An approach to the pathogenesis of inflammatory lung diseases by establishing efficient analytical protocols for multiple mediator gene polymorphisms

1. 趋化因子等基因多态性的鉴定和分析：我们首先分析了日本健康对照者和肺结核患者中一种NRAMP1基因的多种基因多态性。在所分析的多态性中，一个和两个GT重复序列的罕见组合在肺纤维化（PF）患者中更为常见。然后，我们分析了日本健康对照和肺纤维化患者的纤维连接蛋白snp。虽然这些频率在日本人和白种人之间有很大差异，但在PF患者中没有差异。通过检测MCP-1启动子-2518 SNP，我们发现等位基因G在日本人中频率更高，但在PF患者中频率没有差异。不同MCP-1基因型间血浆MCP-1水平无显著差异。对于IL-8基因分析，我们通过直接测序1 Kb启动子区域寻找新的多态性，发现该SNP在败血症患者中频率分布为1:3:6，在健康对照组中频率分布为16:47:37 （a / a: a /T:T/T）。(1)大鼠辐射纤维化模型：分析趋化因子MCP-1、mip - 1α、β、MIP-2、淋巴趋化因子和fractalkine的基因表达，发现辐照后4周MCP-1、TARC mRNA表达增强，8周MCP-1、mip - 1α表达略有增加。2)小鼠烧伤引发脓毒症和ARDS模型：未启动组72小时生存率为100%，烧伤启动组72小时生存率为0%。血浆细胞因子的增加在未启动组呈单峰，在烧伤启动组呈双峰。治疗1周后，烧伤组肝脏IL-18含量明显高于假手术组。

1. Identifiticantion and analysis of chemokine, etc gene polymorphisms: Initially we analyzed various gene polymorphisms of a NRAMP1 gene in Japanese healthy controls and patients with pulmonary tuberculosis. Among polymorphisms analyzed, the rare combination of one and two GT repeats was more frequent in patients with pulmonary fibrosis (PF).We then analyzed fibronectin SNPs in Japanese healthy controls and patients with pulmonary fibrosis. Although these frequency was quite different between Japanese and Caucasians, it was not different in PF patients.By examining MCP-1 promoter -2518 SNP, we found that allele G was more frequent in Japanese, but its frequency was not different in PF patients. Plasma MCP-1 levels was not different among MCP-1 genotypes.For IL-8 gene analysis, we searched new polymorphisms by directly sequencing 1 Kb promoter region and found that the frequency distribution of this SNP was 1:3:6 in a sepsis patient and 16:47:37 (A/A:A/T:T/T) in healthy controls.2. Analysis of various mediator genes expression in animal models: (1) A rat model of radiation fibrosis: We analyzed the gene expressions of chemokines, MCP-1, MIP-1alpha, beta, MIP-2, lymphotactin, and fractalkine and found the augmented expression of MCP-1, TARC mRNA at four weeks after irradiation, and a slightly increased expression of MCP-1, MIP-1alpha at 8 weeks.2) A mouse model of burn-primed sepsis and ARDS:The 72 hour survival was 100% in no priming group and 0% in a bum primed model. The plasma cytokine increase was single-peaked in no priming group and two-peaked in burn-primed group. Furthermore, liver IL-18 content was significantly higher in burn-primed group than in sham group one week after the treatment.