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目的 探索器官纤维化形成中调控Ⅰ型胶原基因高水平转录的启动片段及TGF β、PDGF BB、IGF 1等细胞因子对其活性的影响。方法 从人α2 (Ⅰ )胶原基因转录起始点上游 - 2 .4kb至+5 8bp的片段中 ,取长度不等的片段作为启动子与含氯霉素乙酰基转移酶 (CAT)报告基因的质粒组成5个重组体 ,转染上述重组体至正常人原代培养皮肤成纤维细胞 ,测定细胞CAT表达水平以比较各重组体的启动子活性 ,同时加入细胞因子 ,以测定其对Ⅰ型胶原启动序列的影响。结果 除 - 12 9～ +5 8bp序列外 ,其余 4个重组体CAT表达水平均较高 ,其中 - 2 2 92～ +5 8bp、- 1476～ +5 8bp序列具较强启动CAT表达活性 ,- 339～ +5 8bp、- 6 16～ +5 8bp片段次之。TGF β、IGF 1均能在一定程度上调人α2(Ⅰ )胶原基因启动活性。结论 人α2 (Ⅰ )胶原基因片段 - 2 2 92～ +5 8bp、- 1476～ +5 8bp、- 339～ +5 8bp有高启动活性 ,是进一步研究纤维化相关DNA结合蛋白的重要调控靶序列。TGF β、IGF 1促进胶原表达 ,与其上调胶原基因启动活性有关
OBJECTIVE: To explore the effects of high-level transcriptional promoter of TGF-β1, TGFβ, PDGF BB, IGF-1 and other cytokines on the activity of fibrosis in organ fibrosis. METHODS: The fragments ranging in length from 2.4 kb to +5.8 bp upstream of the transcriptional start point of human α2 (Ⅰ) collagen gene were used as promoters and plasmids containing the reporter gene of chloramphenicol acetyltransferase (CAT) The recombinant plasmids were transfected into primary recombinant human dermal fibroblasts, and the CAT levels were measured to compare the promoter activity of each recombinant, and the addition of cytokines to determine the type I collagen promoter Effect of sequence. Results The expression levels of CAT in four other recombinant plasmids were higher than those of - 12 9 ~ +5 8bp. Among them, -292 ~ +5 8bp and - 1476 ~ +5 8bp exhibited strong CAT activity, 339 ~ +5 8bp, - 6 16 ~ +5 8bp fragment second. Both TGF β and IGF 1 could up-regulate the activation of α2 (Ⅰ) collagen gene to a certain extent. Conclusion The human α2 (Ⅰ) collagen gene fragment with high activity of 2292 ~ +5 8bp, - 1476 ~ +5 8bp and - 339 ~ +5 8bp has high activity and is an important regulatory target for further study of fibrosis related DNA binding protein . TGF β, IGF 1 promote collagen expression, with its up-regulation of collagen gene promoter activity