HIGH MOLECULAR PROTEINS RESPONSIBLE FOR INTRACTABLE INFLAMMATORY DISEASES-IDENTIFICATION AND GENE EXPRESSION-

目前尚不清楚哪些功能分子在包括胶原病在内的顽固性炎症性疾病的发展中起关键作用，尽管在这些疾病中已经指出了自身免疫的几个参数。本研究以MRL/MP-LPR/LPR(MRL/LPR)小鼠为模型，研究了与胶原病发生发展相关的高分子蛋白质及其基因表达。这种品系的小鼠会自发地发展为致命性肾小球肾炎、动脉炎和关节炎，与免疫紊乱诱导基因LPR的表达有关，LPR最近被证实是Fas抗原缺失突变。这些小鼠在同一个体中患上所有这些疾病，但除了LPR基因外，它们还需要MRL遗传背景。我们澄清了通过将MRL与非自身免疫倾向小鼠杂交，这些疾病的背景基因可以相互遗传分离。考虑t…从这些MRL杂交鼠中，我们鉴定了导致MRL/LPR小鼠胶原疾病的蛋白：肾小球肾炎的IgG3亚类和肉芽肿性动脉炎的肿瘤坏死因子亚类。在对IgG3的进一步分析中，我们成功地获得了针对正常或SCID小鼠的肾炎IgG3产生克隆，而且这些克隆在组织病理学表现上产生了规律性的变化。对这些克隆的免疫球蛋白基因分析表明，每个致肾病的IgG3来自不同的B细胞前体，并使用不同的VH生殖系基因。此外，其中一种IgG3与VH区的体细胞突变无关，表明VH区的体细胞突变不是肾炎性抗体产生所必需的。此外，该胚系VH基因与在非自身免疫倾向的小鼠中发现的基因相同。因此，与肾炎性抗体相关的胚系VH基因可能不存在易于自身免疫的特异性等位基因。关于肿瘤坏死因子在动脉炎的发生发展中，在MRL和非自身免疫易感小鼠中没有肿瘤坏死因子-外显子4的RFLP。然而，在MRL杂交小鼠中，动脉炎个体的肿瘤坏死因子和IL-1βmRNA水平呈正相关，而其他巨噬细胞相关细胞因子mRNA水平如LIF、M-CSF和Eta-1之间则不相关。这一事实表明，巨噬细胞的特殊潜能在MRL/LPR小鼠动脉炎的发生中是必需的，并受其背景基因的限制。进一步研究导致肾炎的IgG3可变区序列，包括恒定区的关联，以及产生肾炎性抗体的B细胞克隆的诱导机制，需要进一步的研究。从MRL杂交小鼠建立的易患动脉炎的小鼠品系将有助于进一步研究动脉炎的遗传基础和相关基因。较少

It is still unclear what functional molecules are critical for the development of intractable inflammatory diseases including collagen disease, although several parameters of autoimmunity have been pointed out in these diseases. This project was performed to identify high molecular proteins responsible for the development of collagen disease and to analyze their gene expression by using a murine model, MRL/Mp-lpr/lpr (MRL/lpr) mice. This strain of mice spontaneously develops a lethal glomerulonephritis, arteritis and arthritis, associated with the expression of the immunological disorder-inducing gene, lpr, which is recently clarified to be Fas antigen deletion mutant. These mice develop all of these diseases in the same individual, but MRL genetic background is required for them in addition to the lpr gene.We clarified that the background genes for these diseases are able to be genetically segregated each other by using the MRL hybrid mice with non-autoimmune-prone mice. Considering t … More he facts from these MRL hybrid mice, we identified the responsible proteins for collagen disease in MRL/lpr mice; IgG3 subclass for glomerulonephritis and TNF for granulomatous arteritis.In further analyses of IgG3, we succeeded in obtaining nephritogenic IgG3 producing clones against normal or SCID mice, and moreover these clone generated regular variations of lupus nephritis in histopathological manifestations. Immunoglobulin gene analysis of these clones clarified that each nephritogenic IgG3 is derived from a different B cell precursor and used a different VH germline gene. Moreover, one of these IgG3 was not associated with somatic mutation in the VH region, indicating that somatic mutation in the VH region is not required for the development of nephritogenic antibody. Furthermore, this germline VH gene was identical to that found in a non-autoimmune-prone mice. Thus, there may be no autoimmune-prone specific al- lelism in the germline VH gene relevant to the nephritogenic antibody.Regarding TNF on the development of arteritis, there is no RFLP of TNF-exon 4 among MRL and non- autoimmune-prone mice. However, it was clear that the individuals with arteritis among the MRL hybrid mice manifest a positive correlation between TNF and IL-1 beta mRNA level, but not between other macrophage-related cytokine mRNA such as LIF, M-CSF and Eta-1. This fact indicates that particular potential of macrophages are required for the development of arteritis in MRL/lpr mice, restricted with their background genes.Further studies of the sequences in the variable regions of IgG3 responsible for the nephritogenicity, including the association of the constant regions, and the induction mechanisms of nephritogenic antibody- producing B cell clones are required. An arteritis-prone strain of mice established from the MRL hybrid mice will be useful for further studies of the genetic basis of arteritis and of the responsible genes. Less