3-O-C12-HSL通过结合PPARγ调控树突状细胞功能的机制研究

Pseudomonas aeruginosa (PA) is a common opportunistic pathogen that often induced immune escape for the infection of PA. Preliminary studies have indicated that the signal molecular N-3-Oxododecanoyl-L-homoserine lactone (3-O-C12-HSL,C12), which secreted by PA, inhibiting human dendritic cells maturation，which is one of the most important way that PA induced immune escape. Preliminary studies have indicated that C12, which secreted by PA, inhibited human monocyte derived-dendritic cells (Mo-DCs) maturation. But the receptor of C12 and how to regulate downstream signaling pathways which inhibited Mo-DCs maturation remains unclear. It were known that: ①it can inhibit Mo-DCs maturation when peroxisome proliferator activated receptor γ (PPARγ) and ligand binded, ②we demonstrated that PPARγ was a potential receptor of C12 by molecular docking and ligand screening tests in vitro. Therefore, we proposed that C12 inhibited the maturation of human Mo-DCs by binding and activating PPARγ and then regulating its downstream signaling pathways. This research was 1）to study the kinetic constant and which functional area of PPARγ that C12 combined, 2）to build the C12 mutant strain, from two aspects(C12 monomer and C12 mutant strains), to stuy the effect of C12 on maturation and function of Mo-DCs though PPARγ, 3）to study the impacts of C12 on the PPARγ activity and downstream signaling pathways. This research can enrich the pathogenesis of the PA and provide new strategy for the prevention and treatment of PA infection.

铜绿假单胞菌(PA)诱导机体产生免疫逃逸是其致病的重要方式。前期研究表明PA信号分子3-O-C12-HSL(C12)阻碍人单核细胞衍生树突状细胞(Mo-DCs)成熟，参与免疫逃逸。但C12阻碍Mo-DCs成熟的机制未明。文献报道Mo-DCs中过氧化物酶增殖物激活受体γ（PPARγ)与配体结合后阻碍Mo-DCs成熟。同时，我们前期通过分子对接技术及体外配体筛选实验发现C12与PPARγ直接结合。为此，我们提出C12通过与Mo-DCs中PPARγ结合，调控PPARγ活性及信号通路，影响Mo-DCs成熟与功能。本项目拟：①明确C12与PPARγ结合的动力学常数和结合的功能区；②构建C12突变菌株，从C12单体及PA整体水平两个方面，明确C12通过PPARγ调节Mo-DCs成熟与功能；③阐明C12对PPARγ表达水平、活性及信号通路的影响。本课题为发现PA免疫逃逸的新靶点提供依据。

铜绿假单胞菌(PA)诱导机体产生免疫逃逸是其致病的重要方式。前期研究表明PA信号分子3-O-C12-HSL(C12)阻碍人单核细胞衍生树突状细胞(Mo-DCs)成熟，参与免疫逃逸。但C12阻碍Mo-DCs成熟的机制未明。文献报道Mo-DCs中过氧化物酶增殖物激活受体γ（PPARγ)与配体结合后阻碍Mo-DCs成熟。同时，我们前期通过分子对接技术及体外配体筛选实验发现C12与PPARγ直接结合。因此，我们提出C12通过与Mo-DCs中PPARγ结合，调控PPARγ活性及信号通路，影响Mo-DCs成熟与功能。为此，我们构建PPARγ配体结合区肽段，应用表面等离子共振技术证实C12与PPARγ结合并获得结合参数。在此基础上，构建PPARγ过表达载体，应用PPARγ过表达载体及GW9662（PPARγ的拮抗剂）处理Mo-DCs，发现C12通过PPARγ调节JNK MAPK信号通路、抑制Mo-DCs成熟及功能。本课题为发现PA免疫逃逸的新靶点及制备针对C12的特异性药物提供实验依据。