小檗碱改善多囊卵巢综合征伴胰岛素抵抗大鼠子宫组织局部胰岛素抵抗的作用机制研究

Study on the mechanism of berberine in improving local insulin resistance in uterine tissue of rats with polycystic ovary syndrome and insulin resistance

目的探讨小檗碱对多囊卵巢综合征(PCOS)伴胰岛素抵抗(IR)大鼠血清甾体激素和糖脂代谢指标的影响及可能作用机制。方法将60只雌性SD大鼠随机分为空白组、PCOS组、小檗碱组和二甲双胍组,每组15只。PCOS组、小檗碱组和二甲双胍组大鼠采用胰岛素联合人绒毛膜促性腺激素(HCG)建立PCOS伴IR模型。造模成功后,小檗碱组和二甲双胍组分别给予盐酸小檗碱混悬液、二甲双胍溶液灌胃,空白组和PCOS组给予等量生理盐水灌胃,均连续31 d。进行葡萄糖耐量试验(OGTT),检测各组大鼠血清甾体激素和糖脂代谢指标水平,计算胰岛素抵抗指数(HOMA-IR),苏木素-伊红染色(HE)观察大鼠子宫形态,蛋白质印迹(Western blot)法检测子宫组织中IRS1、AKT、GSK3β蛋白表达水平,实时荧光定量PCR(Real-Time PCR)法检测子宫组织中GLUT4 mRNA表达水平。结果与空白组比较,PCOS组大鼠OGTT各时间段血糖水平及血清促黄体生成素(LH)、17-羟孕酮(17-OHP)、总睾酮(T)、雄烯二酮(ASD)、三酰甘油(TG)、总胆固醇(TC)、低密度脂蛋白胆固醇(LDL-C)、空腹胰岛素(FINS)水平和HOMA-IR、LH/卵泡刺激素(FSH)均明显升高(P均<0.05),性激素结合球蛋白(SHBG)和高密度脂蛋白胆固醇(HDL-C)水平均明显降低(P均<0.05),子宫组织肌层厚度、腔上皮细胞高度和腺腔大小均明显增加(P均<0.05);与PCOS组比较,小檗碱组和二甲双胍组大鼠OGTT各时间段血糖水平及血清LH、17-OHP、T、ASD、TG、TC、LDL-C、FINS水平和HOMA-IR、LH/FSH,子宫组织肌层厚度、腔上皮细胞高度、腺腔大小均明显降低(P均<0.05),SHBG水平均明显升高(P均<0.05)。与空白组比较,PCOS组大鼠子宫组织中pIRS1-Ser307和GSK3β蛋白相对表达量均明显升高(P均<0.05),pAKT-S473蛋白和GLUT4 mRNA相对表达量均明显降低(P均<0.05);与PCOS组比较,小檗碱组和二甲双胍组大鼠子宫组织中pIRS1-Ser307和GSK3β蛋白相对表达量均明显降低(P均<0.05),pAKT-S473蛋白和GLUT4 mRNA相对表达量均明显升高(P均<0.05)。结论小檗碱能改善PCOS伴IR大鼠高胰岛素血症、生殖内分泌和糖脂代谢紊乱状态,其可能通过调节子宫胰岛素信号通路IRS1、AKT、GSK3β蛋白和GLUT4 mRNA的表达,改善子宫局部IR,从而改善子宫内膜容受性和子宫异常增生状态。

Objective It is to investigate the effect and possible mechanism of berberine on serum steroid hormones and glucose-lipid metabolism indexes in polycystic ovary syndrome(PCOS)rats with insulin resistance(IR).Methods Sixty female SD rats were randomly divided into blank group,PCOS group,berberine group and metformin group,with 15 rats in each group.The rats in PCOS group,berberine group and metformin group were treated with insulin combined with human chorionic gonadotropin(HCG)to establish models of PCOS with IR.After successful modeling,the berberine group and metformin group were respectively given berberine hydrochloride suspension and metformin solution by gavage,and the blank group and PCOS group were given the same amount of normal saline by gavage,all treated for 31 days.Glucose tolerance test(OGTT)was performed,the levels of serum steroid hormones and glucose-lipid metabolism indexes of the rats in each group were detected,and the insulin resistance index(HOMA-IR)was calculated.Hematoxylin-eosin staining(HE)was used to observe the morphology of rat uterus.The protein expression levels of IRS1,AKT and GSK3βin uterine tissue were detected by Western blot method,and the mRNA expression level of GLUT4 in uterine tissue was detected by real-time quantitative PCR(Real-Time PCR).Results Compared with the blank group,the levels of blood glucose at each time period of OGTT and serum luteinizing hormone(LH),17-hydroxyprogesterone(17-OHP),total testosterone(T),androstenedione(ASD),three acylglycerol(TG),total cholesterol(TC),low-density lipoprotein cholesterol(LDL-C),fasting insulin(FINS),HOMA-IR,LH/follicle stimulating hormone(FSH)of the rats in PCOS group were significantly increased(all P<0.05),the levels of sex hormone-binding globulin(SHBG)and high-density lipoprotein cholesterol(HDL-C)were significantly decreased(all P<0.05),and the thickness of uterine tissue myometrium,the height of luminal epithelial cells and the size of glandular cavity were significantly increased(all P<0.05);compared with the PCOS group,the levels of blood glucose at each time period of OGTT and serum LH,17-OHP,T,ASD,TG,TC,LDL-C,FINS and HOMA-IR,LH/FSH,uterine tissue myometrial thickness,luminal epithelial cell height,and glandular lumen size of the rats in the berberine group and metformin group were significantly decreased(all P<0.05),and SHBG levels were significantly increased(all P<0.05).Compared with the blank group,the relative expressions of pIRS1-Ser307 and GSK3βproteins in the uterine tissue of the PCOS group were significantly increased(all P<0.05),and the relative expressions of pAKT-S473 protein and GLUT4 mRNA were significantly decreased(all P<0.05);compared with the PCOS group,the relative expressions of pIRS1-Ser307 and GSK3βproteins in the uterine tissue of the rats in the berberine group and metformin group were significantly decreased(all P<0.05),and the relative expressions of pAKT-S473 protein and GLUT4 mRNA were significantly increased(all P<0.05).Conclusion Berberine can improve the hyperinsulinemia,reproductive endocrine and glucose and lipid metabolism disorders in rats with PCOS and IR,which may improve the local IR of the uterus by regulating the expressions of IRS1,AKT,GSK3βprotein and GLUT4 mRNA in the uterine insulin signaling pathway,thus to improve endometrial receptivity and uterine dysplasia status.