IL-22及RANKL在骨性关节炎患者中的表达及调节关系

Expression and regulative relationship of RANKL and IL-22 in osteoarthritis patients

目的 探究骨性关节炎患者体内细胞核因子-κB受体活化因子配体(Receptor activator of nuclear factor-κB ligand,RANKL)和白介素-22(Interleukin-22,IL-22)的表达差异和调节作用以及在骨性关节炎进程中诱导关节软骨退化的可能机制。方法 采用酶联免疫法检测骨性关节炎患者与正常人血清、关节液中的RANKL和IL-22;番红O-固绿复染观察骨性关节炎患者和正常人关节软骨的组织学改变,免疫组化检测关节软骨中RANKL和IL-22的表达情况;RT-PCR检测骨性关节炎患者和正常人关节软骨组织中RANKL、IL-22和基质金属蛋白酶3(Matrix metalloproteinase-3,MMP-3)等基因的表达水平;蛋白质印迹法检测骨性关节炎患者和正常人关节软骨中RANKL、IL-22和MMP3蛋白的含量。用不同浓度的重组人IL-22(Recombinant human IL-22,rhIL-22)处理体外培养的骨性关节炎患者和正常人的关节软骨细胞,采用RT-PCR检测干预后软骨细胞RANKL和MMP-3基因的表达水平。结果 骨性关节炎患者血清、关节液中RANKL和IL-22的含量明显高于正常人;骨性关节炎患者的关节软骨中RNAKL、IL-22和MMP-3的含量和蛋白水平也高于正常人。不同浓度rhIL-22处理后的关节软骨细胞中RANKL和MMP-3的信使RNA表达水平增高且与rhIL-22呈一定的剂量依赖关系。结论 RANKL和IL-22的含量在骨性关节炎患者的血清、关节液和软骨组织中的表达都增高,是骨性关节炎病理生理学过程中导致关节软骨退行性改变的重要细胞因子;IL-22可以上调RANKL的表达,激活其受体RANK并通过p38 MAPK/NF-κB通路促进MMP-3的合成从而引起软骨细胞退行性改变及软骨基质降解,参与骨性关节炎中关节软骨退行性改变的发生和发展;RANKL和IL-22可能是诊断和治疗骨性关节炎的一个新靶点。

Objective To explore the difference of expression levels and adjustment relationship of the receptor activator of nuclear factor-kappa B ligand(RANKL)and interleukin-22(IL-22)in osteoarthritis patients,and the possible mechanisms inducting articular cartilage degeneration in the process of OA.Methods Enzyme-linked immunosorbent assay(ELIAS)was used to detect the concentration of RANKL and IL-22 in OA patients’and normal persons’serum and synovial fluid respectively.Histological changes of articular cartilage of OA patients and normal persons were observed by redyeing of safraninO-fast green and the immunohistochemical staining was used to detect the RANKL and IL-22 expression of the articular cartilage.The gene expression levels of RANKL,IL-22 and matrix metalloproteinases 3(MMP-3)in articular cartilage were separately detected with RT-PCR method in both OA patients and normal person.Then,Western blot method was utilized to detect the protein content of RANKL,IL-22 and MMP-3 in OA patients’and normal persons’articular cartilage.To explore the adjustment relationship of RANKL and IL-22,different concentration gradients of human recombinant IL-22 were added in chondrocytes which were harvested from OA patients and normal persons when cultured in vitro.At last,RT-PCR method was used to detect the RANKL and MMP-3 gene expression levels of the chondrocytes after the intervention of rhIL-22.Results The amount of RANKL and IL-22 in OA patients’serum and synovial fluid was significantly higher than normal persons.OA patients not only had higher gene expression levels of RNAKL and IL-22 than normal persons in the articular cartilage,but also higher protein content levels of RANKL,IL-22 and MMP-3 than normal persons’articular cartilage.The expression levels of RANKL and MMP-3 messenger RNA(mRNA)were obviously increased after chondrocytes had been treated with different concentration gradients of rhIL-22. And there was a certain dose-response relationship between rhIL-22 and RANKL. Conclusion The expression levels of RANKL and IL-22 in OA patients’serum, synovial fluid and cartilage tissues are all increased. It indicated thatboth of them are important cytokines leading to articular cartilage degenerative changes in the pathophysiological process of OA.The expression of RANKL is upregulated by IL-22, and its receptor RANK is activated to promote the synthesis of MMP-3 viathe p38 MAPK/NF-κB pathway. Synthesized MMP-3 can cause chondrocyte and cartilage matrix degradation to contribute tothe occurrence and development of articular cartilage degenerative changes in the OA. All in all, RANKL and IL-22 may be anew target for the diagnosis and treatment of OA.