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目的观察RNA干扰免疫蛋白酶体亚基低分子重量蛋白2(LMP2)对脑缺血再灌注大鼠血脑屏障结构相关蛋白闭合蛋白(Occludin)和闭锁小带蛋白(ZO~(-1))表达的影响。方法线栓法制作SD大鼠大脑中动脉阻塞再灌注(MCAO)模型,脑缺血1 h再灌注72 h。按照体重将SD大鼠随机分为3组:假手术组、实验组和对照组,每组10只。在MCAO术前1 h,实验组和对照组分别立体定位地脑内注射慢病毒载体LMP2-shRNA和阴性对照shRNA液体。以免疫印迹法分析Occludin、ZO~(-1)蛋白表达,用伊文思蓝(EB)示踪法监测血脑屏障损伤程度。结果 Occludin蛋白与ZO~(-1)蛋白的灰度值:假手术组、对照组和实验组分别1.76±0.09,1.37±0.12;0.24±0.07,0.26±0.07;0.83±0.11,0.74±0.09,与假手术组比较,对照组差异均有统计学意义(均P<0.001);与对照组比较,实验组差异也均有统计学意义(均P<0.001)。脑组织EB含量:假手术组、对照组和实验组分别(3.32±0.14),(11.41±0.21),(6.23±0.28)μg·g~(-1),与假手术组比较,对照组差异有统计学意义(P<0.001);与对照组比较,实验组差异也有统计学意义(P<0.001)。结论沉默LMP2基因表达可显著改善大鼠脑缺血后血脑屏障透通性。
Objective To observe the expression of Occludin and ZO - 1 in blood brain barrier (BBB) of rats with cerebral ischemia reperfusion injury by RNA interference (RNAi) proteasome subunit low molecular weight protein 2 (LMP2) Impact. Methods The middle cerebral artery occlusion and reperfusion (MCAO) model of middle cerebral artery occlusion (SDAO) was established by thread occlusion. SD rats were randomly divided into 3 groups according to body weight: sham operation group, experimental group and control group, 10 rats in each group. At 1 h before MCAO, the experimental group and the control group were stereotaxically injected with lentiviral vector LMP2-shRNA and negative control shRNA liquid respectively. The expression of Occludin and ZO ~ (-1) protein was analyzed by immunoblotting, and the damage of blood-brain barrier was monitored by Evans blue (EB). Results The gray values of Occludin protein and ZO -1 protein were 1.76 ± 0.09, 1.37 ± 0.12, 0.24 ± 0.07, 0.26 ± 0.07, 0.83 ± 0.11, 0.74 ± 0.09, Compared with the sham group, the differences in the control group were statistically significant (both P <0.001); compared with the control group, the experimental group differences were also statistically significant (P <0.001). The content of EB in brain tissue in sham group, control group and experimental group were (3.32 ± 0.14), (11.41 ± 0.21) and (6.23 ± 0.28) μg · g -1, respectively. Compared with sham operation group, There was statistical significance (P <0.001). Compared with the control group, there was also significant difference in the experimental group (P <0.001). Conclusion Silencing LMP2 gene expression can significantly improve the permeability of blood-brain barrier after cerebral ischemia in rats.