TLR2介导的Treg细胞塑型在溃疡性结肠炎肠粘膜Treg/Th17细胞间失衡中的作用机制研究

Ulcerative colitis(UC) is an organ-specifically autoimmune disease with unclear pathogenesis. Intestinal tract plays an important role in the oral tolerance, and Treg cell is the important immune tolerance cell. It is concerned the role of Treg in UC. Recent studies failed to draw consistent conclusions regarding the role of Treg in UC. However, immune imbalance between Treg and Th17 cell is a characteristic of UC. The functional heterogeneity and differentiation dynamics can be clearly shown by separating Treg cells into three subsets based on the expression of FoxP3 and CD45RA. We have found that increased CD45RA-FoxP3lo T cells could explain increased Treg with dysfunction in the patients with SSc, which could secrete IL-17 but without immunosuppressive capacity. Based on these, we could find the reason of Treg/Th17 imbalance and increased Treg with dysfunction of UC. TLR2 stimulation drives human Treg into a Th17-like phenotype with reduced suppressive function. Recently report showed that the productions of TLR2 by lamina propria mononuclear cells from UC patients were increased. Thus, we think activation of TLR2 signal could induce the abnormal of Treg differentiation and immunosuppressive capacity, which is the reason of imbalance between Treg and Th17 cells in UC. In this study, we will evaluate the phenotype and function of Treg subpopulations in all of PBMC, mucosa of terminal ileum and colon on the patients of UC, the effect of activating or blocking TLR2 in the differentiation and immunosuppressive capacity of Treg subpopulation in vitro, and confirm the role of TLR2-/- Treg on the pathogenesis of UC mice. Our subject aimed to demonstrate the reason and mechanism of Treg/Th17 imbalance in UC, and to lay the theoretical foundation of new therapeutic targets.

溃疡性结肠炎（UC）是一种机制不明，根治困难的自身免疫性疾病，严重影响患者的生活质量。具有免疫抑制功能的Treg细胞在其发病中的作用值得关注。Treg和Th17细胞功能和分化上彼此拮抗，而UC患者Treg和Th17细胞同时升高，其原因并不清楚。我们前期对硬皮病患者Treg细胞亚群的研究发现CD45RA-FoxP3low细胞显著增多，该群细胞没有抑制功能却能分泌IL-17。TLR2能够诱导Treg细胞产生IL-17并丧失功能。因此我们假设，TLR2信号介导的Treg亚群异常是造成UC患者Treg和Th17细胞同时升高的原因。本课题拟观察UC患者的肠粘膜细胞中Treg亚群的变化，探讨激活和阻断TLR2信号对Treg亚群的影响，并以小鼠UC模型作为研究对象，观察TLR2-/-Treg细胞在UC发病中的作用，以阐明UC患者Treg/Th17细胞失衡的原因及其机制，并为发现新的治疗靶点奠定理论基础。

研究背景：Treg与Th17细胞的平衡关系被打破是溃疡性结肠炎的一个重要的免疫异常表现，Treg细胞分为三个亚群，其中FrIII亚群没有免疫抑制功能，而且有向Th17分化的潜能。TLR2在Treg的分化中发挥着重要的作用。.研究目的：探讨TLR2介导的Treg细胞亚群的变化在DSS诱导的UC模型鼠中Treg/Th17细胞失衡及溃疡性结肠炎发病中的作用。.研究结果：.1 UC模型鼠结肠病理HE染色可见明显结肠黏膜受损，TLR2抗体可以使UC模型小鼠的病情（DAI评分）及结肠黏膜损伤得到缓解。.2 UC模型鼠在外周血、肠系膜淋巴结、回肠固有层、结肠固有层中，Treg细胞水平均显著升高；而Treg细胞Foxp3 mRNA表达水平和IL-10表达水平均低于正常组。在外周血和结肠固有层黏膜中，UC模型鼠Th17细胞水平显著升高。TLR2抗体可以改善外周血及结肠固有层粘膜中的Treg及Th17细胞水平。UC模型鼠中FrI亚群水平均显著高于正常组，FrII细胞水平均低于正常组，FrIII亚群水平均显著高于正常组；FrIII亚群中Foxp3 mRNA表达水平低于正常组；IL-17A、RORC mRNA表达水平高于正常组。在UC模型鼠的结肠固有层黏膜中Treg细胞、Treg细胞亚群、Th17细胞、CD4+CD25+FoxP3+IL-17A+细胞数量水平高于其他组织中的水平。.3 TLR2抗体可以逆转肠系膜淋巴结及结肠固有层粘膜中FrI、FrII及FrIII的水平。UC模型鼠CD4+CD25+FoxP3+IL-17A+细胞水平高于正常组，TLR2抗体可以改善这一现象。.研究结论：.1.UC模型鼠存在Treg细胞和Th17细胞数量失衡，这一现象尤其突出地表现在结肠粘膜组织中。.2. Treg细胞FrIII亚群细胞存在CD4+Foxp3+IL-17A+双阳细胞特质，可能是Treg细胞和Th17细胞失衡的原因。.3. TLR2抗体可以使UC模型小鼠的病情（DAI评分）及结肠黏膜损伤得到缓解。.4.TLR2抗体有助于通过改善Treg各亚群及CD4+Foxp3+IL-17A+双阳细胞的水平，逆转UC小鼠结肠固有层Treg细胞与Th17细胞的失衡。.科学意义：TLR2介导的Treg细胞塑型异常及Treg/Th17细胞失衡在UC发病中的作用，并进一步为TLR2作为UC治疗的靶点提供理论依据。