间充质干细胞促进GARP表达调控Tregs治疗狼疮的机制研究

Allogeneic mesenchymal stem cells (MSCs) transplantions exhibit therapeutic effects in human systemic lupus erythematosus (SLE) patients and MRL/lpr mice. Regulatory T cells (Tregs) and TGF-β levels in SLE were increased in regulation by MSCs in vitro and in vivo, but the underlying mechanisms remain largely unknown. Glycoprotein A repetitions predominant (GARP) is a transmembrane protein that expresses highly on activated Tregs, promotes the secretion and activation of TGF-β, and controls Foxp3 via a positive feedback loop in Tregs, thus enhances the Tregs’ immunosuppressive function. Our previous study found that MSCs could promote the expression of GARP in peripheral blood mononuclear cells (PBMCs) of SLE patients, but the mechanism of MSCs regulation of GARP is not clear. In this study, we are going to detect the expressions of GARP in SLE and observe the correlation between GARP and disease activities, determine the changes of Tregs before and after MSCs transplantations in MRL/lpr mice or GARP deficient lupus prone mice, survey the effects of MSCs on Tregs in SLE and GARP silent or overexpressed Tregs through co-culture assay, so as to investigate the role of GARP in the pathogenesis and treatment of SLE, and to explore the role and mechanisms of GARP regulation of MSCs in Tregs, and elucidate the specific mechanisms of MSC-induction of host immune tolerance in the treatment of SLE.

异基因间充质干细胞（MSCs）移植治疗系统性红斑狼疮（SLE）疗效显著，体内外均可上调调节性T细胞（Tregs）及TGF-β水平，但机制不明。GARP（glycoprotein A repetitions predominant）高表达于活化Tregs，促进TGF-β分泌与活化，正反馈促进Tregs Foxp3表达，增强Tregs免疫抑制功能。我们研究发现MSCs可促进SLE患者外周血单个核细胞GARP表达，但MSCs调控GARP的机制尚不清楚。本研究通过检测狼疮GARP表达与疾病活动相关性；MRL/lpr狼疮鼠及GARP缺陷狼疮鼠MSCs移植前后Tregs变化；GARP沉默/过表达的狼疮Tregs与MSCs共培养后Tregs细胞及分子水平变化，拟明确GARP在SLE发病及治疗中的作用，探讨GARP在MSCs调节Tregs中的作用及机制，阐明MSCs诱导宿主免疫耐受治疗SLE的具体机制。

异基因间充质干细胞（MSCs）移植治疗系统性红斑狼疮（SLE）疗效显著，体内外均可上调调节性T细胞（Tregs）及TGF-β水平，但机制不明。GARP（glycoprotein A repetitions predominant）高表达于活化Tregs，促进TGF-β分泌与活化，正反馈促进Tregs Foxp3表达，增强Tregs免疫抑制功能。本研究拟明确MSCs是否通过促进GARP表达调控Tregs治疗狼疮。流式细胞术检测SLE患者外周血GARP+Tregs水平，发现GARP+Tregs在SLE患者外周血高于健康对照，非活动期SLE高于活动期SLE。MSCs移植及体外共培养实验发现MSCs体内外对Tregs GARP无显著上调作用，与预实验结果不符。考虑SLE患者外周血T细胞处于高活化状态，而GARP表达与T淋巴细胞活化相关，狼疮Tregs GARP升高可能是机体对自身免疫反馈调节的结果，并非MSCs治疗调节狼疮Tregs的关键分子。文献报道肝激酶B1（LKB1）缺陷影响Tregs数量，增加凋亡相关标志物表达。Tregs LKB1条件性敲除小鼠发生严重自身免疫病，大多数Tregs不表达Foxp3。因此考虑LKB1可能参与调节狼疮T细胞或Tregs发育。实验过程中发现SLE患者及MRL/lpr狼疮鼠CD4+T细胞、CD4+ Naïve T细胞LKB1表达均较健康对照显著下降；MRL/lpr狼疮鼠脾脏CD4+ Naïve T细胞体外诱导iTregs细胞LKB1表达较C57BL/6鼠降低；MSCs移植治疗MRL/lpr狼疮鼠有效，上调Tregs，上调脾脏CD4+ Naïve T细胞LKB1表达及mTor水平；MSCs体外也可上调MRL/lpr狼疮鼠脾脏CD4+ Naïve T细胞LKB1表达。表明CD4+T、CD4+ Naïve T细胞LKB1是狼疮发病及治疗中的一关键分子，MSCs通过上调CD4+T细胞LKB1表达起到一定治疗作用，为临床应用MSCs移植治疗SLE提供了依据。