Type 1 diabetes: the role of commensal microbiota

描述（由申请人提供）：1型糖尿病（T1D）是一种使人衰弱的自身免疫性疾病，在发达国家的发病率正在增加。许多事实表明微生物参与了T1D的调节。在初步研究中，我们已经确定：a。缺乏MyD 88衔接子的非肥胖糖尿病（NOD）小鼠在常规无特定病原体（SPF）条件下饲养时对自发性T1D具有抗性; B. MyD88 KO NOD小鼠未显示对胰岛抗原的全身耐受性，但在局部胰腺淋巴结中显示T细胞对此类抗原的耐受性; c.当使相同的动物无菌（GF）时，它们以约100%的频率发展糖尿病; d.当GF MyD88 KO动物用确定的微生物群重建时，糖尿病发病率降低。 总之，这些数据表明，肠道中的肠道（非致病性）微生物影响了T1D的发展。MyD88的可能作用是控制肠道微生物。当MyD88缺失时，肠道微生物群的变化随之而来。为了支持这一观点，我们对正常NOD和MyD88 KO动物的肠道微生物群进行了宏基因组分析（16S rRNA基因测序），发现了它们之间显著且有意义的差异。利用基因敲除和条件敲除小鼠、无菌技术和宏基因组测序方法，我们将实现以下目标1。揭示控制肠道微生物的MyD88依赖性机制，从而促进T1D。 我们将测试TLR 2的可能作用以及B细胞和肠上皮细胞在控制肠道微生物群中的作用，并通过条件性消除MyD88来研究MyD88 KO小鼠对T1D的抗性的细胞基础。2.阐明MyD88独立的机制，通过该机制，药物引起T细胞耐受。 我们将测试不同的模式识别受体（PRR）在预防T1D中的作用。我们将使用基因表达分析的结果：测试PLN细胞亚群中已鉴定基因的表达，以及CCR5在微生物群预防T1D中的重要性。我们将讨论抗原呈递细胞（APC）和调节性T细胞（Treg）在微生物群诱导的局部耐受中的作用。3.研究肠道微生物群在预防T1D中的作用。 我们将测试一个假设，即特定的细菌可以预防T1D，并测试选定的微生物群落本身或与免疫抑制剂联合逆转GF小鼠临床糖尿病的能力。

DESCRIPTION (provided by applicant): Type 1 diabetes (T1D) is a debilitating autoimmune disease with the incidence on the increase in developed countries. Many facts point at microbial involvement in the regulation of T1D. In preliminary studies we have established that: a. non-obese diabetic (NOD) mice lacking MyD88 adaptor were resistant to spontaneous T1D when housed in conventional specific-pathogen-free (SPF) conditions; b. MyD88 KO NOD mice did not show systemic tolerance to islet antigens, but did show tolerance of T cells to such antigens in the local pancreatic lymph nodes; c. when the same animals were made germ-free (GF), they developed diabetes with the frequency of about 100%; d. when GF MyD88 KO animals were reconstituted with the defined microflora, the diabetes incidence was reduced. Taken together the data suggested that commensal (not pathogenic) microbes in the gut influenced the development of T1D. The likely role of MyD88 is to control commensal microbes. When MyD88 is missing, the changes in gut microbiota ensue. To support that idea, we have performed the metagenomic analysis of the gut microbiota (16S rRNA genes sequencing) from normal NOD and MyD88 KO animals, and discovered significant and meaningful differences between them. Taking advantage of knock-out and conditional knock- out mice, the germ-free technology and metagenomic sequencing approach, we will pursue the following aims 1. Uncover MyD88-dependent mechanisms that control commensal microbes, thus promoting T1D. We will test a possible role of TLR2 as well as the role of B cells and intestinal epithelial cells in control of intestinal microbiota and study the cellular basis for resistance of MyD88 KO mice to T1D by conditional elimination of MyD88. 2. Elucidate MyD88-independent mechanisms by which commensals cause T cell tolerance. We will test different Pattern Recognition receptors (PRRs) for their role in prevention of T1D. We will use the results of a gene expression analysis: test expression of identified genes in cellular subsets in PLN and an importance of CCR5 in prevention of T1D by microbiota. We will address the role of antigen presenting cells (APC) and regulatory T cells (Treg) in local tolerance induced by microbiota. 3. Study the role of intestinal microbiota in prevention of T1D. We will test a hypothesis that particular bacteria protect against T1D and test the ability of selected microbial communities to reverse clinical diabetes in GF mice by themselves or in combination with immunosuppression.