黄芪葛根联用对KKAy小鼠心脏组织GRP78、CHOP和PUMA表达的影响

Effects of astragalus and puerarin on the expression of GRP78,CHOP and PUMA in heart tissues of KKA mice

目的探讨黄芪注射液联合葛根素注射液通过缓解内质网应激(ERS)、抑制心肌细胞凋亡,改善2型糖尿病(T2DM)动物KKA^(y)小鼠心脏损害,防治糖尿病心肌病的机制。方法 11周龄的KKA^(y)糖尿病模型小鼠按血糖随机分为模型组和黄芪葛根组(黄芪注射液3 mL/kg联合葛根素注射液1.3 mL/kg进行治疗),同周龄雄性C57BL/6J小鼠作为正常对照组,适应性饲养1周后,于12周龄开始实验干预,在20、24和28周龄分批处理小鼠。电镜观察心肌形态改变, RT-PCR及Western Blot检测心脏组织葡萄糖调节蛋白78(GRP78)、C/EBP同源蛋白(C/EBP homologous Protein, CHOP)、p53凋亡上调基因(p53 up-regulate modulator of apoptosis, PUMA)基因和蛋白的表达。结果透射电镜显示,模型组小鼠心肌细胞细胞核、内质网和线粒体肿胀,黄芪葛根组较模型组情况显著改善;RT-PCR和Western Blot结果显示,与正常组相比,3个周龄段模型组GRP78的基因和蛋白表达水平均明显升高,黄芪葛根组GRP78的基因和蛋白表达较模型组均显著下降,差异具有统计学意义(P<0.05)。CHOP和PUMA的基因和蛋白表达,3个周龄段模型组较正常组均增加(P<0.05),24、28周龄黄芪葛根组较模型组均下降((P<0.05)。结论黄芪注射液与葛根素注射液联用对糖尿病小鼠心肌细胞超微结构具有保护作用,可缓解内质网应激,抑制心肌细胞凋亡,这可能是其防治糖尿病心肌病的机制之一。

Objective To investigate the mechanism of astragalus injection combined with puerarin injection in improving the cardiac function of type 2 diabetic(T2 DM) KKA^(y) mice by alleviating endoplasmic reticulum stress(ERS) and inhibiting cardiomyocyte apoptosis. Methods 11-week-old KKA^(y) diabetes model male mice were randomly divided into model group and astragalas and puerarin group(astragalus injection 3 mL/kg combined with puerarin injection 1.3 mL/kg) according to their blood sugar. Male C57 BL/6 J mice of the same age were used as the normal control group. After one week of adaptive feeding, experimental intervention was started at 12 weeks of age, and mice were processed in batches at 20, 24 and 28 weeks of age. The morphological changes of myocardium were observed under electron microscope. RT PCR and Western Blot were used to detect the gene and protein expressions of glucose-regulated protein 78(GRP78), C/EBP homologous protein(CHOP), and p53 up-regulate modulator of apoptosis(PUMA) in heart tissue. Results Transmission electron microscopy showed that the nuclei, endoplasmic reticulum and mitochondria of the model group were swollen;compared with that, those in the astragalus and puerarin group were significantly better. The results of Real-time PCR and Western Blot showed that compared with the normal group, the GRP78 gene and protein expression levels of the 20-, 24-and 28-week-old mice in the model group were significantly higher, and the gene and protein expression of GRP78 in the astragalus and puerarin group were significantly lower than that of the model group(P<0.05). The gene and protein expressions of CHOP and PUMA were increased in the 20-, 24-and 28-week-old mice in the model group, compared with the normal group(P<0.05), and they were both lower in the 24-week-old and 28-week-old astragalus and puerarin groups compared with the model group(P<0.05). Conclusion Astragalus injection combined with puerarin injection seems to have a protective effect on the ultrastructure of diabetic cardiomyocytes. The way it works may be related to alleviating endoplasmic reticulum stress and inhibiting cardiomyocyte apoptosis.