Role of inflammation, innate resistance, and immunity in carcinogenesis.

该项目的几个目标已经取得了进展，特别是TLR信号在调节成纤维细胞和树突状细胞增殖和存活中的作用，对人DC细胞因子产生的调节，浆细胞样DC在调节口服耐受性中的作用，NK细胞对弓形虫感染中Th1分化的影响，TNF， MyD88和IL-18在结肠炎依赖性癌变中的作用。肝浆细胞样树突状细胞有助于口服诱导CD8+ T细胞耐受（免疫。2008;9:464-75）。肝脏被认为有助于全身T细胞对口服吸收抗原的耐受性，尽管其耐受性作用的确切机制尚不清楚。在半抗原特异性接触敏感性（CHS）模型中，我们发现肝脏是一个口腔抗原呈递的部位，肝树突状细胞（DC）可以耐受小鼠随后的CD8+ T细胞启动。肝DC的耐受性潜能局限于浆细胞样DC (pDC)，通过暴露于半抗原而增强，并且需要CD4+ T细胞。最后，体内pDC的消耗消除了口服耐受性，恢复了半抗原特异性CD8+ T细胞和CHS反应。因此，pDC具有耐受性，在口腔耐受性中起重要作用。这些研究目前正在扩展，以分析肝pDC在其他耐受模型中的作用，特别是在结肠炎和癌变模型中是否需要pDC来诱导TNBS耐受。我们培育了在pDC上特异性表达人白喉毒素受体和绿色荧光蛋白的小鼠。这些小鼠可用于pDC的鉴定和体内消耗，并将用于pDC的体内鉴定和消耗实验。除了口服耐受模型外，我们已经建立了肠固有层中tlr9激活的pDC阻止未成熟CD4 T细胞在FoxP3+ Treg中的转化，我们正在将我们的研究扩展到其他pDCs参与免疫调节的模型。刚地弓形虫感染TAP1基因缺陷小鼠的NK细胞ifn - γ产生受损，导致CD4+ T细胞启动缺陷和死亡率升高[J] .中华检验医学杂志，2007,24(4):591- 592。为了研究CD8+ T细胞介导的弓形虫体内控制是否需要Transporter Associated with Antigen Processing (TAP)1，我们比较了TAP1-/-、CD8-/-和野生型（WT）小鼠对弓形虫感染的抗性。令人惊讶的是，TAP1-/-小鼠比CD8-/-或WT小鼠更容易感染一种无毒的寄生虫菌株。TAP1-/-小鼠的抗性降低与激活和产生ifn - γ的CD4+ T细胞频率降低相关。有趣的是，与WT对照组相比，感染TAP1-/-的小鼠表现出产生ifn - γ的自然杀伤（NK）细胞的频率降低，NK细胞消耗后，CD8-/-和WT小鼠都以与TAP1-/-动物相同的动力学屈服于感染，并表现出受损的CD4+T细胞ifn - γ反应。总之，这些结果揭示了TAP1在诱导ifn - γ中以前未被认识的作用&amp；#61472；产生NK细胞，并首次证明了该细胞群在启动CD4+ T淋巴细胞对弓形虫感染的反应中的功能。这些研究现在正在扩展，以了解TAP1-/-和β 2微球蛋白-/-在NK细胞这一功能中的差异，以及NKT细胞在调节NK细胞活性中的可能作用。先天抵抗和促炎性细胞因子在癌变中的作用。一项非常广泛的研究已经开始研究炎症受体和细胞因子在皮肤和结肠化学致癌中的作用。在我们早期观察到MyD88-/-对皮肤癌变具有抗性之后，我们现在发现MyD88在耐辐射皮肤细胞和造血细胞中都需要表达，并且MyD88-/-角化细胞在体外Ras转化后其基因表达模式发生了很大改变，这表明这种基因表达模式的改变可能是在化学癌变反应中缺乏息肉形成的原因。在结肠中，MyD88缺乏导致对DSS诱导的结肠炎的易感性增加，令人惊讶的是，这与对化学诱导的癌变的易感性高度增加有关。MyD88表型的部分原因是MyD88-/-细胞中缺乏通过IL-18受体传递的信号。我们还观察到，结肠炎依赖性癌变需要TNF，但有趣的是，由肠细胞产生的TNF是癌变所必需的，而T细胞和巨噬细胞产生的TNF具有抗肿瘤作用。这些结果表明，TNF，一种最初被描述为具有抗肿瘤活性的分子，后来被描述为致癌的必需因子，确实具有双重作用，其抗肿瘤和促肿瘤活性取决于其产生的细胞，并可能取决于其在癌变过程中由不同细胞类型的时间产生。

Progress has been made in several of the aims of the project, in particular on the role of TLR signaling in regulating proliferation and survival of fibroblasts and dendritic cells, on the regulation of cytokine production from human DC, on the role of plasmacytoid DC in regulating oral tolerance, on the effect of NK cells on Th1 differentiation in Toxoplasma Gondii infection, role of TNF, MyD88 and IL-18 in colitis-dependent carcinogenesis. Hepatic plasmacytoid dendritic cells contribute to orally induced CD8+ T cell tolerance (Immunity. 2008; 9:464-75). The liver is thought to contribute to systemic T cell tolerance to orally absorbed antigens, although the precise mechanism of its tolerogenic effect is unclear. Here we show that the liver is a site of oral antigen presentation and that hepatic dendritic dells (DC) can tolerize mice to subsequent CD8+ T cell priming, in a model of hapten-specific contact sensitivity (CHS). The tolerogenic potential of liver DC is confined to plasmacytoid DC (pDC), is enhanced by exposure to hapten, and requires CD4+ T cells. Finally, in vivo depletion of pDC abrogated oral tolerance and restored hapten-specific CD8+ T cell and CHS responses. Thus, pDC are tolerogenic and play an essential role in oral tolerance. These studies are now being extended to analyze the role of liver pDC in other model of tolerance and in particular in testing whether pDC are require to induce TNBS tolerance in a model of colitis and carcinogenesis. We have generate a mouse expressing the human Dyptheria Toxin Receptor and the Green Fluorescent protein specifically on pDC. These mice can be used for the identication and in vivo depletion of pDC and they will be used in experiments aimed to identify and deplete pDCs in vivo. In addition to the models of oral tolerance we have established that TLR9-activated pDC in the intestinal lamina propria prevent the conversion of naive CD4 T cells in FoxP3+ Treg and we are estending our studies to other models in which pDCs are involved in immunoregulation. Toxoplasma gondii infected TAP1 deficient mice display impaired NK cell IFN-gamma production leading to defective CD4+ T cell priming and increased mortality (J Exp Med. 2007, 204:2591-602). To investigate if Transporter Associated with Antigen Processing (TAP)1 is required for CD8+ T cell mediated control of Toxoplasma gondii in vivo, we compared the resistance of TAP1-/-, CD8-/- and wild-type (WT) mice to infection with the parasite. Surprisingly TAP1-/- mice displayed greater susceptibility than either CD8-/- or WT mice to infection with an avirulent parasite strain. The decreased resistance of the TAP1-/- mice correlated with a reduction in the frequency of activated and IFN-gamma-producing CD4+ T cells. Interestingly, infected TAP1-/- mice showed a reduced frequency of IFN-gamma producing natural killer (NK) cells relative to that of WT controls, and after NK cell-depletion both CD8-/- and WT mice succumbed to infection with the same kinetics as TAP1-/- animals and displayed impaired CD4+T cell IFN-gamma responses. Together, these results reveal a previously unappreciated role for TAP1 in the induction of IFN-gamma producing NK cells and provide the first demonstration of the function of this cell population in the priming of CD4+ T lymphocyte responses to T. gondii infection. These studies are now being extended to understand the difference between the TAP1-/- and beta2microglobulin-/- in this function of NK cells and the possible role of NKT cells in the regulation of NK activity. Innate resistance and pro-inflammatory cytokines in carcinogenesis. A very extensive investigation has been initiated to study the role of inflammatory receptors and cytokines in skin and colon chemical carcinogenesis. Following our early observation that MyD88-/- are resistant to skin carcinogenesis, we have now showed that MyD88 expression is required both in radioresistant skin cells as well as in hematopoieitc cells and that MyD88-/- keratinocytes are very altered in their gene expression pattern after Ras transformation in vitro, suggesting that this altered pattern of gene expression may be responsible for the lack of polyps formation in response to chemical carcinogenesis. In the colon, MyD88 deficiency resulted in an increased susceptibility to DSS induced colitis that surprisingly was associated with a highly increased susceptibility to chemically induced carcinogenesis. The MyD88 phenotype was in part due to lack of signaling through the IL-18 receptor in the MyD88-/- cells. We also observed that TNF is required for colitis-dependent carcinogenesis but, interestingly, TNF produced by the enterocytes was required for carcinogenesis whereas T cells and macrophages produced TNF had an antitumor effect. These results indicate that TNF, a molecule described originally for its anti-tumor activity but then described as a required factor for carcinogenesis, has indeed a dual role and its anti-tumor and pro-tumor activity depend by which cells it is produced and probably by its temporal production by different cell types during the carcinogenesis processes.