芪棱汤对脑缺血/再灌注大鼠μ-钙依赖蛋白酶-1 mRNA的影响

Effects of Qileng decoction(芪棱汤) on calpain-1 mRNA expression in rats with focal cerebral ischemia-reperfusion injury

目的探讨芪棱汤及其拆方对局灶性脑缺血/再灌注(I/R)损伤大鼠神经元的保护作用,并阐明芪棱汤及其拆方对μ-钙依赖蛋白酶-1(calpain-1)mRNA的调节机制。方法将SD大鼠随机分为正常对照组、假手术组、模型组、芪棱汤去养阴药组、芪棱汤组;后3组又随机分为缺血3 h再灌注6、24、36、48和72 h组,每组4只。运用改良线栓法制备大鼠局灶性脑I/R损伤模型,并进行神经行为学评分。采用原位杂交法观察芪棱汤及其拆方对calpain-1 mRNA表达的影响。结果正常对照组和假手术组神经行为学评分正常,且仅有微量calpain-1 mRNA阳性表达。芪棱汤去养阴药组和芪棱汤组I/R后各时间点大鼠神经行为学评分较模型组显著下降,以芪棱汤组为佳,差异均有统计学意义(P<0.05或P<0.01)。I/R损伤发生后,calpain-1 mRNA表达增高,并在再灌注发生后24 h和48 h达峰值,形成双峰;芪棱汤及其拆方calpain-1 mRNA阳性表达于I/R后各时间点均低于模型组,其中芪棱汤再灌注各时间点的阳性表达又低于相应时间点的芪棱汤去养阴药组(P<0.05或P<0.01)。结论芪棱汤及其拆方可通过抑制calpain-1 mRNA的表达,减少calpain-1的合成,从而减轻脑I/R发生后calpain激活所导致的神经元损伤,发挥神经元保护作用。

Objective To explore the neuron protection of Qileng decoction （QLD,芪棱汤） on rats with focal cerebral ischemia-reperfusion injury, and elucidate its mechanism of regulating the expression of calpain-1 mRNA. Methods Spregue-Dawley（SD） rats were randomly divided into five groups including blank control group, sham-operation control group, model group, QLD group, and Qileng Qu Yangyin decoction （QLQYYD, 芪棱汤去养阴药, a modified QLD in which the nourishing Yin drugs were deleted） group. The rats in the last three groups were subjected to ischemia for 3 hours followed by 6, 24, 36, 48 and 72 hours of reperfusion, there were 4 rats in each group. The modified thread thrombotic method was applied to establish the model of rat with focal cerebral ischemia-reperfusion, and the neurologic scores were evaluated at the different cerebral reperfusion time points. The expression of calpain-1 mRNA was examined by in situ hybridization histochemistry （ISH）. Results The neurographic scores in blank control group and sham-operation control group were normal, and there was very low level of calpain-1 mRNA positive expression in these two groups. The neurographic scores in QLD treatment group and QLDWNY treatment group were significantly decreased compared with the score in the model group, especially better in QLD treatment group （P〈0.05 or P〈0.01）. The level of the calpain-1 mRNA expression in the model group was higher after ischemia-reperfusion injury, and peaked at 24 and 48 hours after reperfusion forming double peaks, the positive expressions in QLD group and QLQYYD group were significantly lower than that in model group, and the level of the calpain-1 mRNA expression in QLD group was lower than that in QLQYYD group at different corresponding time points （P〈0.05 or P〈0.01）. Conclusion QLD and QLQYYD can restrict the synthesis of calpain-1 mRNA after the focal cerebral ischemia-reperfusion injury by restricting the expression of calpain-1 mRNA, thus play a role in protecting the neuron after focal cerebral ischemiareperfusion injury. It proves that both QLD and QLQYYD can protect the neurons after the injury, and QLD has better therapeutic effect than that of QLQYYD.