黄连素在脊髓损伤中对线粒体的保护作用及机制

目的探讨黄连素对脊髓损伤(SCI)后线粒体氧化损伤的作用和可能机制。方法将36只C57小鼠随机分为假手术组、SCI组(伤后立即腹腔注射10 mg/kg生理盐水)和黄连素组(SCI后立即腹腔注射10 mg/kg黄连素),每组12只。使用PSI-IH脊髓打击器建立小鼠SCI模型,于损伤后24 h处死小鼠,取脊髓组织。使用全自动酶标仪检测各组小鼠脊髓组织线粒体内丙二醛(MDA)、还原型谷胱甘肽(GSH)和超氧化物歧化酶(SOD)的变化;用蛋白质印迹法检测脊髓组织caspase-3、cleaved caspase-3的表达及细胞质内和线粒体内细胞色素C(Cyt C)的表达;用免疫荧光双标染色法检测脊髓组织中神经细胞凋亡情况。结果与假手术组相比,SCI组小鼠脊髓组织线粒体内MDA水平升高,GSH、SOD水平降低;细胞质内Cyt C和脊髓组织中caspase-3、cleaved caspase-3表达水平增高,线粒体内Cyt C表达水平降低;脊髓组织中神经元凋亡比例增高;差异均有统计学意义(P 0.05)。与SCI组相比,黄连素组小鼠脊髓组织线粒体内MDA水平降低,SOD和GSH水平增高;细胞质内Cyt C和脊髓组织中caspase-3、cleaved caspase-3表达水平降低,线粒体内Cyt C表达水平增高;脊髓组织中神经细胞凋亡比例减少;差异均有统计学意义(P 0.05)。结论黄连素可减轻SCI小鼠脊髓组织中神经细胞凋亡,这可能与其抑制线粒体氧化损伤、减少Cyt C释放、降低凋亡蛋白表达有关。

Protective effect and mechanism of berberine on mitochondria in spinal cord injury

Objective To investigate the effects and possible mechanisms of berberine on the oxidative damage of mitochondria after spinal cord injury (SCI). Methods Thirty-six C57 mice were randomly divided into 3 groups (n=12):sham group, SCI group (immediately received 10 mg/kg saline injection intraperitoneally once post injury) and berberine group (immediately received 10 mg/kg berberine injection intraperitoneally once post injury). SCI model was established using the PSI-IH spinal cord impactor. The mice were sacrificed and the spinal cord tissue was taken out 24 h after injury. Malondialdehyde (MDA), reduced glutathione (GSH) and superoxide dismutase (SOD) in spinal cord mitochondria in each group were measured by automatic enzyme-linked immunosorbent assay. Cytochrome C (Cyt C) expression in cytoplasm and mitochondria and expressions of caspase-3 and cleaved caspase-3 in the spinal cord were analyzed by Western blotting. The neural apoptosis in the spinal cord was detected by immunofluorescent double labeling. Results Compared with the sham group, the level of MDA increased, while the levels of SOD and GSH decreased in the SCI group; the expression level of Cyt C in mitochondria decreased, while the expression of Cyt C in the cytoplasm, and the expressions of caspase-3 and cleaved caspase-3 in the spinal cord increased in the SCI group; the apoptosis rate of spinal neurons increased in the SCI group; the differences were all statistically significant(P<0.05). Compared with the SCI group, the level of MDA decreased, and the level of SOD and GSH increased in the berberine group; the level of Cyt C in mitochondria increased, and the level of Cyt C in the cytoplasm, and the levels of caspase-3 and cleaved caspase-3 in the spinal cord decreased in the berberine group; the apoptosis rate of the spinal neurons decreased; the differences were all statistically significant(P<0.05). Conclusion Berberine can inhibit the neuron apoptosis in SCI mice, which may be due to inhibition of mitochondrial oxidative damage, decrease of Cyt C releaseand activation of apoptotic molecules.