Regulation of Cutaneous Inflammation by local gd T Cells

描述（由申请人提供）：在鸡、小鼠和人类等多种物种中，大量的T细胞组成性地与体表上皮相关，如肠道、泌尿生殖道和皮肤。这种上皮内淋巴细胞（iel）通常富集于T细胞受体(TCR) γ / δ +细胞中，通常具有有限的组织相关抗原受体多样性；因此，小鼠皮肤中的大多数IELs，也称为树突状表皮T细胞（DETC），表达明显同质的Vgamma5/Vdelta1 TCRs，值得注意的是它们缺乏连接（CDR3区域）多样性。联合使用不同菌株的TCRd-/-（敲除）小鼠，并通过过继转移对这些小鼠进行选择性重构，表明Vgamma5+ DETC，而不是其他γ / δ细胞，是几种生理相关的皮肤炎症反应的有效下调因子，包括局部的、遗传依赖性的、tcrα / β + T细胞依赖性的、环境依赖性的慢性皮炎，这些慢性皮炎具有人类特应性皮炎的几个特征。该项目的长期目标是利用这个强大的实验模型来确定局部T细胞调节局部组织内全身免疫反应的机制。本项目的具体目标是：1。使用免疫组织学、体外流式细胞术和微阵列分析详细表征通常由DETC下调的皮肤炎症；B)通过细胞因子拮抗剂或阻断抗体治疗或第二次“敲除”基因突变，易感的δ -/-小鼠与δ -/-小鼠的表型比较也缺乏选定的促炎细胞和/或分子2。为了通过基因表达序列分析（SAGE）来表征“静息”DETC和“激活”DETC在体外表达的基因，通过各种体内和体外激活状态下DETC的定量RT-PCR来验证这些分析，并将这些基因表达模式与肠道相关的γ / δ +和α / β + IELs以及系统性CD8+ α / β +“幼稚”和“记忆”T细胞的表达模式进行比较。3. 通过重组δ -/-受体，使胎儿胸腺DETC前体缺乏候选抗炎分子，研究选择的DETC候选抗炎因子/效应分子。4. 利用全基因组微卫星定位来确定控制自发性皮炎易感性/抗性的遗传区间，这些遗传区间在一些d-/-小鼠中发生，但在其他d-/-小鼠中没有发生，随后进行额外的研究（同源系的发展，该区间内基因的差异表达分析），旨在明确识别调节皮肤炎症的基因。

DESCRIPTION (provided by applicant): In species as diverse as chickens, mice, and humans, substantial numbers of T cells are constitutively associated with body surface epithelia, such as the gut, genitourinary tract, and the skin. Such intraepithelial lymphocytes (IELs) are commonly enriched in T cell receptor (TCR)gamma/delta+ cells, frequently with limited tissue associated antigen receptor diversity; thus, most IELs in mouse skin, also known as dendritic epidermal T cells (DETC), express strikingly homogeneous Vgamma5/Vdelta1 TCRs notable for their lack of junctional (CDR3 region) diversity. Combined use of different strains of TCRd-/- (knockout) mice and selective reconstitution of such mice via adoptive transfer has shown that Vgamma5+ DETC, but not other gamma/delta cells, are potent down-regulators of several physiologically relevant, cutaneous inflammatory responses, including a localized, genetically-dependent, TCRalpha/beta+ T cell-dependent environmentally-dependent, chronic dermatitis that shares several features of human atopic dermatitis. The long-term goal of this project is to utilize this powerful experimental model to define the mechanisms by which local T cells regulate the effects of systemic immune responses within local tissues. The specific aims of this project are: 1. To characterize in detail the cutaneous inflammation normally down-regulated by DETC) using: a) immunohistology, ex vivo flow cytometry, and microarray analysis; b) phenotypic comparisons of susceptible delta-/- mice with delta-/- mice also deficient in selected pro-inflammatory cells and/or molecules by virtue either of treatment with cytokine antagonists or blocking antibodies, or of a second "knockout" genetic mutation 2. To characterize the genes expressed by "resting" DETC and DETC "activated" in vitro by serial analysis of gene expression (SAGE), validate such analyses by quantitative RT-PCR of DETC in various in vivo and in vitro activation states, and compare such gene expression patterns with those both of gut-associated gamma/delta+ and alpha/beta+ IELs, and of systemic CD8+ alpha/beta+ "naive" and "memory" T cells. 3. To investigate selected candidate DETC anti-inflammatory cytokines/effector molecules by reconstituting delta-/- recipients with fetal thymic DETC precursors rendered deficient in candidate anti-inflammatory molecules. 4. To utilize genome-wide microsatellite mapping to identify the genetic interval(s) controlling susceptibility/resistance to the spontaneous dermatitis that develops in some, but not other d-/- mice, followed by additional studies (development of congenic lines, differential expression analyses of genes contained within this interval) directed at definitive identification of the gene(s) that regulate cutaneous inflammation.