黄连温胆汤调控SCFAs-GPR41/43-GLP1信号通路干预2型糖尿病模型大鼠的作用机制

Mechanism and Intervention Effect of Huanglian Wendan Tang(黄连温胆汤)on Type 2 Diabetes Mellitus in Rats by Regulating SCFAs-GPR41/43-GLP1 Signaling Pathway

目的:基于SCFAs-GPR41/43-GLP1信号通路研究黄连温胆汤对2型糖尿病模型大鼠的干预机制。方法:选择SPF级SD雄性大鼠80只,先随机分10只为正常对照组,其余大鼠以高糖高脂乳剂10 mL/kg灌胃,监测血糖1次/w, 8 w后,腹腔注射STZ 35 mg/kg诱导2型糖尿病模型,5 d后选取空腹血糖值大于11.10 mmol/L造模成功的大鼠随机分为模型对照组、二甲双胍0.25 g/kg组和黄连温胆汤3、6、12 g/kg组。各组灌胃给予相应的药物或生理盐水,1次/d,连续给药4 w后,取材检测。生化试剂检测大鼠葡萄糖耐量(OGTT)、血脂全套指标;气相色谱法测定大鼠结肠内容物短链脂肪酸(SCFAs)含量;ELISA法检测大鼠结肠组织GLP1、PYY的含量;RT-qPCR法检测大鼠结肠组织G蛋白偶联受体41(Gpr41)、Gpr43、胰高血糖素样肽-1(Glp1)、肽YY(Pyy) mRNA的表达;Western blot法检测大鼠结肠组织GPR41、GLP1、PYY蛋白表达。结果:与正常对照组相比,模型对照组大鼠OGTT试验中血糖含量明显升高,血脂中TC、TG、LDL-C、HbA1c含量显著升高,HDL-C含量显著下降(P<0.01);结肠内容物SCFAs中乙酸、丙酸、正丁酸的含量均显著下降(P<0.01);结肠组织GLP1、PYY含量显著降低,Gpr41、Gpr43、Glp1、Pyy mRNA和GPR41、GLP1、PYY蛋白表达显著下调(P<0.01);与模型对照组比较,黄连温胆汤3、6、12 g/kg组和二甲双胍0.25 g/kg组大鼠OGTT血糖含量明显降低,血脂中TC、TG、LDL-C、HbA1c含量明显降低,HDL-C含量明显升高(P<0.05或P<0.01);结肠内容物乙酸、丙酸、正丁酸的含量明显升高(P<0.05或P<0.01);结肠组织GLP1、PYY含量明显升高,Gpr41、Gpr43、Glp1、Pyy mRNA和GPR41、GLP1、PYY蛋白表达明显上调(P<0.05或P<0.01)。结论:黄连温胆汤抗2型糖尿病作用机制可能与增加SCFAs,调控SCFAs-GPR41/43-GLP1信号通路,促进GLP1、PYY的分泌,降低体内血糖、血脂,改善机体胰岛素抵抗相关。

Objective:To study the intervention effect and mechanism of Huanglian Wendan Tang(黄连温胆汤)on type 2 Diabetes Mellitus(T2DM)in rats through SCFAs-GPR41/43-GLP1 signaling pathway.Methods:A total of 80 male SD rats with SPF grades were selected.10 rats were randomly divided into the normal control group,and the other rats were fed with high-sugar and high-fat emulsions of 10 mL/kg.Their blood glucose was monitored once a week.After eight weeks,the models of T2DM were established by intraperitoneal injection of 35 mg/kg STZ.Five days after modeling,rats with fasting blood glucose(FBG)value more than 11.10 mmol/L were selected and randomly divided into the model control group,group with 0.25 g/kg metformin,and groups with 12,6,and 3 g/kg Huanglian Wendan Tang.Rats were given corresponding drugs or normal saline once a day for four weeks,and then the samples were taken for detection.The oral glucose toler-ance test(OGTT)and a full set of blood lipids in rats were detected by a biochemical test.The levels of short-chain fatty acids(SCFAs)in the colon contents of rats were detected by gas chromatography.The mRNA levels of G-protein-coupled receptors 41 and 43(Gpr41 and Gpr43),glucagon-like peptide-1(Glp1),and peptide YY(Pyy)in the colon tissue of rats were detected by RT-qPCR.The levels of GLP1 and PYY in the colon tissue of rats were detected by enzyme-linked immunosorbent assay(ELISA).Western blot(WB)was used to determine the protein expression of GPR41,GLP1,and PYY in the colon tissue of rats.Results:Compared with that of the normal control group,the blood glucose level in the OGTT of the model control group rats was significantly increased,and the levels of TC,TG,LDL-C,and HbA1c in blood lipids were significantly increased,while the level of HDL-C was significantly reduced(P<0.01).In colon contents,the levels of acetic acid,propionic acid,and ethylacetic acid of SCFAs were significantly decreased(P<0.01).The mRNA levels of Gpr41,Gpr43,Glp1,Pyy,the protein expressions of GPR41,GLP1,and PYY,and levels of GLP1 and PYY in colon tissue were significantly decreased(P<0.01).Compared with that of the model control group,the blood glucose content in the OGTT of the rat groups with 12,6,and 3 g/kg Huanglian Wendan Tang and 0.25 g/kg metformin were significantly reduced,and the levels of TC,TG,LDL-C,and HbA1c in blood lipids were significantly reduced,while the level of HDL-C was significantly increased(P<0.05 or P<0.01).In colon contents,the levels of acetic acid,propionic acid,and ethylacetic acid were significantly increased(P<0.05 or P<0.01).The levels of GLP1 and PYY,the mRNA levels of Gpr41,Gpr43,Glp1,and Pyy,and the protein expressions of GPR41,GLP1,and PYY in colon tissue were significantly upregulated(P<0.05 or P<0.01).Conclusions:The anti-type 2 diabetes mellitus effect of Huanglian Wendan Tang may be related to increasing SCFAs,regulating SCFAs-GPR41/43-GLP1 signalING pathway,promoting GLP1 and PYY productions,reducing the levels of blood glucose and lipids,and improving insulin resistance.