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参芎化瘀胶囊对全脑缺血大鼠海马CA1区RHOA和ROCK-Ⅱ表达的影响
Influence on the Expression of RHOA and ROCK-Ⅱafter Global Cerebral Ischemia-reperfusion in Hippocampus byShenxiong Huayu Capsules
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摘要: 目的 探讨参芎化瘀胶囊预处理对全脑缺血大鼠海马CA1区重组蛋白A（small GTP binding protein A, RHOA）和RHO激酶2（RHO associated protein kinase-2, ROCK-Ⅱ）表达的影响。方法 96只SD雄性大鼠分3组，每组32只，造模前7 d开始灌胃，至处死日早晨结束。正常对照组（生理盐水灌胃）、全脑缺血模型组（生理盐水灌胃）和参芎化瘀胶囊+全脑缺血组（参芎化瘀胶囊0.048 g/kg体质量，溶于0.5 mL双蒸水中，每日1次，灌胃0.3 mL/100 g体质量）。后两组采用改良的Pulsinelli四血管闭塞法制作大鼠全脑缺血模型，分别在造模成功后1、3、7、14 d四个时间点采用水迷宫行为学检测评价学习记忆能力，处死大鼠后取脑组织HE 染色观察组织病理学的变化，免疫组化法观察RHOA 和ROCK-Ⅱ 表达，蛋白印迹分析检测RHOA 和ROCK-Ⅱ 蛋白含量。结果 和正常对照组比较，模型组大鼠各个时间点逃避潜伏期时间延长（P<0.05），参芎化瘀胶囊治疗后大鼠逃避潜伏期时间缩短（P<0.05），但仍长于正常对照组（P<0.05）。HE染色显示，与正常组比较，模型组大鼠海马CA1 区神经元1~14 d存活神经元逐渐减少；参芎化瘀胶囊治疗后各个时间点存活神经元增加（P<0.05），但仍少于正常组（P<0.05）；免疫组化和蛋白印迹分析显示，正常对照组RHOA 和ROCK-Ⅱ表达不明显，模型组先升高后下降。参芎化瘀胶囊组所有时间RHOA 和ROCK-Ⅱ 表达均较模型组降低（P<0.05），但仍高于正常组（P<0.05）。结论 参芎化瘀胶囊改善全脑缺血引起的神经元损伤，降低海马CA1 区RHOA 和ROCK-Ⅱ 的表达。Abstract: Objective To investigate the effect of Shenxiong Huayu capsule on the expression of hippocampal CA1 recombinant protein A (small GTP binding protein A, RHOA) and ROCK-2 (RHO associated protein kinase-2, ROCK-Ⅱ). Methods Clean SD male rats (n=96), divided into three groups with 32 rats for each group, gavage was applied 7 days before modeling until the morning of the day to put to death. The groups included the normal control group (normal saline), global cerebral ischemia model group (normal saline) and Shenxiong Huayu capsule+global cerebral ischemia group (Shenxiong Huayu capsule 0.048 g/kg, was dissolved in 0.5 mL double distilled water, once a day, orally 0.3 mL/100 g). Modified Pulsinelli four-vessel occlusion model was constructed in global cerebral ischemia model and Shenxiong Huayu treatment groups and at 1, 3, 7, 14 d after successful modeling, water maze learning test was applied to evaluate the memory abilities of different groups, histopathological changes in HE staining, expression and protein content of RHOA and ROCK-Ⅱ in immunohistochemical staining and Western blot was observed. Results At each time point, escape latency in model group was prolonged (P <0.05) when compared with that in normal control group, and that in Shenxiong Huayu was shorter (P <0.05) than that of model group, but still longer (P <0.05) than that of normal control group. staining showed that, compared with the normal group, model hippocampal CA1 reduced gradually from 1 d to 14 d; an increased survival neurons (P <0.05) in Shenxiong Huayu treatment group at each time points was observed, but still less than that in normal group (P <0.05); immunohistochemistry and Western blot analysis demonstrated that the expression of RHOA and ROCK-Ⅱ in normal control group was not obvious, in model group was decreased after an initial increasing, and that in Shenxiong Huayu treatment group was lower than that of model group (P <0.05), but still higher than that in normal group (P <0.05). Conclusion Shenxiong Huayu capsule improve neuronal damage induced by global ischemia, decreased the expression of hippocampal CA1 region of RHOA and ROCK-Ⅱ.
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