Elucidation of Pathoptysiological Roles of Immune Chemokines

该项目的目的是阐明免疫趋化因子的病理生理作用，主要针对淋巴细胞和树突状细胞。今年的主要成就如下：(1)我们发现，促炎细胞因子如IL-1和tnf - α能诱导正常人表皮角质形成细胞表达LARC/CCL20，这是一种已知的趋化因子，可选择性地吸引未成熟树突状细胞和α4β7阳性记忆/效应T细胞vccr6。我们还发现，特应性皮炎（AD）的病变皮肤通常含有产生LARC的表皮角质形成细胞和表皮层下广泛积累的CCR6+细胞。此外，我们发现来自AD患者的血浆样本通常含有升高的LARC水平。总之，这些结果支持LARC在AD中的重要病理作用。(2)我们发现，IL-1和tnf - α等促炎细胞因子能有效诱导粘膜上皮细胞中LARC的表达。此外，我们分析了这些细胞因子对LARC启动子的激活机制，发现转录起始位点-96 ~ -87bp的NF-kB位点对LARC启动子的诱导至关重要。(3)我们发现，TARC/CCL17和MDC/CCL22均可通过CCR4吸引th2型记忆/效应T细胞和寻皮CLA+记忆/效应T细胞，在AD患者血浆中显著升高。此外，我们发现血小板中含有TARC，其含量在AD患者的血小板中显著升高。我们进一步发现IFN-γ诱导正常人表皮角质形成细胞表达TARC和MDC。总的来说，TARC和MDC可能在AD的发病机制中起重要作用。此外，IFN-γ也被认为是Th1趋化因子的有效诱导剂，它可能通过诱导一组趋化因子同时吸引Th1和Th2细胞而成为AD发病的关键因素。(4)我们发现，在感染白色念珠菌的慢性炎症皮肤和淋巴结中，很大一部分成熟树突状细胞产生MDC，并与表达ccr4的T细胞聚集。因此，MDC可能在DC-T细胞相互作用中发挥重要作用。(5)我们发现，在卵清蛋白诱导的哮喘小鼠模型中，抗mtarc单克隆抗体有效地阻断了肺淋巴细胞和嗜酸性粒细胞的浸润，并提高了甲基苯丙胺诱导的气道阻力。因此，TARC可能在哮喘中发挥重要作用。(6)我们发现多发性硬化症（MS）患者脑脊液中表达CCR5或CXCR3的淋巴细胞频率在加重期间升高，而表达CCR5的淋巴细胞频率在缓解后降低。这些结果支持了ms发病机制中th1偏倚的免疫应答。(7)我们在亚美尼亚仓鼠中制备了针对小鼠趋化因子如MDC、LARC和SLC/CCL21的单克隆抗体，但无法获得具有中和活性的抗体。(8)我们将tarc敲除小鼠和αE整合素敲除小鼠反复回交到BALB/c和C57BL/6小鼠中，获得了同基因菌株。少

The aim of the project has been to elucidate the pathophysiological roles of the immune chemokines, which are mainly directed to lymphocytes and dendritic cells. The main accomplishments of this year are as follows. (1) We showed that the proinflammatory cytokines such as IL-1 and TNFα potently induced normal human epidermal keratinocytes to express LARC/CCL20, the chemokine known to selectively attracts immature dendritic cells and α4β7-positive memory/effector T cells vua CCR6. We also showed that the lesional skin of atopic dermatitis (AD) often contained epidermal keratinocytes producing LARC and extensive accumulation of CCR6+ cells beneath the epidermal layer. Furthermore, we showed that plasma samples from AD patients often contained elevated levels of LARC. Collectively, these results support an important pathologenic role of LARC in AD. (2) We showed that expression of LARC was potently induced in mucosal epithelial cells by proinflammatory cytokines such as IL-1 and TNFα. Fur … More thermore, we analysed the activation mechanism of the LARC promoter by these cytokines and showed that an NF-kB site from -96 to -87bp from the transcriptional initiation site was essential for the induction of the LARC promoter. (3) We showed that TARC/CCL17 and MDC/CCL22, both known to attract Th2-type memory/effector T cells and skin-seeking CLA+ memory/effector T cells via CCR4, were significantly evevated in the plasma of AD patients. Furthermore, we found that platelets contained TARC and its contents were dramatically elevated in platelets from AD patients. We further showed that IFN-γinduced normal human epidermal keratinocytes to express TARC and MDC. Collectively, TARC and MDC may play important roles in the pathogenesis of AD. Furthermore, IFN-γ, which is also known to be apotent inducer of Th1-attracting chemokines, may be a critical factor in AD pathogenesis by inducing a set of chemokines attracting both Th1 and Th2 cells. (4) We showed that a large fraction of mature dendritic cells produced MDC and were clustered with CCR4-expressing T cesss in chronically inflamed skin infected with Candida albicans as well as in lymph node. Thus, MDC may play an important role in DC-T cell interactions. (5) We showed that in an asthmatic model in mice induced by priming with and subsequent inhalating of ovalbumin, the treatment with anti-mTARC monoclonal antibody efficiently blocked lung infiltrating of lymphocytes and eosinophils and elevation of methacholine-induced airway resistance. Thus, TARC is likely to play an important role in asthma. (6) We showed that in patients with multiple sclerosis (MS), the frequencies of lymphocytes expressing CCR5 or CXCR3 was increased in the cerebrospinal fluid during exacerbation, while the frequencies of CCR5-expressinglymphocytes were decreased after remission. These results support the Th1-biased immune responses in the pathogenesis of MS. (7) We generated monoclonal antibodies to mouse chemokines such as MDC, LARC and SLC/CCL21 in Armenisan hamsters but could not get antibodies with neutralizing activities. (8) We have repeatedly backcrossed TARC-knockout mice and αE integrin-knockout mice into BALB/c and C57BL/6 mice and developed the syngeneic strains. Less