STAT3 Mediates the Differential Effects of Oncostatin M and TNFα on RA Synovial Fibroblast and Endothelial Cell Function
STAT3 Mediates the Differential Effects of Oncostatin M and TNFα on RA Synovial Fibroblast and Endothelial Cell Function
复制标题
DOI：
10.3389/fimmu.2019.02056
复制
发表时间：
2019-08-28
影响因子：
7.3
通讯作者：
McGarry, Trudy
中科院分区：
文献类型：
--
作者：
Hanlon, Megan M.;Rakovich, Tatsiana;McGarry, Trudy
目的：抑瘤素M（Oncostatin M，OSM）是一种多效性细胞因子，属于gp 130/IL-6细胞因子家族，与自身免疫性疾病的发病机制密切相关。在这里，我们调查的机制，其协同相互作用与TNF α调节细胞的生物能量学和侵袭性功能的滑膜细胞从患者RheumatoArthritis.Methods：原代RA滑膜成纤维细胞（RAFLS）和人脐静脉内皮细胞（HUVEC）与OSM单独或联合TNF α培养。通过实时PCR和ELISA定量促炎细胞因子、血管生成生长因子和粘附分子。通过Transwell侵袭室、Matrigel管形成测定和粘附结合测定来定量侵袭、血管生成和细胞粘附。使用Seahorse XFe 96分析仪评估细胞生物能量学。关键代谢基因采用实时荧光定量PCR和蛋白质印迹法检测GLUT-1、HK 2、PFKFB 3、HIF 1 α、LDHA、PKM 2和转录因子STAT 3的表达。OSM差异性地调节RAFLS和HUVEC中的促炎介质，与观察到的IL-8和GRO α的抑制相反，IL-6、MCP-1、ICAM-1和VEGF都被显著诱导，对于VCAM-1观察到相反的作用，这取决于细胞类型。在功能上，OSM显著诱导血管生成网络形成、粘附和侵袭机制。这伴随着细胞生物能量谱的变化，其中OSM显著增加了ECAR/OCR比率，有利于糖酵解，这是通过诱导葡萄糖转运蛋白GLUT-1和关键糖酵解酶（HK 2，PFKFB 3，HIF 1 α）实现的。OSM与TNF α协同作用以差异性地调节RAFLS和HUVEC中的促炎机制。有趣的是，OSM差异协同TNF α调节代谢重编程，其中诱导糖酵解活性伴随线粒体呼吸和ATP活性的衰减在RAFLS中得到证实，但在HUVEC中没有。最后，我们确定了一种机制，即OSM与TNF α的组合只在RAFLS中诱导STAT 3的转录活性，在HUVEC.Conclusion中没有观察到任何影响：STAT 3介导OSM和TNF α对RAFLS和EC功能的差异作用。靶向OSM或下游信号通路可能会导致新的潜在治疗或辅助策略，特别是对于那些对TNFi反应不佳的患者。
Objectives: Oncostatin M (OSM), a pleiotropic cytokine and a member of the gp 130/IL-6 cytokine family, has been implicated in the pathogenesis of autoimmune diseases. Here we investigate the mechanisms by which its synergistic interactions with TNF alpha regulate the cellular bioenergetics and invasive function of synovial cells from patients with Rheumatoid Arthritis.Methods: Primary RA synovial fibroblasts (RAFLS) and human umbilical vein endothelial cells (HUVEC) were cultured with OSM alone or in combination with TNF alpha. Pro-inflammatory cytokines, angiogenic growth factors and adhesion molecules were quantified by real-time PCR and ELISA. Invasion, angiogenesis and cellular adhesion were quantified by Transwell invasion chambers, Matrigel tube formation assays, and adhesion binding assays. Cellular bioenergetics was assessed using the Seahorse XFe96 Analyser. Key metabolic genes (GLUT-1, HK2, PFKFB3, HIF1 alpha, LDHA, PKM2) and transcription factor STAT3 were measured using real-time PCR and western blot.Results: OSM differentially regulates pro-inflammatory mediators in RAFLS and HUVEC, with IL-6, MCP-1, ICAM-1, and VEGF all significantly induced, in contrast to the observed inhibition of IL-8 and GRO alpha, with opposing effects observed for VCAM-1 depending on cell type. Functionally, OSM significantly induced angiogenic network formation, adhesion, and invasive mechanisms. This was accompanied by a change in the cellular bioenergetic profile of the cells, where OSM significantly increased the ECAR/OCR ratio in favor of glycolysis, paralleled by induction of the glucose transporter GLUT-1 and key glycolytic enzymes (HK2, PFKFB3, HIF1 alpha). OSM synergizes with TNF alpha to differentially regulate pro-inflammatory mechanisms in RAFLS and HUVEC. Interestingly, OSM differentially synergizes with TNF alpha to regulate metabolic reprogramming, where induction of glycolytic activity with concomitant attenuation of mitochondrial respiration and ATP activity was demonstrated in RAFLS but not in HUVEC. Finally, we identified a mechanism, whereby the combination of OSM with TNF alpha induces transcriptional activity of STAT3 only in RAFLS, with no effect observed in HUVEC.Conclusion: STAT3 mediates the differential effects of OSM and TNF alpha on RAFLS and EC function. Targeting OSM or downstream signaling pathways may lead to new potential therapeutic or adjuvant strategies, particularly for those patients who have sub-optimal responses to TNFi.