Treg在特应性皮炎样GVHD发病中的优势漂移及分子机制

In early 2013, we reported atopic dermatitis-like graft-versus-host disease (AD-like GVHD) for the first time in the world. It's a novel form of cutaneous GVHD and was characterized by generalized dermatitis, intense pruritus and association with internal organ's rejection. Since it's a new disease, the mechanism has not been studied yet. Regulatory T cells (Tregs) have the properties of maintaining immune homeostasis and suppressing immune response.In our pilot study, we found increased Tregs in the peripheral blood of AD-like GVHD patients and in infiltrates of skin lesions, suggesting that the dominant drift of Tregs be involved in the pathogenesis of the disease. In this study, we aimed to investigate the role of Tregs in AD-like GVHD through a co-cultured system of Tregs and other related cells in vitro, to investigate the phenotypes, cytokines profile, antibody expression and signaling pathway of Tregs by means of cellular biology and molecular biology, and to investigate the effect of Tregs on the patients' skin barrier function (ultrastructure of stratum cornium, filaggrin expression and transepidermal water loss). Furthermore, the interactions between Tregs and other cells will also be studied. This study will help to clarify the possible mechanism of Tregs in the pathogenesis of AD-like GVHD and might help in developing measures for prevention, early recognition and treatment of this novel disease.

我们于2013年在国际上首先报告了一种新的移植物抗宿主病- - 特应性皮炎样GVHD（AD样GVHD）,临床特征包括泛发性皮炎、剧烈瘙痒、伴发内脏排异等，由于是新发现的GVHD类型，因此发病机制的研究还为空白。调节性T细胞（Treg）具有维持免疫自稳和抑制免疫应答的特性，我们在前期研究中发现AD样GVHD患者常有血循环中Treg数量增加，皮损中有Treg细胞浸润，提示在AD样GVHD中Treg发生了优势漂移。本课题拟采用细胞共培养体系，通过分离和培养患者的Treg及相关细胞，构建AD样GVHD体外细胞模型，应用细胞与分子生物学等技术，研究不同条件下Treg的表型、细胞因子分泌模式、抗体表达、信号通路、与其他细胞间的相互作用，及其对患者皮肤屏障功能（角层超微结构、丝聚蛋白表达和经皮水丢失）等的影响，阐明Treg在AD样GVHD发病中的作用机制，为AD样GVHD的预防、早期识别和治疗提供新的思路。

移植物抗宿主病（GVHD）是异基因造血干细胞移植等移植术后常见的并发症，属于难治性疾病。GVHD有多种类型，我们在国际上首先报告了一种新的GVHD亚型——特应性皮炎样GVHD（AD样GVHD）。实验中我们选取了19例AD样GVHD患者、8例扁平苔藓样GVHD（LP样GVHD）、24例AD和15例健康志愿者作为研究对象，利用流式细胞术、免疫荧光法分别检测了外周血、皮肤组织中T细胞亚群的表达，利用ELISA检测血清中、RT-PCR法检测皮肤组织中各细胞因子的水平，利用皮肤组织通过免疫组化、Western Blot、RT-PCR法检测其皮肤屏障功能及相关分子的表达。研究发现与健康对照相比，1. AD样GVHD、AD患者外周血嗜酸性粒细胞升高，而LP样GVHD患者未见外周血嗜酸性粒细胞升高；2. AD样GVHD、AD样GVHD、AD患者血清总IgE升高；3. AD样GVHD、LP样GVHD患者外周血Th2细胞升高、皮肤组织中有较多Th2细胞浸润；4.AD样GVHD患者外周血中Treg细胞升高，皮肤组织中有较多的Treg细胞浸润，且Foxp3蛋白的表达也有升高；5. AD样GVHD、LP样GVHD、AD患者TEWL升高；6.AD样GVHD和LP样GVHD患者皮肤组织中filaggrin、involucrin、loricrin表达增加；7.AD样GVHD、LP样GVHD和AD患者皮肤组织中TSLP表达升高，与TSLP相关的KLK5、PRA2蛋白表达也升高。综上，AD样GVHD和LP样GVHD是以Th2为主的免疫反应，皮肤屏障功能破坏，但是不同的是AD样GVHD中嗜酸性粒细胞和Treg细胞的表达升高，可能与疾病发生发展有关。