PGC1β对类风湿关节炎破骨细胞PPARγ通路激活的关键调控作用研究

The mechanisms of osteoclast (Oc) differentiation and activation is critical for preventing joint destruction and reducing disability in rheumatoid arthritis (RA). Our results of previous National Natural Science Foundation of China has proved that the activation of PPARγ pathway can inhibit RA-Oc differentiation and activation. However, clinical studies showed that PPARγ agonist therapy for patients with active RA was effective but not ideal, suggesting that there may be other important regulatory factors for PPARγ pathway activation in RA. Recent research found that the activation of PPARγ pathway in osteoclasts depended on PPARγ coactivator 1β (PGC1β) under physiological condition. We found that PGC1β mRNA expression in peripheral blood monocytes (PBMs, regarded as pre-Oc) from RA patients was significantly lower than in those from healthy controls, indicating that PGC1β may play a key regulatory role in PPARγ pathway activation of RA-Oc. To investigate whether the lack of PGC1β could hinder PPARγ agonist from inhibiting RA-Oc differentiation and activation and joint destruction in collagen-induced arthritis (CIA) mice, we will set up Lentiviral- PGC1β-RNAi to transfect PBMs in vitro and generated conditional PGC1β knock-out CIA mice which were deleted PGC1β in hematopoietic lineages. Furtherly, we will apply the luciferase reporter gene technology and chromatin immunoprecipitation assay to identify PGC1β transcriptional signaling pathways in RA-Oc. The aim of this study was to further elucidate the mechanism of osteoclast differentiation, activation and joint destruction in RA, to determine whether PGC1β be a new target preventing RA joint destruction and provide experimental evidence for exploring new drug regulating PGC1β.

弄清破骨细胞（Oc）形成及活化调控机制是阻断类风湿关节炎（RA）骨破坏、降低致残的关键。我们前一个国家自然已证实PPARγ通路激活可抑制RA-Oc形成及活化，临床研究PPARγ激动剂治疗RA有效但不理想，提示可能存在其他重要调控因素。新近发现生理状态下Oc的PPARγ通路激活需共激活分子PGC1β的参与。我们前期研究发现RA患者Oc前体PGC1β表达降低，提示PGC1β可能在RA-Oc的PPARγ通路激活过程中起关键调控作用。本课题拟分别从体外（慢病毒PGC1β-RNAi转染RA-Oc前体细胞）和体内（造血细胞系特异性条件敲除PGC1β小鼠）探讨缺乏PGC1β能否阻碍PPARγ激动剂对RA-Oc形成、活化及CIA小鼠关节骨破坏的抑制作用；再用荧光素酶报告基因技术和ChIP明确PGC1β转录调控的信号通路，旨在明确PGC1β能否成为阻断RA骨破坏的新靶点并为开发调控PGC1β药物提供依据。

类风湿关节炎（RA）是一种常见的以进行性关节破坏为特征的自身免疫性疾病。RA关节破坏的严重程度主要受破骨细胞的数量及功能影响。我们前期已证实过氧化物酶体增殖物激活受体（PPAR）γ通路激活可抑制破骨细胞形成及活化，临床研究发现PPARγ激动剂治疗RA有效但不理想。PPARγ辅激活因子1（PGC1）是一类重要的转录调控因子，广泛参与各种生理过程。条件敲除破骨细胞前体PGC1β的小鼠用PPARγ激动剂干预后，破骨细胞数量、功能及小鼠骨量均无明显改变，提示缺乏PGC1β的破骨细胞PPARγ通路不能被激活，PGC1β在生理状态下破骨细胞形成中具有更重要的调控作用。因此，本项目拟明确PGC1β是否在RA破骨细胞形成和活化过程中起关键调控作用，并阐明PGC1β在RA破骨细胞形成和活化过程中的调控机制。..本研究首先探讨诱导破骨细胞形成的上游通路分子RANKL的调控机制，发现RA成纤维样滑膜细胞（FLS）中高表达PGC1β，并通过上调NF-κB转录促进RANKL的表达。研究成果已发表在Arthritis Research & Therapy杂志上（影响因子：4.269，JCR二区）。然后探讨RA破骨细胞中PGC1β的调控作用及机制。结果发现RA患者外周血单核细胞（破骨细胞前体）表达PGC1β升高且与患者骨破坏程度呈正相关。在RANKL诱导下，高表达的PGC1β可促进体外培养的外周血单核细胞形成成熟破骨细胞。进一步研究发现PGC1β可促进破骨细胞标志基因以及关键信号通路分子NFATc-1的表达。应用NFATc-1活性抑制剂VIVIT干预过表达PGC1β的外周血单核细胞证实PGC1β通过NFATc-1通路促进破骨细胞标志基因的表达。通过免疫共沉淀和荧光素酶报告基因技术发现PGC1β可直接与NFATc-1启动子结合，上调NFATc-1转录表达，阐明了PGC1β促进下游破骨细胞标志基因表达，进而促进破骨细胞形成和活化的具体调控机制。研究结果在欧洲风湿病学年会、南京国际免疫论坛等会议上发言或壁报交流，申请发明专利2项。..本研究结果明确了PGC1β在RA破骨细胞形成和活化过程中发挥关键调控作用，阐明了PGC1β在RA破骨细胞形成和活化过程中的具体调控机制，为临床RA 治疗寻找有效抑制关节破坏的“治本”药提供了新的靶点，具有重要的临床意义及应用前景。