Modeling and Analysis of the Role of Microbiota Metabolites in T-Cell Differentiation

 描述(申请人提供)：调节性T细胞(Tregs)包括一类异质性淋巴细胞，能够通过分泌细胞因子和调节树突状细胞功能来促进外周组织的免疫耐受。目前存在的方法是在有转化生长因子-b(诱导或iTregs)存在的情况下选择性地体外扩增Tregs，并转移到患者身上，用于治疗炎症性肠病、移植物抗宿主病和其他以过度炎症为特征的病理疾病的炎症抑制。虽然iTregs的采用转移在临床上有望是安全的，但在这一治疗策略的翻译中仍然存在诸如等主要障碍。例如， 转移到急性或慢性炎症患者体内的iTregs可以从抗 炎症时，iTreg表型转变为促炎表型，Th17表型受存在于微环境中的炎性细胞因子的影响，从而负向促进炎症状态。因此，当iTregs被引入炎症微环境中时，产生具有稳定调节表型和功能的iTregs是很重要的。几项研究将体内Treg的发育与肠道(微生物区系)中丰富而多样的微生物种群的存在联系起来。虽然微生物区系在生理免疫耐受中的作用尚不清楚，但一个流行的假说是，微生物区系产生特定的因子，促进Treg诱导并将肠道免疫调节到耐受状态。我们之前证明了吲哚，一种源自饮食色氨酸的微生物组代谢物，存在于健康小鼠和人类的胃肠道中，可以减轻炎症指标。我们实验室未发表的数据还表明，在吲哚和iTreg倾斜的条件下进行体外培养后，CD4+CD25-幼稚T细胞戏剧性地扩展为Foxp3+iTregs。然而，iTreg的稳定性和功能可以被几种促炎和抗炎细胞因子协同增强或减弱，因此，系统地预测微生物代谢产物调节炎症环境中Treg稳定性和功能的关系将有助于我们理解Treg的免疫生物学，并促进Treg细胞的治疗。我们的总体假设是色氨酸衍生的微生物代谢产物(TDMM)在体内诱导具有增强稳定性的Treg表型。利用接触不同色氨酸来源微生物区系代谢物的Treg和Th17数据，我们建议建立神经网络模型，用于在吲哚存在下的体外Treg诱导和体内转移后的稳定性，并使用该模型生成关于最佳Treg诱导、功能和稳定性的可测试预测。具体的 目的是：(1)在体外全面测定TDMM诱导的iTregs和Th17细胞的表型和功能；(2)建立TDMM在体外对iTreg诱导和Th17衰减以及体内表型维持的影响的模型；(3)确定TDMM诱导的Tregs和TDMM减毒的Th17细胞转移到淋巴细胞减少和实验性结肠炎小鼠体内后的功能和稳定性。

 DESCRIPTION (provided by applicant): Regulatory T cells (Tregs) comprise a heterogeneous class of lymphocytes that are able to promote immune tolerance in peripheral tissue through cytokine secretion and modulation of dendritic cell function. Methods currently exist to selectively expand Tregs in vitro in presence of TGF-b (induced or iTregs) and transfer to patients for therapeutic inhibition of inflammation in those suffering from inflammatory bowel disease, graft- versus-host disease and other pathologies characterized by excessive inflammation. Although the adoptive transfer of iTregs has the promise to be safe in the clinic, major hurdles such as still exist in the translation of this therapeutic strategy. For example, the iTregs transferred into a patient with acute or chronic inflammation could transition from the anti inflammatory, iTreg phenotype to a pro-inflammatory, Th17 phenotype under the influence of inflammatory cytokines present in the microenvironment, and thereby, negatively contribute to the inflammatory state. Therefore, it is important to generate iTregs that possess a stable regulatory phenotype and function when introduced into an inflammatory microenvironment. Several studies link in vivo Treg development to the presence of an abundant and diverse microbial population in the intestinal tract (the microbiota). Although the microbiota's role in physiologic immune tolerance is poorly understood, a prevalent hypothesis is that the microbiota produces specific factors that promote Treg induction and modulate gut immunity towards a tolerant state. We previously demonstrated that indole, a microbiota metabolite derived from dietary tryptophan and present in the GI tract of both healthy mice and humans, attenuate indicators of inflammation. Unpublished data from our lab also show that, after conditioning in vitro in the presence of indole and iTreg-skewing conditions, CD4+ CD25- naïve T cells dramatically expand into Foxp3+ iTregs. However, iTreg stability and function can be synergistically increased or attenuated by several pro- and anti-inflammatory cytokines, and therefore, the ability to systematically predict the relationship of microbiota metabolite regulatin of Treg stability and function in inflammatory environments would contribute to our understanding of Treg immunobiology and advance Treg cell-based therapy. Our overall hypothesis is that tryptophan derived microbiota metabolites (TDMMs) induce a Treg phenotype with enhanced stability in vivo. Using Treg and Th17 data from exposure to different tryptophan derived microbiota metabolites, we propose to develop neural network models for Treg induction in vitro and stability post-transfer in vivo in the presence of indole, and use the model for generating testable predictions on optimal Treg induction, function and stability. The specific aims are: (1): To comprehensively determine the phenotype and function of TDMM-induced iTregs and Th17 cells in vitro; (2): Model the effect of TDMM on iTreg induction and Th17 attenuation in vitro and phenotype maintenance in vivo; and (3): To determine the function and stability of TDMM-induced Tregs and TDMM- attenuated Th17 cells after transfer to lymphopenic mice and mice with experimentally induced colitis.