA inhibitor, C3 transferase, were administered to clarify the involvement of the PKCα/RhoA/MLC2 pathway following ICH. Post-ICH assessments including functional tests, brain edema measurements, Evans blue extravasation, western blotting and immunohistochemical assays were performed. TRPV4 inhibition remarkably ameliorated neurological symptoms, brain edema, and neuronal death, as well as BBB disruption, 24-72 h following ICH. Meanwhile, TRPV4 blockade preserved the expression of adherens and tight junction proteins, as well as BBB integrity, by inhibiting stress fiber formation, which might be correlated with the regulation of components of the PKCα/RhoA/MLC2 pathway. Furthermore, adherens and tight junction protein degradation induced by GSK1016790A treatment in naive rats was also related to PKCα/RhoA/MLC2-pathway-mediated stress fiber formation. Based on these findings, therapeutic interventions targeting TRPV4 may represent a novel approach to ameliorate secondary brain injury following ICH.
血脑屏障（BBB）破坏及随后的脑水肿在脑出血（ICH）后观察到的继发性神经元死亡和神经功能障碍中起重要作用。在先前的研究中，瞬时受体电位香草酸受体4（TRPV4），一种钙通透性机械敏感通道，被证明可在多种类型细胞中诱导细胞毒性，并在协调屏障功能中发挥作用。在本研究中，我们探讨了TRPV4在ICH诱导的脑损伤中的作用，特别研究了其对BBB破坏的影响。将自体动脉血注入大鼠基底神经节以模拟ICH。成年雄性Sprague Dawley大鼠被随机分为假手术组和实验组，用于研究ICH后TRPV4表达的时间进程。给予选择性TRPV4拮抗剂HC-067047和TRPV4 siRNA以评估TRPV4抑制的效果。将TRPV4激动剂GSK1016790A给予未处理的大鼠以验证TRPV4诱导的BBB破坏的参与情况。给予PKC抑制剂二盐酸盐（H7）和选择性RhoA抑制剂C3转移酶以阐明ICH后PKCα/RhoA/MLC2途径的参与情况。进行了包括功能测试、脑水肿测量、伊文思蓝外渗、蛋白质印迹和免疫组织化学分析在内的ICH后评估。TRPV4抑制在ICH后24 - 72小时显著改善了神经症状、脑水肿、神经元死亡以及BBB破坏。同时，TRPV4阻断通过抑制应力纤维形成来维持黏附连接和紧密连接蛋白的表达以及BBB完整性，这可能与PKCα/RhoA/MLC2途径成分的调节相关。此外，GSK1016790A处理在未处理大鼠中诱导的黏附连接和紧密连接蛋白降解也与PKCα/RhoA/MLC2途径介导的应力纤维形成有关。基于这些发现，针对TRPV4的治疗干预可能代表一种改善ICH后继发性脑损伤的新方法。
