养阴通脑颗粒对大鼠脑缺血/再灌注损伤后Caspase-3、Bcl-2、Bax mRNA表达的影响

Effects of Yangyin Tongnao Granules on the expression of Caspase-3, Bax, Bcl-2 mRNA in rats after cerebral ischemia/reperfusion injury

目的:观察养阴通脑颗粒(YYTN)对缺血/再灌注(I/R)损伤大鼠的保护作用及其可能的作用机制。方法:采用线栓法建立大鼠局灶性脑缺血模型,缺血90min后再灌注,分假手术组、I/R模型组、YYTN高、中、低剂量组,再灌注后24h灌胃给药,连续7d。再灌注后第1、3、7天,采用Longa法进行神经功能评分;缺血后第7天,进行TUNEL神经细胞凋亡染色,并通过实时荧光定量PCR技术分析海马组织中Caspase-3、Bcl-2和Bax mRNA表达的变化,采用ΔΔCt法计算mRNA相对表达水平。结果:术后第3、7天,与I/R模型组相比,YYTN高、中剂量组Longa评分显著降低(P<0.01,P<0.05)。TUNEL染色后,各组海马区均可见TUNEL阳性细胞,与I/R模型组比较,YYTN高、中、低剂量组均能显著降低凋亡细胞平均光密度值(P<0.01,P<0.05);YYTN高剂量组Caspase-3 mRNA相对表达明显降低(P<0.01),YYTN高、中剂量组Bax mRNA相对表达水平明显降低(P<0.01,P<0.05),YYTN各剂量组Bcl-2 mRNA相对表达水平明显升高(P<0.01,P<0.05)。结论:YYTN可明显抑制神经元凋亡,对I/R损伤具有保护作用,其作用机制可能与降低I/R损伤后大鼠海马区Caspase-3和Bax mRNA表达,促进Bcl-2 mRNA表达有关。

Objective： To observe the protective effect of Yangyin Tongnao Granules（YYTN） on ischemia/reperfusion（I/R） injury rats and its possible mechanism. Methods： Focal cerebral ischemia model was induced by occlusion of the right middle cerebral artery for 90 min using the intraluminal filament model（MCAO）. Rats were divided into the sham-operation group, I/R model group, the YYTN high, middle, low dose groups. Medication was performed 24 h after reperfusion once daily for 7 consecutive days. The neurological function was assessed using Longa method at the 1, 3 and 7 after reperfusion. Seven days after reperfusion, TUNEL staining was implemented to detect the apoptosis of nerve cells; Real-time PCR was used to investigate the Caspase-3, Bcl-2 and Bax gene. Based on β-actin as a reference gene, ΔΔCt method was adopted to calculate the relative gene expression. Results： Compared with I/R model group, the neurological function score of YYTN high and middle dose groups were significantly lower at days 3 and 7（P〈0.01, P〈0.05）. After TUNEL staining, the hippocampus of each group was observed in TUNEL positive cells. Compared with the I/R model group, YYTN high, middle and low dose groups could significantly reduce the apoptosis of neurons in the average optical density（P〈0.01, P〈0.05）. Real-Time PCR results showed that, compared with the I/R model group, YYTN high dose group Caspase-3 mRNA expression decreased significantly（P〈0.01）, YYTN high, middle dose groups Bax mRNA expression decreased significantly（P〈0.01, P〈0.05）, each YYTN group Bcl-2 mRNA relative expression level increased significantly（P〈0.01, P〈0.05）. Conclusion： YYTN can inhibit the apoptosis of hippocampus neurons in ischemia reperfusion injury, its mechanism may be related to the decreased mRNA expression of Caspase-3, Bax, and elevating the mRNA expression of Bcl-2 in hippocampus of ischemia reperfusion injury rats.