黄芪甲甙对Balb/C小鼠CVB3病毒性心肌炎心肌细胞凋亡及Bcl-2/Bax基因和蛋白表达的影响

目的]探讨黄芪甲甙对Balb/C小鼠CVB3病毒性心肌炎心肌细胞凋亡及Bcl-2/Bax基因与蛋白表达的影响.方法]Balb/C小鼠50只随机分5组（每组10只）,A组空白对照组：腹腔无菌注射不含病毒的Eagle＇s培养基0.1 mL,30 min后,以生理盐水0.1 mL灌胃,共7 d;B组病毒性心肌炎对照组：小鼠每只腹腔注射0.1 mL内含50%组织感染率（TCID50） 为1×10^5的CVB3病毒Eagle＇s培养基,30 min后,以生理盐水0.1 mL灌胃,共7 d;黄芪甲甙低、中、高剂量干预组（分别为C、D、E组）,在腹腔注射0.1 mL内含TCID50 为1×10^5的CVB3病毒Eagle＇s培养基,30 min后,用黄芪甲甙具体剂量分别为0.07、0.2、0.6 mg/（kg·d）]0.1 mL灌胃,共7 d.采用缺口末端标记法（TUNEL）检测心肌凋亡细胞,免疫组织化学检测Bcl-2、Bax蛋白的表达,反转录聚合酶链反应（RT-PCR）检测Bcl-2、Bax基因的表达,并利用图像分析系统测量平均光密度值进行半定量分析.结果]与A组比较,B组心肌细胞凋亡发生率增高（0.57±0.16vs 0.06±0.02,P〈0.01）;抑制凋亡因子Bcl-2基因（0.52±0.12 vs 0.76±0.11,P〈0.01）及蛋白（6.08±1.15 vs 12.38±3.05, P 〈0.01）表达下降,而促进凋亡因子Bax基因（0.79±0.12 vs 0.61±0.14, P 〈0.01）及蛋白（6.21±1.52 vs 3.01±0.75, P 〈0.01）表达增强,Bcl-2/Bax mRNA比值降低（0.58±0.14vs0.87±0.12,P〈0.05）.与B组比较,E组CVB3病毒性心肌炎心肌细胞的凋亡指数（0.09±0.03vs 0.57±0.16,P〈0.05）降低,Bcl-2基因（0.74±0.12 vs 0.52±0.12,P〈0.05）及蛋白水平（11.82±2.96 vs 6.08±1.15, P 〈0.05）表达增强,Bax基因（0.63±0.13 vs 0.79±0.12,P〈0.05）及蛋白（3.15±0.72 vs 6.21±1.52,P〈0.05）水平表达下降.结论]黄芪甲甙在Balb/C小鼠CVB3病毒性心肌炎中抗凋亡作用机制可能是促进抑制凋亡基因Bcl-2表达,而抑制促凋亡Bax基因表达.

Effect of Astragaloside on the Apoptosis of Cardiocytes and Expressions of Bcl-2/Bac Gene and Protein in Balb/C Micewith CVB3-viral Myocarditis

Objective] To explore the effect of Astragaloside on the apoptosis of cardiocytes and the expressions of Bcl-2/ Bax gene and protein in Balb/C mice with CVB3-viral myocarditis. Methods] Fifty Balb/C mice were randomized into five groups with 10 in each. Normal control group（group A） was given 0.1 mL of Eagle＇s medium without viruses by intraperitoneal injection, and lavaged with 0. lmL saline after 30rain for 7d. Viral myocarditis control group（group B） was injected intraperitoneally with 0. lmL Eagle＇s medi;um containing CVB3-virus with 1 ×l0^5 TCID50 and lavaged with 0.1 mL saline after 30 min for 7 d. Low-dose intervention group（group C）, middle-dose intervention group（group D） and high-dose intervention group（group E） were injected 0. lmL Eagle＇s medium containing CVB3 virus With 1×10^5 TCID50 intraperitoneally, and after 30min, received the lavage of 0.1 mL 0.07 mg/（kg· d）, 0.2 mg/（kg · d） and 0.6 mg/（kg ·d） Astrogaloside for 7 d, respectively. TUNEL was used to detect the apoptosis of cardiocytes, and immunohistochemistry method was used to detect the levels of Bax and Bcl- 2 proteins, and reverse transeription-polymerase chain reaction（RT-PCR） was used to detect the expressions of Bcl-2 and Bax genes, and image analysis system was used to measure the average optical for the semiquantitative analysis. Results] Compared with group A, the apoptosis of eardiocytes in group B was significantly increased（0. 57±0. 16 vs 0. 06±0. 02, P 〈 0.01）, and the expressions of Bcl-2 gene and protein decreased（0.52±0.12 vs 0.76±0.11 and 6.08±1.15 vs 12.38±3.05, respectively, all P 〈0.01）, and the expressions of Bax gene and protein increased（0. 79 ± 0.12 vs 0.61 ± 0.14 and 6.21±1.52 vs 3.01±0.75, respectively, all P 〈0.01）, and the ratio of Bcl-2/Bax mRNA decreased（0.58±0.14 vs 0.87±0. 12, P 〈0. 05）. Compared with group B, the apoptosis index of cardiocytes of CVB3-viral myocarditis in group E reduced（0. 09±0. 03 vs 0.57±0.16, P 〈0.05）, and the expressions of Bcl-2 gene and protein increased（0.74±0.12 vs 0.52±0.12 and 11.82±2. 96 vs 6.08±1.15, respectively, all P G0. 05）, and the expressions of Bax gene and protein reduced（0.63±0. 13 vs 0. 7920. 12 and 3.15±0.72 vs 6.21±1.52, respectively, all P〈0.05）. Conclusion] Astragaloside can reduce the apoptosis of cardiocytes in Balb/C mice with CVB3-viral myocarditis. Astragaloside can promote the expression of BcP2 and inhibit the expression of Bax, which may be its mechanism.