摘要
目的基于胰岛素受体底物2(insulin receptor substrate 2,IRS2)/磷脂酰肌醇-3-激酶(phosphatidylinositol-3-kinase,PI3K)/叉头框蛋白O4(forkhead box O4,FOXO4)信号通路探讨复方仙草颗粒抑制糖尿病肾病(diabetic nephropathy,DN)大鼠肾足细胞上皮-间充质转化(epithelial-mesenchymal transition,EMT)的作用机制。方法采用单侧肾切除、高糖高脂饲料喂养及一次性ip链脲佐菌素制备DN模型大鼠,另设置对照组和假手术组,将造模成功的大鼠随机分为模型组、厄贝沙坦(15.75 mg/kg)组和复方仙草颗粒组低、中、高剂量(315.0、472.5、945.0 mg/kg)组,各给药组ig相应药物,连续6周。检测各组大鼠空腹血糖、肾脏指数(kidney index,KI)、尿微量白蛋白/肌酐比值(urinary albumin-to-creatinine ratio,UACR)、尿素氮(blood urea nitrogen,BUN)水平;采用苏木素-伊红(HE)染色法观察肾组织病理变化;采用透射电镜(TEM)观察肾足细胞超微结构变化;采用qRT-PCR和Western blotting检测大鼠肾组织足细胞裂孔膜蛋白(nephrin)、P-钙黏蛋白(P-cadherin)、α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)、Desmin、成纤维细胞特异蛋白-1(fibroblast specific protein-1,FSP-1)及IRS2/PI3K/FOXO4信号通路相关因子mRNA和蛋白表达。结果与对照组比较,模型组大鼠KI、UACR、BUN水平均显著升高(P<0.05),肾小球肥大,肾小管广泛扩张,上皮细胞变性脱落,足突融合或丢失;肾组织nephrin、P-cadherin表达显著降低(P<0.05),α-SMA、Desmin、FSP-1表达显著升高(P<0.05),IRS2/PI3K/FOXO4信号通路相关因子mRNA和蛋白表达均显著降低(P<0.05)。与模型组比较,复方仙草颗粒组大鼠KI、UACR、BUN水平显著降低(P<0.05),肾脏病理结构和足细胞超微结构明显改善;肾组织nephrin、P-cadherin表达显著升高(P<0.05),α-SMA、Desmin、FSP-1表达显著降低(P<0.05),IRS2/PI3K/FOXO4信号通路相关因子mRNA和蛋白表达均显著升高(P<0.05)。结论复方仙草颗粒能够通过调节IRS2/PI3K/FOXO4信号通路改善DN大鼠肾功能,抑制DN大鼠足细胞EMT,减轻足细胞损伤。
Objective To investigate the mechanism of Compound Xiancao Granules(复方仙草颗粒)on inhibiting epithelial-mesenchymal transition(EMT)of renal podocytes in diabetic nephropathy(DN)rats based on insulin receptor substrate 2(IRS2)/phosphatidylinositol-3-kinase(PI3K)/forkhead box O4(FOXO4)signaling pathway.Methods DN model rats were prepared by unilateral nephrectomy,high sugar and high fat diet and one-time ip streptozocin,control group and sham-operated group were set up.Successfully modeled rats were randomly divided into model group,irbesartan(15.75 mg/kg)group,Compound Xiancao Granules low-,medium-and high-dose(315,472.5,945 mg/kg)groups,and each administration group was ig corresponding drugs for six weeks.Fasting blood glucose,kidney index(KI),urinary albumin-to-creatinine ratio(UACR)and blood urea nitrogen(BUN)of rats in each group were measured;Hematoxylin-eosin(HE)staining was used to observe histopathology changes in renal;Transmission electron microscopy was used to observe ultrastructural changes in renal pedicle cells;qRT-PCR and Western blotting were used to detect mRNA and protein expressions of nephrin,P-cadherin,α-smooth muscle actin(α-SMA),Desmin,fibroblast specific protein-1(FSP-1)and IRS2/PI3K/FOXO4 signaling pathway related factors.Results Compared with control group,KI,UACR and BUN in model group were significantly increased(P<0.05);Glomerular hypertrophy,extensive tubular dilatation,epithelial cell degeneration and shedding,fusion or loss of peduncle were observed;Nephrin and P-cadherin expressions in renal were significantly decreased(P<0.05),α-SMA,Desmin and FSP-1 expressions were significantly increased(P<0.05),mRNA and protein expressions of IRS2/PI3K/FOXO4 signaling pathway related factors were significantly decreased(P<0.05).Compared with model group,KI,UACR and BUN in Compound Xiancao Granules groups were significantly decreased(P<0.05),renal pathological structure and podocyte ultrastructure were significantly improved;Nephrin and P-cadherin expressions in renal were significantly increased(P<0.05),α-SMA,Desmin and FSP-1 expressions were significantly decreased(P<0.05),mRNA and protein expressions of IRS2/PI3K/FOXO4 signaling pathway related factors were significantly increased(P<0.05).Conclusion Compound Xiancao Granules can improve kidney function in DN rats by regulating IRS2/PI3K/FOXO4 signaling pathway,inhibit EMT of podocytes in DN rats and reduce podocyte injury.
作者
刘丹宁
黄国东
杨鑫勇
周嫦艳
王亚南
LIU Dan-ning;HUANG Guo-dong;YANG Xin-yong;ZHOU Chang-yan;WANG Ya-nan(Guangxi University of Chinese Medicine,Nanning 530000,China;Guangxi International ZhuangMedicineHospital,Nanning530000,China)
出处
《中草药》
CAS
CSCD
北大核心
2022年第21期6795-6804,共10页
Chinese Traditional and Herbal Drugs
基金
国家自然科学基金资助项目(81960913)
广西中医药大学2021年研究生教育创新计划项目区级课题(YCSW2021235)。
