来源：风湿病与关节炎,2020,9（9）:1-6,12.

梅花入骨丹通过Toll样受体4/核转录因子-κB信号通路调节大鼠退变软骨细胞外基质代谢紊乱的作用机制（基础研究）

林家钟¹，黄艳峰²，林　翔¹，刘蔚楠¹，付长龙²，刘献祥²，王荣茂¹

　　【摘　要】目的：探讨梅花入骨丹通过Toll样受体4/核转录因子-κB（TLR4/NK-κB）信号通路调节退变软骨细胞外基质代谢紊乱的作用机制。方法：体外培养大鼠软骨细胞并鉴定，采用CCK-8法检测梅花入骨丹水提物（BCAE）对体外软骨细胞的增殖-毒性，筛选药物作用的最佳浓度和时间。采用10 ng·mL^-1白细胞介素-1β（IL-1β）建立软骨细胞炎症退变模型，将软骨细胞分为空白对照组、模型对照组、BCAE组、抑制剂TAK-242组和抑制剂PDTC组，分别进行相应干预后，qRT-PCR及Western blot法检测各组基质金属蛋白酶-1（MMP-1）、MMP-3、MMP-9、MMP-13、TLR4、IKKβ、IκBα、p-p65、ColⅡ的mRNA、蛋白表达情况。结果：与空白对照组比较，模型对照组MMP-1、MMP-3、MMP-9、MMP-13、TLR4、IKKβ、p-p65的mRNA及蛋白的表达明显升高（P < 0.01），而IκBα、ColⅡ表达下降（P < 0.01）；与模型对照组比较，TAK-242组、PDTC组、BCAE组MMP-1、MMP-3、MMP-9、MMP-13、TLR4、IKKβ、p-p65的mRNA及蛋白的表达明显减少（P < 0.01），而IκBα、ColⅡ表达升高（P < 0.01）。结论：梅花入骨丹通过TLR4/NK-κB信号通路抑制MMPs的关键调控因子表达，从而抑制IL-1β介导的退变软骨细胞外基质代谢紊乱，发挥抗炎和保护软骨细胞的作用。

　　【关键词】　骨关节炎；梅花入骨丹；基质金属蛋白酶；软骨细胞；Toll样受体4；核转录因子-κB；信号通路

Mechanism of Meihua Rugu Dan（梅花入骨丹）Regulating Extracellular Matrix Metabolism Disorder of Degenerative Cartilage in Rats Through TLR4/NK-κB Signaling Pathway

LIN Jia-zhong,HUANG Yan-feng,LIN Xiang,LIU Wei-nan,FU Chang-long,LIU Xian-xiang,WANG Rong-mao

　　【ABSTRACT】Objective:To investigate the mechanism of Meihua Rugu Dan（梅花入骨丹）regulating extracellular matrix metabolism disorder of degenerative cartilage through TLR4/NK-κB signaling pathway.Methods:Rat chondrocytes were cultured in vitro and identified.CCK-8 method was used to detect the proliferation toxicity of BCAE on chondrocytes in vitro,and the optimal concentration and administration time of medicine were screened.Inflammatory degeneration model of chondrocytes were established with 10 ng·mL^-1 of IL-1β.Chondrocytes were divided into a normal group,a model group,a BCAE group,a TAK-242 group and a PDTC group.After intervention,qRT PCR and Western blot methods were performed to detect the mRNA and protein expressions of MMP-1,MMP-3,MMP-9,MMP-13,TLR4,IKKβ,IκBα,p-p65 and ColⅡ.Results:Compared with the normal group,the mRNA and protein expressions of MMP-1,MMP-3,MMP-9,MMP-13,TLR4,IKKβ,p-p65 in the model group significantly increased（P < 0.01）,while the expressions of IκBα and ColⅡ decr-eased（P < 0.01）.Compared with the model group,the mRNA and protein expressions of MMP-1,MMP-3,MMP-9,MMP-13,TLR4,IKKβ,p-p65 in the TAK-242 group,PDTCgroup and BCAE group significantly decreased（P < 0.01）,while the expression of IκBα and ColⅡ increased（P < 0.01）.Conclusion:Meihua Rugu Dan can inhibit the expression of key regulatory factors of MMPs through TLR4/NK-κB signaling pathway,thus inhibiting the metabolic disorder of extracellular matrix of degenerative cartilage mediated by IL-1β,playing an anti-inflammatory role and protecting chondrocytes.

　　【Keywords】 osteoarthritis;Meihua Rugu Dan（梅花入骨丹）;matrix metalloproteinase;chondrocytes;TLR4;NK-κB;signaling pathway