摘要
目的 :探讨地塞米松对人α1( )原胶原基因调控序列的作用。方法 :用组织块法原代培养人皮肤成纤维细胞并进行传代培养作为本研究的模型细胞。 (1)采用 Brd U掺入的 EL ISA法 ,测定经不同浓度地塞米松处理 2 4h后成纤维细胞的增殖情况。 (2 )构建含长短不一的人α1( )原胶原基因 5′侧翼序列与氯霉素乙酰基转移酶 (CAT)报告基因的 3种重组质粒 ;以脂质体法转染人皮肤成纤维细胞 ;EL ISA法测定不同浓度地塞米松作用 2 4h后 ,转染了 3种重组质粒的成纤维细胞的 CAT表达量。结果 :(1)在加入 2 % FCS或 10 % FCS的培养条件下 ,1× 10 - 9～ 1× 10 - 4 m ol/ L 地塞米松处理 2 4h后 ,各浓度组成纤维细胞增殖值之间以及与对照组之间差异均无显著性 (P>0 .0 5 )。 (2 ) 3种重组体转染成纤维细胞 ,并经地塞米松处理 2 4h后 ,转染细胞的 CAT相对表达量测定 :地塞米松组与对照组之间、地塞米松较高浓度组 (1× 10 - 5 m ol/ L)与较低浓度组 (1× 10 - 6mol/ L)之间 ,差异均有显著性 (P<0 .0 5 )。结论 :地塞米松在 1× 10 - 9～ 1× 10 - 4 mol/ L 浓度范围内 ,对所研究的模型细胞——人皮肤成纤维细胞增殖无明显作用。地塞米松对胶原基因启动序列具有负性调控作用 ,且存在剂量依赖关系。
Objective: To investigate the effects of dexamethasone on the promoter activities of human α1(Ⅰ) procollagen gene. Methods: Fibroblasts from human skin were primarily cultured and subcultured. (1)The effects of dexamethasone on the human skin fibroblasts were determined by BrdU incorporation into DNA of fibroblast. (2)Three plasmids containing various lengths of 5′flank sequence of human α1(Ⅰ) procollagen gene and CAT as reporter gene were constructed, and were transfected into the human skin fibroblasts by FuGENE transfection reagent. The effects of dexamethasone on 3 plasmids were determined by CAT ELISA. Results: (1)After 24 h of treatment with 1×10 -9 1×10 -4 mol/L dexamethasone in DMEM containing 2% and 10% FCS, BrdU incorporation into DNA showed no difference ( P >0.05). (2)The 3 plasmids were transfected into fibroblasts and then treated with 10 -5 mol/L and 10 -6 mol/L dexamethasone for 24 h, relative CAT value were different between dexamethasone and control, higher dexamethasone(1×10 -5 mol/L) and lower dexamethasone(1×10 -6 mol/L) ( P <0.05). Conclusion: Dexamethasone has no effects on the proliferation of human skin fibroblasts, and it has negative effect on the promoter activity of human α1(Ⅰ) procollagen gene, which is dose dependent. [
出处
《第二军医大学学报》
CAS
CSCD
北大核心
2000年第8期718-720,共3页
Academic Journal of Second Military Medical University
基金
国家自然科学基金!资助项目 (39870 76 1)
