基于TLR4/NF-κB、Raf/MEK通路探讨加味升降散抑制糖尿病大鼠肾脏炎症和纤维化的作用机制  

Mechanism of Jiawei Shengjiang Powder(加味升降散) in Inhibiting Renal Inflammation and Fibrosis in Diabetic Rats Based on TLR4/NF-κB and Raf/MEK Pathways

作　　者：张尧 高飞[1,2] 宋瑞婧 张欣欣 毛竞宇 檀金川[1,2] Zhang Yao;Gao Fei;Song Ruijing;Zhang Xinxin;Mao Jingyu;Tan Jinchuan(Hebei University of Chinese Medicine,Shijiazhuang 050200;The First Affiliated Hospital of Hebei University of Chinese Medicine,Hebei Provincial Hospital of Traditional Chinese Medicine,Shijiazhuang 050011)

机构地区：[1]河北中医学院,石家庄050200 [2]河北中医学院第一附属医院河北省中医院,石家庄050011

出　　处：《中药药理与临床》2022年第5期20-27,共8页Pharmacology and Clinics of Chinese Materia Medica

基　　金：河北省自然科学基金面上项目(编号:H2019423037)。

摘　　要：目的:探讨加味升降散对糖尿病肾病(Diabetic nephropathy,DN)大鼠肾脏Raf/MEK通路蛋白的影响及其炎症和纤维化改善的可能机制。方法:DN大鼠随机分为模型对照组、厄贝沙坦0.014 g/kg组、加味升降散4.4、8.7、17.5 g/kg组,另设正常对照组,各组分别灌胃相应药物或生理盐水,1次/d,连续给药8 w。检测大鼠尿蛋白含量,血清中Glu、Bun、Scr含量、白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、C-反应蛋白(CRP)含量;HE、Masson染色和电镜观察肾脏病理变化;免疫组化(IHC)检测肾组织核因子-κB蛋白(NF-κB)、髓样分化因子(MyD88)、Toll样受体4(TLR4)、α-平滑肌肌动蛋白(α-SMA)、转化生长因子-β1(TGF-β1)、E-钙黏附蛋白(E-cadherin)的表达;蛋白免疫印迹法(Western blot)检测肾组织Raf蛋白激酶(Raf)、丝裂原活化细胞外信号调节激酶(MEK)、细胞外调节蛋白激酶(ERK)、核因子κB(NF-κB)蛋白的表达。结果:与正常对照组比较,模型对照组大鼠尿蛋白含量显著升高,血清Glu、Bun、Scr、IL-6、TNF-α、CRP含量显著升高(P<0.01),模型对照组大鼠肾组织细胞产生了病理损伤;与模型对照组比较,各给药组大鼠尿白蛋白含量显著降低,血清Glu、Bun、Scr、IL-6、TNF-α、CRP含量显著降低(P<0.01);在HE、Masson染色和电镜下观察可见给予厄贝沙坦和加味升降散治疗后,肾组织细胞的病理损伤逐渐改善。与正常对照组比较,模型对照组大鼠肾脏NF-κB、MyD88、TLR4、α-SMA、TGF-β1、p-Raf、p-MEK、p-ERK、p-NF-κB蛋白表达明显上调,E-cadherin蛋白表达明显下调(P<0.05);与模型对照组比较,厄贝沙坦组和加味升降散8.7、17.5 g/kg组大鼠肾脏NF-κB、MyD88、TLR4、α-SMA、TGF-β1、p-Raf、p-MEK、p-ERK、p-NF-κB蛋白表达明显下调,E-cadherin表达明显上调(P<0.05)。结论:加味升降散可明显减轻糖尿病肾病大鼠肾组织损害,其机制可能与下调TLR4/NF-κB、Raf-MEK通路蛋白表达,抑制肾脏炎症状�Objective:To investigate the effect of Jiawei Shengjiang Powder on the proteins in the Raf/mitogen-activated protein kinase kinase(MEK)pathway in the kidney of rats with diabetic nephropathy(DN)and its underlying mechanism in improving inflammation and fibrosis.Methods:DN rats were randomly divided into a normal group,a model group,an irbesartan(0.014 g/kg)group,and low-,medium-,and high-dose Jiawei Shengjiang Powder(4.4 g/kg,8.7 g/kg,and 17.5 g/kg)groups.Rats were given corresponding drugs or normal saline by gavage,once a day for 8 weeks.The content of urine protein and the serum levels of Glu,Bun,Scr,interleukin-6(IL-6),tumor necrosis factor-α(TNF-α),and CRP were detected.Hematoxylin-eosin(HE)staining,Masson staining,and transmission electron microscopy(TEM)were used to observe pathological changes in the kidney.Immunohistochemistry(IHC)was used to detect the protein expression of nuclear factor-κB(NF-κB),myeloid differentiation factor 88(MyD88),Toll-like receptor 4(TLR4),α-smooth muscle actin(α-SMA),transforming growth factor-β1(TGF-β1),and E-cadherin in renal tissues.Western blot was used to detect the protein expression of Raf kinase,MEK,extracellular-regulated kinase(ERK),and NF-κB in renal tissues.Results:Compared with the normal group,the model group showed increased content of urine protein,elevated serum levels of Glu,Bun,Scr,IL-6,TNF-α,and CRP(P<0.01),and pathological damage in renal tissues of rats.Compared with the model group,all groups with drug intervention displayed decreased content of urine protein and reduced serum levels of Glu,Bun,Scr,IL-6,TNF-α,and CRP(P<0.01).As revealed by HE staining,Masson staining,and TEM,the pathological damage was improved after treatment with irbesartan and Jiawei Shengjiang Powder.Compared with the normal group,the model group exhibited up-regulated protein expression of NF-κB,MyD88,TLR4,α-SMA,TGF-β1,p-Raf,p-MEK,p-ERK,and p-NF-κB in the kidney and down-regulated expression of E-cadherin(P<0.05).Compared with the model group,the irbesartan group and

关 键 词：加味升降散 糖尿病肾病 有丝分裂原活化蛋白激酶 炎症 纤维化 

分 类 号：R285.5[医药卫生—中药学]