自噬在T细胞发育和Th细胞分化及稳定性作用机制的研究

Autophagy influences many aspects of innate and adaptive immunity. Autophagy adjusts cellular biomass and function in response to diverse stimuli, including infection. Autophagy plays specific roles in shaping immune system development, fueling host innate and adaptive immune responses. Although great progress has been made in understanding of autophagy in innate immunity in last decades, only early progress has been made in understanding the roles of autophagy in adaptive immunity, this area warrants more study. Some of the questions that have recently been asked in this area include: the role of autophagy in the polarization of TH17 cells and possibly other T cell subsets, along with the role of autophagy in general lymphocyte homeostasis and differentiation. In this proposal, we propose to address the effect of autophagy gene ATG16L1 on T cell homeostasis and T helper cell subset differentiation in a comprehensive fashion. By cross ATG16L1f/f conditional knock out mice with different Cre mice, we will analyze the effect of ATG16L1 on T cell development, determine whether ATG16L1 involves regulating T helper cell subset differentiation. In order to investigate if ATG16L1 regulate T helper cell lineage commitment, ATG16L1 will delete only after Th17 cell differentiation by crossing the ATG16L1f/f mice with Th17 specific Cre mice. Further study will performed with this mouse to investigate if ATG16L1 play a role in maintenance of Th17 cell stability and function. Finally, the mechanism of ATG16L1 regulates T cell homeostasis and T helper cell differentiation will be investigated. We believe autophagy and immunity are fully integrated, and our continuing study of the interface between these processes will be a fruitful area of sci-entific inquiry for many years to come. We believe our project will provide novel insight into an unexplored area of autophagy biology that will not only fuel a better understanding of adaptive immunity, but will facilitate the development of novel approaches to therapeutic manipulate autophagy.

自噬影响着天然免疫和适应性免疫的很多方面。虽然自噬在天然免疫中的研究取得了很多成就，但在适应性免疫中的作用还有待进一步研究。据此，利用自噬条件性敲除小鼠，研究自噬基因Atg16L1在多个方面对于T细胞稳态和辅助性T细胞亚群分化和维持的影响有非常重要的意义。首先通过将Atg16L1基因条件性敲除的小鼠（Atg16L1f/f）和在T细胞发育不同阶段表达Cre酶的小鼠交配，分析Atg16L1基因在T细胞发育中的作用；其次，通过可诱导性敲除（ATG16L1f/f/ER-T2-Cre)小鼠,研究自噬对辅助性T细胞分化的影响；最后，将重点研究Atg16L1或自噬在维持特定辅助性T细胞亚群Th17和Treg细胞稳定性中的作用和机制。本项目将以独特的角度揭示自噬中的一个未知领域，不仅能更全面的了解适应性免疫，而且将为发展治疗多种与自噬相关疾病的方法提供有利的依据。

自噬对于初始 T 细胞的稳态是非常重要的。然而，自噬尤其是 ATG16L1 在 T 细胞中的作用还不清楚。因此，我们构建了条件性基因敲除小鼠，在特定的细胞中敲除 ATG16L1 基因来研究其在 T 细胞中的生理功能。结果发现： ATG16L1 依赖性的自噬对于 T 细胞稳态的维持是必需的，敲除ATG16L1的小鼠脾脏中CD4和CD8细胞的比例明显上调，并且CD4+ 和CD8+T细胞的活化程度增加，进一步研究发现Treg细胞中敲除自噬相关基因ATG7后小鼠脾脏中Treg的细胞比例增加。在炎症条件下自噬缺陷小鼠的T细胞能够产生更多的IL-17。众所周知，在肠炎的发生发展过程中，T细胞和固有免疫细胞都发挥重要的作用，我们采用Atg16L1 T300A KI的小鼠研究了巨噬细胞的变化。发现KI小鼠的巨噬细胞表现出自噬诱导的缺陷，相应地，KI小鼠在细菌感染过程中对细菌的清除也更加缓慢，并产生了更多的促炎性细胞因子IL-1β， Salmonella 诱导的肠炎也更加严重。目前，自噬相关基因ATG16L1在辅助性 T细胞的分化和功能以及稳定性维持的作用和机制正在深入研究中。我们的研究将为了解自噬调控适应性免疫的分子机制，为临床疾病的治疗和控制提供有利的指导。此外，我们做了部分ID蛋白对T细胞分化调控的影响。发现在流感感染中，id3可以促进Th1细胞的分化，这种调控作用是依赖于E蛋白的； E蛋白可以通过直接结合到Th1细胞的关键性的转录因子T-bet的转录调控区，调控T-bet的表达，进而调控Th1细胞的分化。