MECNHANISMS OF CELL ADHESION AND TRAFFICKING IN EXPERIMENTAL ILEITIS

项目4由Klaus Ley博士领导，研究SAMP1/YitFc(SAMP)小鼠肠道T细胞黏附和炎性细胞运输的机制。最初的项目旨在检验这样的假设：1)SAMP小鼠表达与T细胞、中性粒细胞和其他白细胞的运输有关的黏附分子，2)从SAMP小鼠移植的CD4+T细胞的接受者发生回肠炎需要白细胞和/或内皮黏附分子，以及3)CD4+T细胞使用特定的黏附分子定位于SAMP小鼠的末端回肠并导致疾病。在过去的四年中，关于血管内皮细胞黏附分子VCAM-1、ICAM-1、MAdCAM-1、P-选择素和Pnad在SAMP回肠炎症区域显著上调的研究取得了重要进展。治疗研究表明，在过继转移模型中，阻断α4整合素、血管细胞黏附分子-1和细胞间黏附分子-1的组合或MAdCAM-1和L-选择素的组合显著改善疾病严重程度。L-选择素在促进炎症中的重要作用这一发现代表了关于这种分子的一个新的和意想不到的观察，因为L-选择素被认为表达在幼稚的T细胞上。然而，L-选择素的抑制可能会限制炎性髓系细胞的募集，以及幼稚、记忆和调节性T细胞，或它们的组合。此外，我们还发现B细胞在SAMP回肠炎模型中起致病作用。在下一个资助期，项目4将根据最近的两项发现，将重点放在L-选择素及其配体之一PSGL-1上：1)论证L-选择素的重要性 2)初步证据表明PSGL-1在回肠炎的血管内皮细胞中表达。 回肠末端的固有层微血管。基于这些发现，本项目将首先使用免疫染色、激光捕获显微解剖和实时RT-PCR检测回肠固有层和黏膜下层炎症的内皮细胞是否表达L-选择素配体PSGL-1。将使用单抗阻断PSGL-1，以及通过将SAMP小鼠与现有PSGL-1缺陷小鼠使用速度基因进行杂交，来研究PSGL-1/L-选择素黏附途径的功能作用。最后，将研究L-选择素表达细胞在SAMP过继转移模型中对回肠炎严重程度的影响。这些研究旨在了解致病淋巴细胞、幼稚淋巴细胞和调节性淋巴细胞在SAMP小鼠炎症回肠中的转运机制，并提供合理的 通过单一或联合阻断克罗恩病患者的肠道特异性黏附分子来改善疾病严重程度的基础。

Project 4, headed by Dr. Klaus Ley, studies the mechanisms of intestinal T cell adhesion and inflammatory cell trafficking in SAMP1/YitFc (SAMP) mice. The original project was designed to test the hypothesis that: 1) SAMP mice express adhesion molecules relevant to the trafficking of T cells, neutrophils, and other leukocytes, 2) leukocytes and/or endothelial adhesion molecules are required for the development of ileitis in recipients of CD4+ T cells transferred from SAMP mice, and 3) CD4+ T cells use specific adhesion molecules to home to the terminal ileum of SAMP mice and cause disease. In the past four years, important progress has been made with respect to the discovery that the endothelial adhesion molecules VCAM-1, ICAM-1, MAdCAM-1, P-selectin, and PNAd are significantly upregulated in inflamed areas of the SAMP ileum. Therapeutic studies have shown that blocking alpha 4 integrins, a combination of VCAM-1 and ICAM-1, or a combination of MAdCAM-1 and L-selectin significantly ameliorates disease severity in the adoptive transfer model. The finding that L-selectin is important in promoting inflammation represented a novel and unexpected observation regarding this molecule, since L-selectin is thought to be expressed on naive T cells. However, L-selectin inhibition may limit the recruitment of inflammatory myeloid cells, as well as naive, memory, and regulatory T cells, or a combination thereof. In addition, we discovered that B cells play a pathogenic role in the SAMP model of ileitis. For the next funding period, Project 4 will focus on L-selectin and one of its ligands, PSGL-1, based on two recent discoveries: 1) demonstration of the importance of L-selectin in mediating ileitis, and 2) preliminary evidence that PSGL-1 is expressed in endothelial cells of lamina propria microvessels in the terminal ileum. Based on these discoveries, this project will first test whether the L-selectin ligand PSGL-1 is expressed in inflamed endothelial cells of the ileal lamina propria and submucosa using immunostaining, laser capture microdissection, and real-time RT-PCR. The functional role of the PSGL-1/L-selectin pathway of adhesion will be studied using monoclonal antibodies to block PSGL-1, as well as by crossing SAMP mice with existing PSGL-1 deficient mice using speed congenics. Finally, the nature of L-selectin expressing cells that contribute to ileitis severity in the SAMP adoptive transfer model will be studied. These studies are designed to understand the mechanisms of trafficking of pathogenic, naive and regulatory lymphocytes in the inflamed ileum of SAMP mice and to provide a rational basis for ameliorating disease severity by single or combined blockade of intestinal-specific adhesion molecules in patients with Crohn's disease.