[束缚应激通过激活Rho/ROCK信号通路诱导大鼠杏仁核血脑屏障损伤]
[Restraint stress induces blood-brain barrier injury in rat amygdala by activating the Rho/ROCK signaling pathway].
作者信息
Xu G, Gao A, Cong B
机构信息
Department of Forensic Medicine, National Police University for Criminal Justice, Baoding 071000, China.
College of Forensic Medicine, Hebei Medical University, Hebei Key Laboratory of Forensic Medicine, Shijiazhuang 050017, China.
出版信息
Nan Fang Yi Ke Da Xue Xue Bao. 2024 Mar 20;44(3):411-419. doi: 10.12122/j.issn.1673-4254.2024.03.01.
OBJECTIVE
To investigate the role of Rho/ROCK signaling pathway in mediating restraint stress-induced blood-brain barrier (BBB) injury in the amygdala of rats.
METHODS
Sixty male SD rats were randomized equally into control group (with food and water deprivation for 6 h per day), restraint stress group (with restraint for 6 h per day), stress + fasudil treatment (administered by intraperitoneal injection at 1 mg/100 g 30 min before the 6-h restraint) group, and fasudil treatment alone group. The elevated plus-maze test was used to detect behavioral changes of the rats, serum corticosterone and S100B levels were determined with ELISA, and Evans Blue leakage in the brain tissue was examined to evaluate the changes in BBB permeability. The changes in expression levels of tight junction proteins in the amygdala were detected using immunofluorescence assay and Western blotting, and Rho/ROCK pathway activation was detected by Pull-down test and Western blotting. Ultrastructural changes of the cerebral microvascular endothelial cells were observed using transmission electron microscopy.
RESULTS
Compared with those in the control group, the rats in restrain stress group and stress+fasudil group showed obvious anxiety-like behavior with significantly increased serum corticosterone level (<0.001). Compared with those in the control group and stress+fasudil group, the rat models of restrain stress showed more obvious Evans Blue leakage and higher S100B expression (<0.01) but lower expressions of tight junction proteins in the amygdala. Pull-down test and Western blotting confirmed that the expression levels of RhoA-GTP, ROCK2 and P-MLC 2 were significantly higher in stress group than in the control group and stress + fasudil group (<0.05). Transmission electron microscopy revealed obvious ultrastructural changes in the cerebral microvascular endothelial cells in the rat models of restrain stress.
CONCLUSION
Restraint stress induces BBB injury in the amygdala of rats by activating the Rho/ROCK signaling pathway.
目的
探讨Rho/ROCK信号通路在介导束缚应激诱导大鼠杏仁核血脑屏障（BBB）损伤中的作用。
方法
将60只雄性SD大鼠随机均分为对照组（每天禁食禁水6小时）、束缚应激组（每天束缚6小时）、应激+法舒地尔治疗组（在6小时束缚前30分钟按1mg/100g腹腔注射给药）和单纯法舒地尔治疗组。采用高架十字迷宫试验检测大鼠行为变化，用ELISA法测定血清皮质酮和S100B水平，检测脑组织伊文思蓝渗漏情况以评估BBB通透性变化。采用免疫荧光法和蛋白质印迹法检测杏仁核紧密连接蛋白表达水平变化，用Pull-down试验和蛋白质印迹法检测Rho/ROCK通路激活情况。用透射电子显微镜观察脑微血管内皮细胞超微结构变化。
结果
与对照组相比，束缚应激组和应激+法舒地尔组大鼠出现明显的焦虑样行为，血清皮质酮水平显著升高（<0.001）。与对照组和应激+法舒地尔组相比，束缚应激大鼠模型伊文思蓝渗漏更明显，S100B表达更高（<0.01），但杏仁核紧密连接蛋白表达更低。Pull-down试验和蛋白质印迹法证实，应激组RhoA-GTP、ROCK2和P-MLC 2表达水平显著高于对照组和应激 + 法舒地尔组（<0.05）。透射电子显微镜显示，束缚应激大鼠模型脑微血管内皮细胞有明显超微结构变化。
结论
束缚应激通过激活Rho/ROCK信号通路诱导大鼠杏仁核BBB损伤。
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