黄芪甲苷对链脲佐菌素诱导糖尿病肾病大鼠的作用机制

Mechanism of Astragaloside Ⅳ on diabetic nephropathy induced by Streptozotocin in rats

目的探讨黄芪甲苷(AS-Ⅳ)对链脲佐菌素(STZ)诱导糖尿病肾病(DN)大鼠的作用机制。方法选择SPF级雄性6周龄SD大鼠30只,随机取6只大鼠作为对照组,剩余24只大鼠按65 mg/kg腹腔注射STZ(溶于0.1 mol/L柠檬酸缓冲液,pH 4.4)制备DN模型,对照组腹腔注射等体积0.1 mol/L柠檬酸缓冲液,将模型大鼠随机分为模型组、AS-Ⅳ组、胰岛素(INS)组、AS-Ⅳ+INS组各6只,分别给予AS-Ⅳ10 mg/kg、INS 5 U、AS-Ⅳ10 mg/kg+INS 5 U灌胃,对照组和模型组灌胃等体积1%羧甲基纤维素钠溶液,1次/d,干预8周。对比各组血糖、体质量、尿蛋白、BUN、Scr、HbA1c,血清和肾组织一氧化氮(NO)水平,肾组织中内皮型一氧化氮合酶(eNOS)、磷酸化一氧化氮合成酶丝氨酸1177位点[peNOS(Ser1177)]和苏氨酸495位点[p-eNOS(Thr495)]蛋白表达。结果与对照组比较,模型组血糖均升高(P<0.05);AS-Ⅳ组、INS组、AS-Ⅳ+INS组从第8周开始血糖值均低于模型组(P<0.05);但AS-Ⅳ组、INS组、AS-Ⅳ+INS组间血糖比较差异无统计学意义(P>0.05)。与对照组比较,所有造模大鼠体质量低(P<0.05),尿蛋白、BUN、Scr、HbA1c高(P<0.05);与模型组比较,AS-Ⅳ组、INS组、AS-Ⅳ+INS组体质量高(P<0.05),尿蛋白、BUN、Scr、HbA1c低(P<0.05);与INS组比较,AS-Ⅳ+INS组尿蛋白、Scr低(P<0.05)。与对照组比较,模型组血清和肾组织NO水平低(P<0.05);与模型组比较,AS-Ⅳ组、INS组、AS-Ⅳ+INS组血清和肾组织NO水平高(P<0.05);与INS组比较,AS-Ⅳ+INS组肾组织NO水平高(P<0.05)。与对照组比较,模型组肾组织中p-eNOS(Ser1177)蛋白表达低(P<0.01);与模型组比较,AS-Ⅳ组、AS-Ⅳ+INS组肾组织中p-eNOS(Ser1177)蛋白表达高(P<0.01)。结论黄芪甲苷可能通过eNOS/NO通路对STZ诱导DN大鼠发挥治疗,且作用位点可能在Ser1177。

Objective To explore the protective mechanism of astragalosideⅣ(AS-Ⅳ)through endothelial nitric oxide synthase(eNOS)/nitric oxide(NO)pathway in kidney of diabetic rats.Methods Thirty SPF male SD rats aged 6 weeks were selected.Except of the control group(n=6),the remaining 24 rats were intraperitoneally injected with 65 mg/kg STZ(dissolved in 0.1 mol/L citric acid buffer,pH 4.4)to make DN model,while the control group was intraperitoneally injected with the same volume of 0.1 mol/L citric acid buffer.The model rats were randomly divided into model group,AS-Ⅳgroup,insulin(INS)group and AS-Ⅳ+INS group with 6 rats in each group.AS-Ⅳ10 mg/kg,INS 5 U and AS-Ⅳ10 mg/kg+INS 5 U were given by gavage respectively,and the control group and the model group were given the same volume of 1%sodium carboxymethyl cellulose solution once a day for 8 weeks.The blood sugar,body weight,urine protein,BUN,Scr,HbA1c,nitric oxide(NO)levels in serum and kidney,and the sites of endothelial nitric oxide synthase(eNOS),phosphorylated nitric oxide synthase serine 1177[P-Enos(SER 1177)]and threonine 495[P-Enos(THR 495)]in kidney were compared.Results Compared with the control group,the blood sugar in the model group was increased(P<0.05).The blood sugar levels in As-ⅳgroup,INS group and As-ⅳ+INS group were lower than those in the model group from the 8th week(P<0.05).However,there was no significant difference in blood glucose among As-ⅳgroup,INS group and As-ⅳ+INS group(P>0.05).Compared with the control group,the body weight of the model group was lower(P<0.05),and the urine protein,BUN,Scr and HbA1c were higher(P<0.05).Compared with the model group,AS-Ⅳgroup,INS group and AS-Ⅳ+INS group had higher body mass(P<0.05),but lower urine protein,BUN,Scr and HbA1c(P<0.05).Compared with INS group,urinary protein and Scr in AS-ⅳ+INS group were lower(P<0.05).Compared with the control group,the level of NO in serum and kidney tissue in the model group was lower(P<0.05).Compared with the model group,AS-Ⅳgroup,INS group and AS-Ⅳ+INS group had higher levels of NO in serum and kidney tissue(P<0.05).Compared with INS group,the level of NO in renal tissue in AS-ⅳ+INS group was higher(P<0.05).Compared with the control group,the expression of p-eNOS(Ser1177)protein in renal tissue of model group was lower(P<0.01).Compared with the model group,the expression of p-eNOS(Ser1177)protein in renal tissue of As-Ⅳgroup and As-Ⅳ+INS group was higher(P<0.01).Conclusion AstragalosideⅣmay play a role in treating STZ-induced DN rats through eNOS/NO pathway,and the site of action may be Ser1177.