温经通络汤对膝骨关节炎模型小鼠膝关节软骨组织HMGB1/NF-κB信号通路的影响  

Effect of Wenjing Tongluo Decoction(温经通络汤)on HMGB1/NF-κB Signaling Pathway in Knee Articular Cartilage Tissue of Knee Osteoarthritis Model Mice

作　　者：钱佳佳[1] 许奇 许炜民 黄桂成[1] QIAN Jiajia;XU Qi;XU Weimin;HUANG Guicheng(Nanjing University of Chinese Medicine,Naniing,2210023;The First Affiliated Hospital of Anhui University of Chinese Medicine)

机构地区：[1]南京中医药大学,江苏省南京市210023 [2]安徽中医药大学第一附属医院

出　　处：《中医杂志》2023年第9期939-947,共9页Journal of Traditional Chinese Medicine

基　　金：国家自然科学基金(81774341);江苏高校品牌专业建设工程二期项目。

摘　　要：目的探讨温经通络汤治疗膝骨关节炎的可能作用机制。方法将32只小鼠随机分为假手术组,模型组及温经通络汤低剂量组、高剂量组,每组8只。除假手术组外,其余各组采用前交叉韧带横断术(ACLT)法建立膝骨关节炎模型。术后4周,温经通络汤低剂量、高剂量组分别以温经通络汤20、80 g/(kg·d)灌胃,假手术组及模型组给予0.2 ml/10 g生理盐水灌胃,均每天1次,持续4周。采用HE染色及番红O-固绿(S-O)染色法观察各组小鼠软骨组织病理形态,并根据改良Mankin’s评分标准对病理变化进行评分;Western blot法检测各组小鼠关节软骨组织中基质金属蛋白酶13(MMP13)、Ⅱ型胶原(COLⅡ)及高迁移率族蛋白B1(HMGB1)、核因子κB(NF-κB)通路的蛋白表达情况;qPCR法检测COLⅡ、MMP13及NF-κB mRNA表达;免疫组化法观察软骨中诱导型一氧化氮合酶(iNOS)表达;ELISA法检测血清炎症因子前列腺素E2(PGE2)、白细胞介素6(IL-6)、肿瘤坏死因子α(TNF-α)表达。结果与假手术组比较,模型组关节软骨表面破坏明显,软骨层有明显缺损,改良Mankin’s评分提高,软骨组织中p-IκBα/IκBα值、p-p65/p65值、MMP13及HMGB1蛋白表达,MMP13、NF-κB1及NF-κB2 mRNA表达均显著提高,iNOS阳性细胞表达增加,血清炎症因子IL-6、PGE2和TNF-α表达明显升高,且COLⅡ蛋白及mRNA表达明显降低(P<0.05或P<0.01);与模型组比较,温经通络汤高、低剂量组软骨组织病理形态均改善,关节软骨表面破损减少,改良Mankin’s评分降低,软骨组织中p-IκBα/IκBα值、p-p65/p65值、MMP13及HMGB1蛋白表达,MMP13、NF-κB1 mRNA表达,血清炎症因子IL-6、PGE2和TNF-α表达水平均明显降低,iNOS阳性细胞表达减少,COLⅡ蛋白表达升高(P<0.05或P<0.01);与温经通络汤低剂量组比较,温经通络汤高剂量组改良Mankin’s评分,p-IκBα/IκBα值和MMP13蛋白表达,MMP13、NF-κB1、NF-κB2 mRNA,血清炎症因子IL-6、PGE2和TNF-α表达降低,p-p65/p65Objective To explore the possible mechanism of Wenjing Tongluo Decoction(温经通络汤,WTD)in the treatment of knee osteoarthritis(KOA).Methods Thirty-two mice were randomly divided into sham operation group,model group,WTD low-dose group(Group WTD-L)and high-dose group(Group WTD-H),with eight mice in each group.Except for the Group S,the KOA model was established with anterior cruciate ligament transection(ACLT)method.Four weeks after operation,Group WTD-L and Group WTD-H were given 20 and 80 g/(kg·d)of WTD by gavage,respectively,while Group S and Group M were given 0.2 ml/10 g of normal saline by gavage,once a day for 4 weeks.The pathological morphology of the cartilage tissue of mice were evaluated through HE staining and safranin O-fast green(S-O)staining,and the pathological changes were scored according to the modified Mankin system.Protein expression of matrix metalloproteinase 13(MMP13),collagen(COLⅡ),high mobility group box B1(HMGB1),and nuclear factorκB(NF-κB)pathway in articular cartilage of mice were detected by Western blot.The mRNA expression of COLⅡ,MMP13 and NF-κB were detected by qPCR method.Inducible nitric oxide synthase(iNOS)expression in cartilage was observed by immunohistochemistry,and serum inflammatory factors prostaglandin E2(PGE2),interleukin 6(IL-6),and tumor necrosis factorα(TNF-α)were detected by ELISA.Results Compared to those in the sham operation group,the articular cartilage surface in the model group was damaged significantly with obvious defects of cartilage layer,and the modified Mankin score significantly increased;the protein expression of p-IκBα/IκBα,p-p65/p65,MMP13 and HMGB1 in the cartilage tissue,the mRNA expression of MMP13,NF-κB1 and NF-κB2,positive expression of iNOS cells,expression of serum inflammatory factors IL-6,PGE2 and TNF-α increased,while the protein and mRNA expression of COLⅡ significantly decreased(P0.05 or P0.01).Compared to those in the model group,the pathological morphology of cartilage tissues in Group WTD-L and Group WTD-H were

关 键 词：膝骨关节炎 软骨退变 温经通络汤 HMGB1/NF-κB信号通路 炎症因子 

分 类 号：R285.5[医药卫生—中药学] R-332[医药卫生—中医学]