参七糖络丸调控PI3K/Akt通路改善肥胖型糖尿病小鼠肝细胞凋亡的机制研究

    Study on the Mechanism of Shenqi Tangluo Pill Improving Hepatocyte Apoptosis in Obese Diabetes Mice by Regulating PI3K/Akt Pathway

    • 摘要:
      目的 探讨参七糖络丸调控PI3K/Akt通路改善肥胖型糖尿病肝脏细胞凋亡的作用及机制。
      方法 将60只SPF级7周龄db/db小鼠纳入实验,随机分为模型对照组、阳性对照组和参七糖络丸高、中、低剂量组,另选12只同龄db/m小鼠作为空白对照组。阳性对照组每日给予二甲双胍(0.26 g·kg−1)灌胃,参七糖络丸高、中、低剂量组每日分别给予不同浓度参七糖络丸(76、38、19 g·kg−1)灌胃,空白对照组和模型对照组每日给予等量蒸馏水灌胃,持续干预8周。观察小鼠生存状态并测定体质量、空腹血糖(fasting blood glucose,FBG);采用生化试剂盒检测肝脏AST、ALT含量;采用苏木精-伊红染色、油红O染色和TUNEL染色法观察各组肝脏组织病理变化、脂肪堆积及凋亡情况;采用免疫组织化学染色法分析肝脏组织中Bax、Bcl-2蛋白表达水平;Western blotting检测各组小鼠肝组织中PI3K、p-Akt、Caspase-3蛋白表达水平。
      结果 与空白对照组相比,模型对照组小鼠体质量明显升高(P<0.05);FBG显著升高(P<0.05);肝组织中AST、ALT显著增高(P<0.05);肝脏病理及油红O染色显示肝脏脂肪变性程度显著增加,肝细胞结构紊乱;肝脏细胞凋亡率显著升高(P<0.05);肝脏组织中Caspase-3、Bax蛋白表达水平显著升高(P<0.05);PI3K、p-Akt、Bcl-2蛋白表达水平显著降低(P<0.05);Bax/Bcl-2显著升高(P<0.05)。与模型对照组相比,参七糖络丸高、中剂量组小鼠体质量明显降低(P<0.05);FBG显著降低(P<0.05);肝脏细胞凋亡率显著降低(P<0.05);治疗后各组小鼠的肝脏结构损伤程度明显减轻,脂肪变性程度有所减轻;肝组织中AST、ALT显著降低(P<0.05);肝脏组织中Bax、Caspase-3表达水平显著降低(P<0.05);PI3K、p-Akt、Bcl-2表达水平显著升高(P<0.05);Bax/Bcl-2显著降低(P<0.05)。
      结论 参七糖络丸可以改善肥胖型糖尿病小鼠血糖及肝脏脂肪变性,其作用机制可能与调节PI3K/Akt通路抑制肝细胞凋亡有关。

       

      Abstract:
      OBJECTIVE To investigate the effect and mechanism of Shenqi Tangluo pill on improving apoptosis of liver cells in obese diabetes mellitus by regulating PI3K/Akt pathway.
      METHODS The 60 SPF grade 7-week-old db/db mice were included in the experiment and randomly divided into model control group, positive control group and Shenqi Tangluo pill high-dose, medium-dose and low-dose groups. The 12 db/m mice of the same age were selected as blank control group. Positive control group was given metformin(0.26 g·kg−1) by gavage, Shenqi Tangluo pill high, medium and low dose groups were given different concentrations of Shenqi Tangluo pill(76, 38, 19 g·kg−1) by gavage, blank control group and model control group were given the same amount of distilled water by gavage every day, and the intervention lasted for 8 weeks. The living state of the mice was observed and body weight and fasting blood glucose(FBG) were measured. The contents of AST and ALT in liver were detected by biochemical kit. Hematoxylin-eosin(HE), oil red O and TUNEL staining were used to observe the pathological changes, fat accumulation and apoptosis of liver tissues in each group. The expression levels of Bax and Bcl-2 protein in liver tissues were analyzed by immunohistochemical staining(IHC). The expression level of PI3K, p-Akt, Caspase-3 protein in liver tissues of mice in each group was detected by Western blotting.
      RESULTS  Compared with the blank control group, the body weight of mice in the model control group significantly increased(P<0.05); FBG significantly increased(P<0.05); AST and ALT in liver tissue significantly increased(P<0.05); liver pathology and oil red O staining showed a significant increase in the degree of hepatic steatosis and disordered hepatocyte structure; the apoptosis rate of liver cells significantly increased(P<0.05); the expression levels of Caspase-3 and Bax proteins in liver tissue significantly increased(P<0.05); the expression levels of PI3K, p-Akt, and Bcl-2 proteins significantly decreased(P<0.05); and the Bax/Bcl-2 ratio significantly increased(P<0.05). Compared with the model control group, the body weight of mice in the high- and medium-dose groups of Shenqi Tangluo pill significantly decreased(P<0.05); FBG significantly decreased(P<0.05); the apoptosis rate of liver cells significantly decreased(P<0.05); the degree of liver structural damage in each treatment group was significantly reduced, and the extent of fatty degeneration was alleviated; AST and ALT levels in liver tissue significantly decreased(P<0.05); the expression levels of Bax and Caspase-3 in liver tissue significantly decreased(P<0.05); the expression levels of PI3K, p-Akt, and Bcl-2 significantly increased(P<0.05); and the Bax/Bcl-2 ratio significantly decreased(P<0.05).
      CONCLUSION Shenqi Tangluo pill can improve blood sugar and liver steatosis in mice with obesity diabetes mellitus, and its mechanism may be related to regulating PI3K/Akt pathway and inhibiting hepatocyte apoptosis.

       

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