Tim-3+Tregs通过调节巨噬细胞极化影响急性肺损伤修复的机制研究

T-regulatory cells(Tregs) are considered to play a key role in the resolution and anti-fibrotic stage of acute lung injury (ALI). A few of studies have documented a role of Tregs in the repression of fibrotic diseases by modulating macrophage polarizing into M2c via the anti-inflammatory cytokine IL-10. Whereas the sub-population of Tim3 + Tregs can secrete a large amount of IL-10, the underlying mechanism in the development of ALI remains to be elucidated. Using a clinically relevant two-hit indirect ALI model (first hemorrhage shock priming subsequent to cecal ligation and puncture challenged), we found that adoptive transfer (AT) of splenic wild type (WT)-Tregs induced a significant repression of the indices of lung injury via up -regulating IL-10；The percentage of Tim-3+Tregs was significantly increased in the survivors comparing with non-survivors of ALI patients. Based on those results, we hypothesize that Tim3+Tregs play a pivotal role in promoting the resolution of ALI by inducing M2c cell polarization via IL-10. Therefore, this project is designed to elucidate the molecular mechanism by which Tim3+Tregs regulate the IL-10 mediated macrophage polarization and the repression of inflammation and fibrosis in the late stage of ALI through multiple biological techniques such as ELISA, RT-PCR, flow cytometry and immunohistochemistry. This work would advance our understanding on how Tregs exert their inhibitory effects in life-threatening complications such as ALI and/or ARDS and promote the resolution of iALI in patients.

调节性T细胞（Tregs）在急性肺损伤（ALI）后炎症缓解和抗纤维化过程中起重要作用。Tregs通过分泌IL-10调控巨噬细胞（Mø）向M2c极化，减轻纤维化。共刺激信号分子Tim-3+的Tregs可产生大量IL-10，但该群细胞在ALI中的作用和机制不明。前期研究发现过继输入Tregs增加IL-10分泌，减轻ALI小鼠肺损伤；ALI存活与死亡患者相比，Tregs细胞表达更高的Tim-3。推测Tim-3+Tregs可能通过IL-10诱导M2c细胞产生，促进肺脏修复。本项目拟通过过继输入总Tregs、Tim-3-和Tim-3-Fc预处理Tregs及流式、免疫组化、RT-PCR、Western blotting等技术探讨Tim-3 在促进Tregs细胞免疫调控IL-10诱导M2c极化和促进ALI后炎症消退，减轻肺纤维化中的作用和机制。为促进ALI后肺组织修复，改善预后提供实验依据和治疗靶点。

急性肺损伤（acute lung injury, ALI）及其严重形式急性呼吸窘迫综合征（acute respiratory distress syndrome, ARDS）是导致ICU患者死亡的首要原因。ARDS早期的病理生理改变主要以炎性渗出为主，晚期则以进行性纤维化为特征。尽管近年来对ARDS的认识逐步深入，危重病救治手段也得到快速发展，肺损伤的修复不全和肺纤维化仍然是目前ALI/ARDS患者的主要死因。而导致修复障碍的主要原因是肺损伤后局部炎症的持续存在和继发肺纤维化，但其潜在病理生理分子机制未明。因此探索ALI/ARDS后损伤修复的机制，寻找其防治的新途径和新靶点，是ARDS防治的难点和热点问题。深入开展此领域内的研究工作具有重要的科学意义和临床实用价值。.调节性T细胞（Tregs）在急性肺损伤（ALI）后炎症缓解和抗纤维化过程中起重要作用。Tregs通过分泌IL-10调控巨噬细胞（Mø）向M2c极化，减轻纤维化。共刺激信号分子Tim-3+的Tregs可产生大量IL-10，但该群细胞在ALI中的作用和机制不明。前期研究发现过继输入Tregs增加IL-10分泌，减轻ALI小鼠肺损伤；回顾性分析显示与死亡组相比，ALI存活的病人Tregs表达更高的Tim-3。推测Tim-3+Tregs可能通过IL-10诱导M2c细胞产生，促进肺脏修复。我们首先通过诱导急性肺损伤小鼠模型，比较了两种体外诱导分化的巨噬细胞对急性肺损伤小鼠肺组织炎症消退和纤维化增生的作用。研究结果发现M2（M2a和M2c）细胞均明显减轻急性肺损伤后炎症和纤维增生变化，且M2c作用优于M2a，该作用可能是通过作用于IL-10信号通路实现的。我们进一步通过诱导急性肺损伤小鼠纤维化模型，比较了Tim-3+Tregs和Tim-3-Tregs对急性肺损伤小鼠肺组织炎症消退和纤维化增生的作用。研究结果发现Tim-3+Tregs过继输注明显减轻肺组织纤维化，而Tim-3-Tregs过继输注则无此作用。进一步机制研究发现，Tim-3+Tregs过继输注通过促进肺组织巨噬细胞M2样极化减轻肺损伤和纤维化。研究结果对急性肺损伤的修复、纤维化的防治提供了一个新的思路。