TLRs信号的负性调控分子TAM受体在炎症性肠病中的作用和机制研究

Excessive activation of TLRs signaling plays a crucial role in inflammatory bowel diseases (IBD), but the molecule mechanism of this process is obscure, suppress its excessive activation has become a hot topic in the research of IBD. TAM receptors are negative regulators of TLRs, the combine of TAM and its ligand could suppress the activation of TLRs signaling pathway, and leading to a suppression of the inflammation ultimately. It has been reported that mice lack of TAM receptors can developed more severe colitis than normal. Our previous study found that SIGIRR, a negative regulatory factors of TLRs, has therapeutic effects on IBD. Whether TAM receptors, also act as a TLRs negative regulatory factors, have the same effect, and what’s the possible mechanism. The aims of this project included as follow: 1. Explore the correlation of TAM receptors and TLRs signaling pathway in the development of IBD. 2. Investigate whether the TAM ligand and the exogenous gene encoding TAM receptors have therapeutic effects on IBD and their regulate effects on TLR signaling pathway, through MCS-NPs macrophage targeting delivery system. 3. Further elucidate the mechanisms of TAM signaling on TLRs signaling pathway in macrophage through transcriptome technology.

TLRs信号过度激活在炎症性肠病（IBD）的发病中起重要作用，但其机制不明，抑制它的过度激活是目前IBD防治研究中的方向和热点。TAM受体是TLRs信号的负性调控分子，其与配体结合可抑制TLRs信号激活，最终抑制炎症反应，有研究报道TAM受体缺失小鼠可发生更为严重的结肠炎。我们的前期研究发现TLRs信号通路的负性调控分子SIGIRR对IBD有治疗作用，那么同为TLRs信号通路负性调控的TAM受体是否对IBD也有治疗作用？其作用机制如何？目前尚无报道。为此本项目拟研究：①IBD发病中TAM受体信号和TLRs信号通路的变化及其与疾病的关系；②采用MCS-NPs巨噬细胞靶向递药系统，研究给予编码TAM受体的外源性基因和TAM配体对IBD是否有治疗作用及其对TLRs信号的调控；③采用转录组学技术进一步研究TAM受体信号对巨噬细胞TLRs信号通路负调控的作用机制。为IBD的防治提供新的靶点和方向。

IBD是一病因不明的慢性肠道炎症性疾病，巨噬细胞及TLRs信号过度激活在其中起重要作用，TAM/Gas6信号和Tim-4参与了TLRs信号的调控，但是它们在IBD中的作用和机制尚不清楚。为此，本项目进行了以下研究：TAM/Gas6信号对巨噬细胞中TLRs信号的调控在IBD中的作用；Tim-4在IBD发病中的作用及其临床价值；转录组学分析实验性结肠炎及Tim-4阻断小鼠结肠黏膜中的差异表达基因。结果表明：①IBD患者和实验性结肠炎小鼠肠黏膜中TLRs信号负性调控TAM/Gas6信号被抑制，导致TLRs信号过度激活。②TAM受体和Gas6可以抑制LPS刺激后巨噬细胞内TLRs信号通路的激活及促炎细胞因子的分泌。③UC患者外周血循环CD14+Tim-4+单核细胞异常上调，与疾病严重程度和活动度密切相关，并与Treg细胞、M-MDSC细胞和炎症因子显著相关。④Tim-4在UC患者肠粘膜中高表达，并与疾病的组织学活动、Treg细胞、TLR-4/NF-κB通路及M2型巨噬细胞标志物CD206显著相关。⑤阻断Tim-4可显著改善DSS小鼠的临床症状、DAI和病理组织学炎症水平，且显著抑制TLR4/NF-κB信号通路，使Treg细胞和M1/M2趋于正常化。⑥DSS诱导的实验性结肠炎小鼠肠道菌群丰度显著减少且菌群结构发生显著变化，而Tim-4阻断后肠道微生物丰度有所增加，且菌群结构亦向健康小鼠趋化。⑦实验性结肠炎小鼠肠上皮细胞mRNA表达谱有显著改变，Tim-4对它们有调控作用，这主涉及免疫炎症反应、细胞因子产生调节等相关生物过程和信号通路。以上结果提示：TAM/Gas6信号和Tim-4参与了IBD发病，其机制可能是调控TLR信号、免疫炎症细胞的浸润和活性、及肠道菌群的平衡共同作用，这为以TAM/Gas6信号和Tim-4为靶点治疗IBD提供了实验和理论依据。