基于TLR4/MyD88/NF-κB信号通路探讨小青龙汤对寒饮内蕴型慢性肾小球肾炎大鼠的作用机制  

Mechanism of Xiaoqinglong Decoction(小青龙汤)on Model Rats with Chronic Glomerulonephritis with Internal Retention of Cold Fluid through TLR4/MyD88/NF-κB Signal Pathway

作　　者：赖永健 何丽清 储开博 柴希 柴溥苒 LAI Yongjian;HE Liqing;CHU Kaibo;CHAI Xi;CHAI Puran(Shanxi University of Chinese Medicine,Jinzhong 030619,Shanxi,China)

机构地区：[1]山西中医药大学,晋中山西030619

出　　处：《中华中医药学刊》2023年第11期182-187,I0035,I0036,共8页Chinese Archives of Traditional Chinese Medicine

基　　金：国家自然科学基金面上项目(82274365);山西省自然科学基金面上项目(20210302123228);山西中医药大学科技创新团队项目(2022TD1011);山西中医药大学研究生教育创新项目(2022CX019)。

摘　　要：目的探讨小青龙汤基于Toll样受体4/髓样分化因子88/核转录因子-κB(TLR4/MyD88/NF-κB)信号通路对慢性肾小球肾炎(CGN)大鼠的治疗效果及作用机制。方法50只健康Wistar雄性大鼠,随机选8只作为正常组(Normal),对剩余42只大鼠施加低温环境、冰水游泳、冷食冷饮5周,阳离子化牛血清白蛋白(C-BSA)诱导CGN产生。模型制备成功后,随机分为模型组,小青龙汤低剂量组、小青龙汤中剂量组、小青龙汤高剂量组。小青龙汤灌胃15 d后,收集24 h尿液,麻醉后,腹主动脉取血,取两侧肾脏。进行24 h尿蛋白、尿素氮(BUN)、血清白蛋白(ALB)、血清肌酐(Scr)含量测定;酶联免疫吸附测定试验(ELISA)检测血清肿瘤坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)、白细胞介素-4(IL-4)、白细胞介素-6(IL-6)、白细胞介素-8(IL-8)含量;苏木素-伊红(HE)染色进行肾脏病理组织观察;免疫组化(IHC)和蛋白免疫印迹法(Western blot)检测大鼠肾组织TLR4、MyD88、NF-κB p65的蛋白表达;实时荧光定量聚合酶链式反应(Real-time PCR)检测TLR4、MyD88、NF-κB p65 mRNA表达情况。结果与正常组相比,模型组大鼠24 h尿蛋白含量明显升高(P<0.01);血清中BUN、Scr、TNF-α、IL-1β、IL-4、IL-6、IL-8含量明显升高(P<0.01),ALB含量明显降低(P<0.01);模型组大鼠肾组织有炎性细胞浸润,肾小球结构紊乱,肾小球系膜细胞严重增生,肿胀,部分膜细胞脱落,明显病变;与正常组比较,模型组大鼠肾组织中TLR4、MyD88、NF-κB p65 mRNA和蛋白表达均显著增加(P<0.01)。与模型组比较,小青龙汤高剂量组大鼠24 h尿蛋白含量明显降低(P<0.01);小青龙汤中、高剂量组大鼠血清BUN、Scr含量显著降低(P<0.01),小青龙汤低、中、高剂量组ALB含量明显升高(P<0.05,P<0.01);小青龙汤中、高剂量组大鼠血清TNF-α、IL-1β、IL-4、IL-6、IL-8含量显著降低(P<0.01);小青龙汤中、高剂量组较模型组肾小球系膜细胞增生明显减轻,�Objective To investigate the therapeutic effect and mechanism of Xiaoqinglong Decoction(小青龙汤,XQLD)on model rats with chronic glomerulonephritis(CGN)through the Toll-like receptor 4/myeloid differentiation factor 88/nuclear transcription factor-κB(TLR4/MyD88/NF-κB)signaling pathway.Methods Fifty healthy mal Wistare rats were selected.Eight of them were randomly selected as a normal group,and the remaining ones were subjected to low temperature,swimming in ice water,and cold food and drink for five weeks.CGN modeling was induced by cationized bovine serum albumin(C-BSA).After successful modeling,the remaining rats were randomly divided into a model group,a low-dose Xiaoqinglong Decoction group,a medium-dose Xiaoqinglong Decoction group and a high-dose group Xiaoqinglong Decoction group.After gavaging XQLD for 15 days,24 h urine was collected.After anesthesia,the blood from the abdominal aorta and both kidneys were collected.The contents of 24 h urine protein,urea nitrogen(BUN),serum albumin(ALB),serum creatinine(Scr)were determined.Enzyme-linked immunosorbent assay was used to detect the serum tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),interleukin(IL)-4,IL-6 and IL-8.Hematoxylin-eosin staining was used for renal pathological tissue observation.Immunohistochemistry and Western blotting were used to detect the protein expressions of TLR4,MyD88 and NF-κB p65 in kidney tissues of the rats.Real-time polymerase chain reaction was used to detect the mRNA expressions of TLR4,MyD88 and NF-κB p65.Results Compared with those of the normal group,the 24 h urine protein content of rats in the model group was significantly increased(P<0.01).The contents of BUN,Scr,TNF-α,IL-1β,IL-4,IL-6 and IL-8 in serum increased significantly(P<0.01),and ALB content decreased significantly(P<0.01).The kidney tissue of rats in the model group showed inflammatory cell infiltration,disordered glomerular structure,severe proliferation and swelling of mesangial cells,partial shedding of membrane cells and obvious lesions.T

关 键 词：小青龙汤 TLR4/MyD88/NF-κB信号通路 慢性肾小球肾炎 阳离子化牛血清白蛋白 

分 类 号：R285.5[医药卫生—中药学]