摘要
目的观察壮骨健膝方含药血清对脂多糖(LPS)诱导的兔A型滑膜细胞肝X受体(LXRs)/核因子-κB(NF-κB)信号通路的影响,探讨壮骨健膝方治疗膝骨关节炎(KOA)滑膜炎症的机制。方法制备壮骨健膝方含药血清并采用超高效液相色谱质谱联用技术(UPLC-MS/MS)进行质量控制(龙胆苦苷885 ng/mL、阿魏酸185 ng/mL)。将兔A型滑膜细胞随机分为空白组和LPS组,LPS组经LPS诱导后建立炎症细胞模型,模型鉴定成功后随机分为模型组、壮骨健膝方组、LXRα抑制剂组、核受体辅助抑制因子(N-CoR)抑制剂组,分别予以10%空白血清、10%含药血清、10%含药血清+LXRα抑制剂、10%含药血清+N-CoR抑制剂,空白组予10%空白血清干预。干预24 h后,收集各组细胞及上清液,Western Blot、RT-qPCR检测各组细胞中LXRα、N-CoR、P50、P65蛋白及m RNA的表达,ELISA检测各组上清液中IL-1β、TNF-α、基质金属蛋白酶-3(MMP-3)及MMP-13含量。结果与空白组比较,模型组P50、P65蛋白与mRNA表达及IL-1β、TNF-α、MMP-3、MMP-13含量均升高(P<0.01),LXRα、N-CoR蛋白与mRNA表达下降(P<0.01)。与模型组比较,壮骨健膝方组P50和P65蛋白与mRNA表达及IL-1β、TNF-α、MMP-3、MMP-13含量均降低(P<0.01),LXRα、N-CoR蛋白与mRNA表达升高(P<0.01);LXRα抑制剂组及N-CoR抑制剂组P50、P65蛋白与mRNA表达及IL-1β、TNF-α、MMP-3、MMP-13含量均降低(P<0.01,P<0.05)。与壮骨健膝方组比较,LXRα抑制剂组及N-CoR抑制剂组LXRα、N-CoR蛋白及mRNA表达下降(P<0.01),P50、P65蛋白及mRNA表达升高(P<0.01,P<0.05),两组IL-1β、TNF-α、MMP-3及MMP-13含量升高(P<0.01,P<0.05)。结论壮骨健膝方可通过上调LXRα和N-CoR蛋白及mRNA表达,下调P50、P65蛋白及mRNA表达,从而抑制LXRs/NF-κB信号通路的传导,减少下游炎症相关指标IL-1β、TNF-α、MMP-3及MMP-13的分泌,最终达到控制膝骨关节炎滑膜炎症反应的作用。
Objective To study the mechanism of Zhuanggu Jianxi Decoction(ZGJXD)on knee osteoarthritis(KOA)synovitis via liver X receptors(LXRs)/nuclear factor-kappa B(NF-κB)signaling pathway expression in Lipopolysaccharide(LPS)-induced rabbit type A synoviocytes by intervention of ZGJXD drugcontaining serum.Methods The drug-containing serum of ZGJXD was prepared and ultra performance liquid chromatography tandem mass spectrometry(UPLC-MS/MS)was used for quality control(gentiopicroside885 ng/mL,ferulic acid 185 ng/mL).The rabbit cells of type A synoviocytes were randomly divided into blank group and LPS group.The LPS group was induced by LPS to establish an inflammatory cell model and then divided randomly into model group,ZGJXD group,LXRαinhibitor group,N-CoR inhibitor group.After being identified,10%blank serum,10%drug-containing serum,10%drug-containing serum+LXRαinhibitor,10%drug-containing serum+N-CoR inhibitor were given respectively.The blank group was given 10%blank serum.The expression of LXRα,N-CoR,P50,P65 protein and mRNA in cell were detected by Western Blot and RTqPCR,IL-1β,TNF-α,MMP-3 and MMP-13 content in the supernatant were detected by ELISA after 24 hours intervention.Results Compared with blank group,the expression of P50,P65 protein and mRNA,the content of IL-1β,TNF-α,MMP-3 and MMP-13 in the model group increased(P<0.01),LXRα,N-CoR protein and mRNA expression decreased(P<0.01).Compared with model group,the expression of P50,P65 protein and mRNA and the content of IL-1β,TNF-α,MMP-3 and MMP-13 in ZGJXD group decreased significantly(P<0.01),LXRα,N-CoR protein and mRNA expression increased(P<0.01);P50 and P65 protein and mRNA expression,the content of IL-1β,TNF-α,MMP-3 and MMP-13 decreased in LXRαinhibitor group and N-CoR inhibitor group(P<0.01,P<0.05).Compared with ZGJXD group,LXRαand N-CoR protein and mRNA expression decreased(P<0.01),P50 and P65 protein and mRNA,the content of IL-1β,TNF-α,MMP-3 and MMP-13increased in LXRαinhibitor group and N-CoR inhibitor group(P<0.01,P<0.05).Conclusio
作者
陈鹏
郭洁梅
肖艳
林震
张英杰
张鹏
李保林
苏友新
CHEN Peng;GUO Jie-mei;XIAO Yan;LIN Zhen;ZHANG Ying-jie;ZHANG Peng;LI Bao-lin;SU You-xin(School of Rehabilitation Medicine,Fujian University of Traditional Chinese Medicine,Fuzhou350122;College of Traditional Chinese Medicine,Fujian University of Traditional Chinese Medicine,Fuzhou350122;Key Laboratory of Orthopedics&Traumatology and Rehabilitation of Traditional Chinese Medicine,Ministry of Education,Fuzhou350122;Medical School of Fujian Health College,Fuzhou,350101)
出处
《中国中西医结合杂志》
CAS
CSCD
北大核心
2022年第9期1087-1094,共8页
Chinese Journal of Integrated Traditional and Western Medicine
基金
国家自然科学基金面上项目(No.81774347)
福建省自然科学基金项目(No.2019J01091)。
