E003调控滑膜细胞凋亡干预CIA大鼠发病的作用及机制研究

A resistance against programmed cell death induced by abnormal expression of apoptosis genes in synovial cell may be an important cause of synovial chronic inflammatory thickening in the patients with rheumatoid arthritis. Our previous studies have shown that E003 (daphnetin) significantly suppressed the immune foot swelling and inflammation of the collagen-induced arthritis (CIA, model of rheumatoid arthritis), alleviated the degeneration of articular cartilage cells and induced the apoptosis of synovial cells in CIA rats. However, the cellular and molecular mechanisms of the E003 acting on CIA rats are still poorly understood. In this study, the E003 will be respectively applied to CIA rat synovial cell and CIA rat model. In the presence and absence of E003, the expression of synovial cell apoptosis genes, apoptosis rate of synovial cell, the expression of mRNA and corresponding protein for signal transduction genes in CIA rats will be tested.In vivo the effect before and after the application of caspase inhibitor on apoptosis signal transduction pathway will be observed. This project from the aspects of aleration at various levels,such as mRNA,protern,cell and signal transduction pathways, will be used to analyze the regulation and its mechanism of E003 on apoptosis of synovial cells to intervene the pathogenesis of CIA rats. This study can provide theoretical and experimental basis for the development of new drugs and treatment of rheumatoid arthritis.

滑膜细胞凋亡基因表达异常导致其凋亡受阻可能是类风湿关节炎患者关节滑膜慢性炎性增厚的重要原因。我们的前期研究发现， E003（即瑞香素）可明显抑制CIA大鼠（类风湿关节炎模型）免疫足肿胀及炎症，缓解关节软骨细胞退化变性,并可诱导其滑膜细胞凋亡。但目前对E003上述效应产生的细胞和分子机制知之甚少。本研究将应用E003作用CIA大鼠关节滑膜细胞和CIA大鼠模型，比较E003作用前后和凋亡信号传导途阻断剂作用前后，CIA大鼠滑膜细胞凋亡基因、信号传导通路基因mRNA和对应蛋白的表达情况，滑膜细胞凋亡率，以及动物体内疗效，从基因、分子、细胞、整体水平以及凋亡信号传导通路等多个层面，解析E003调控滑膜细胞凋亡干预CIA大鼠发病的作用及机制。本研究可为研制类风湿关节炎的新药及治疗提供理论和实验依据。

金边瑞香是我省大余县的支柱产业，其主要活性成分瑞香素（E003），在我们的前期研究中已经证实可明显抑制 CIA 大鼠（类风湿关节炎模型）免疫足肿胀及炎症，缓解关节软骨细胞退化变性，并可诱导其滑膜细胞凋亡。本研究从基因、分子、细胞、整体水平以及凋亡信号传导通路等多个生物学层面，解析瑞香素调控滑膜细胞凋亡干预 CIA 大鼠发病的作用及机制。研究结果表明：瑞香素可通过上调CIA大鼠滑膜细胞中FasL、TNF、Bax、Bid、Cyt-c、Caspase-3、Caspase-8、Caspase-9 mRNA表达以及Bax、Caspase-3、Caspase-8、Caspase-9蛋白表达，降低Bcl-2 mRNA及其蛋白表达，诱导CIA大鼠滑膜细胞发生凋亡，从而达到缓解CIA大鼠关节炎症的作用；同时瑞香素对早、中、晚期CIA大鼠均有不同的抑制炎症作用，且抑制早期炎症效果优于中、晚期。而瑞香素诱导细胞凋亡主要是通过Caspase依赖的死亡受体途径与线粒体途径；且以线粒体途径为主。