Golinski Marie-Laure, Vandhuick Thibault, Derambure Céline, Fréret Manuel, Lecuyer Matthieu, Guillou Clément, Hiron Martine, Boyer Olivier, Le Loët Xavier, Vittecoq Olivier, Lequerré Thierry
INSERM, U905 & Normandy University, Institute for Research and Innovation in Biomedicine (IRIB), Rouen, France.
INSERM U905, Université de Rouen, Faculté de médecine - pharmacie, 22 boulevard Gambetta, 76000, Rouen, France.
Arthritis Res Ther. 2015 Dec 26;17:382. doi: 10.1186/s13075-015-0894-9.
B and T cells play a key role in rheumatoid arthritis (RA) pathophysiology. RasGRP1 and RasGRP3 are involved in T and B cell receptors signaling, and belong to gene combination able to predict infliximab responsiveness, leading to the question of RasGRP1 and RasGRP3 involvement in RA.
RasGRP1 and RasGRP3 expression levels were measured by qRT-PCR and/or western-blot in peripheral blood mononuclear cells (PBMCs), in T and B cells from untreated RA patients and in RA patients treated by TNFα inhibitors. T and B cells from healthy controls (HC) were cultured with TNFα, and TNFα receptors neutralizing antibodies to highlight the TNFα effects on RasGRP1 and RasGRP3 pathways. MAPK pathways and apoptosis were respectively analyzed using the Proteome Profiler arrays and flow cytometry.
In PBMCs from RA patients, gene expression levels of RasGRP1 were invariant while RasGRP3 was downregulated under TNFα inhibitors and upregulated under TNFα. In T cells from RA patients, RasGRP1 was decreased and its gene expression level was correlated with disease activity. In T cells from HC, TNFα stimulation increased RasGRP1 gene expression level while it reduced RasGRP1 protein expression level. Bryostatin-1 experiments have confirmed that the TNFα effect observed on T cells proliferation was due to the decrease of RasGRP1 expression. Besides, RasGRP3 expression level increased in PBMCs from RA patients under TNFα and in B cells from HC leading us to conclude that RasGRP3 in B cells was modulated by TNFα.
This study demonstrates RasGRP1 dysregulation in RA patients while RasGRP3 is characterized as a biomarker linked to TNFα inhibitors. After binding to TNFR1, TNFα reduced RasGRP1 protein expression resulting in inhibition of T cell activation.
Clinicaltrials.gov NCT00234234 , registered 04 November 2008; NCT00767325 , registered 05 October 2005.
B细胞和T细胞在类风湿关节炎（RA）的病理生理学中起关键作用。RasGRP1和RasGRP3参与T细胞和B细胞受体信号传导，属于能够预测英夫利昔单抗反应性的基因组合，这引发了RasGRP1和RasGRP3是否参与RA的问题。
通过qRT-PCR和/或蛋白质印迹法检测未治疗的RA患者外周血单个核细胞（PBMC）、T细胞和B细胞以及接受TNFα抑制剂治疗的RA患者中RasGRP1和RasGRP3的表达水平。将健康对照（HC）的T细胞和B细胞与TNFα及TNFα受体中和抗体一起培养，以突出TNFα对RasGRP1和RasGRP3途径的影响。分别使用蛋白质组分析阵列和流式细胞术分析MAPK途径和细胞凋亡。
在RA患者的PBMC中，RasGRP1的基因表达水平不变，而RasGRP3在TNFα抑制剂作用下下调，在TNFα作用下上调。在RA患者的T细胞中，RasGRP1减少，其基因表达水平与疾病活动相关。在HC的T细胞中，TNFα刺激增加了RasGRP1基因表达水平，同时降低了RasGRP1蛋白表达水平。苔藓抑素-1实验证实，观察到的TNFα对T细胞增殖的影响是由于RasGRP1表达的降低。此外，在TNFα作用下RA患者的PBMC以及HC的B细胞中RasGRP3表达水平升高，这使我们得出结论，B细胞中的RasGRP3受TNFα调节。
本研究证明RA患者中RasGRP1失调，而RasGRP3被表征为与TNFα抑制剂相关的生物标志物。TNFα与TNFR1结合后，降低了RasGRP1蛋白表达，导致T细胞活化受到抑制。
Clinicaltrials.gov NCT00234234，于2008年11月4日注册；NCT00767325，于2005年10月5日注册。
