摘要
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 。
Objective To investigate the fragment of high promoter activity in human α2(Ⅰ) procollagen gene and to explore the potential effect of cytokines on promoter activity. Methods Five chimeric genes were constructed with various lengths of sequences between 2.4kb upstream of the initiation of transcription of the human α2(Ⅰ) procollagen gene and 58bp downstream of this site. The sequences were fused to chloramphenicol acetyltransferase(CAT) reporter gene. These recombinant plasmids were transiently transfected into human skin fibroblast by FuGENE 6 transfection method. The putative promoters activities were tested and compared by CAT measurement in transfected cells. Results The highest CAT expression was demonstrated in construction driven by -2292 to +58bp, -1476 to +58bp fragment. TGF-β, IGF-1 increased promoter activity of human α2(Ⅰ) procollagen gene. Conclusion TGF-β, IGF-1 increasedⅠcollagen synthesis by regulating promoter element in procollagen gene.
出处
《中华微生物学和免疫学杂志》
CAS
CSCD
北大核心
2002年第1期58-61,共4页
Chinese Journal of Microbiology and Immunology
基金
国家自然科学基金资助项目 ( 39870 30 1)
