5-氮杂-2-脱氧胞苷对人瘢痕疙瘩成纤维细胞的影响

Effects of 5-aza-2-deoxycytidine on human keloid fibroblasts

摘要:

目的：探讨甲基化酶抑制剂5-氮杂-2-脱氧胞苷对人瘢痕疙瘩成纤维细胞生长的抑制作用及对人瘢痕疙瘩成纤维细胞相关因子的影响。方法：收集人瘢痕疙瘩以及周围正常皮肤标本原代培养成纤维细胞，分为瘢痕疙瘩成纤维细胞5-氮杂-2-脱氧胞苷药物干预组、瘢痕疙瘩成纤维细胞对照组、正常皮肤成纤维细胞5-氮杂-2-脱氧胞苷药物干预组及正常皮肤成纤维细胞对照组4组，用RT-PCR检测各组转化生长因子－β1（transforming growth factor-β1，TGF-β1）mRNA、Smad7mRNA的表达，流式细胞术（flow cytometry，ＦＣＭ）检测各组细胞的周期及凋亡。结果：人瘢痕疙瘩成纤维细胞较正常皮肤成纤维细胞TGF-β1mRNA表达增高（P=0.001），Smad7mRNA表达降低（P=0.001），细胞周期停滞于G0/G1期（P=0.035）及细胞凋亡（P=0.006）比例降低；5-氮杂-2-脱氧胞苷干预人瘢痕疙瘩成纤维细胞后，TGF-β1mRNA表达降低（P=0.003），Smad7mRNA回升（P=0.000），ＦＣＭ显示瘢痕疙瘩成纤维细胞停滞于G0/G1期比例增加（P=0.000）并且细胞凋亡率增加（P=0.047），而正常皮肤成纤维细胞组无明显变化（P均>0.05）。结论：甲基化酶抑制剂5-氮杂-2-脱氧胞苷可影响瘢痕疙瘩形成相关因子的表达并且可抑制瘢痕疙瘩成纤维细胞的增殖及促进其凋亡，而对正常皮肤的成纤维细胞无明显影响。瘢痕疙瘩的发病可能与相关基因甲基化有关，5-氮杂-2-脱氧胞苷可能成为瘢痕疙瘩治疗的新选择。

Abstract:

Objective：To investigate the inhibition effects of methylase inhibitor 5-aza-2-deoxycytidine on human keloid fibroblasts，and the effects of 5-aza-2-deoxycytidine on related cytokines of human keloid fibroblasts. Methods：Human keloid and surrounding normal skin samples were collected for primary cultured fibroblasts. Cells were divided into keloid fibroblast experimental group，normal skin fibroblast experimental group，keloid fibroblast control group and normal skin fibroblast control group. 5-aza-2-deoxycytidine was used to intervene the experimental groups. Expressions of TGF-β1mRNA and Smad7mRNA in each group were detected with RT-PCR. Effects of 5-Aza-2-deoxycytidine on the cell cycle and apoptosis of fibroblasts were analyzed with flow cytometry（FCM）. Results：Compared with those of normal skin fibroblast，expressions of TGF-β1mRNA was increased（P=0.001） but Smad7mRNA were decreased（P=0.001） in keloid fibroblast，meanwhile the proportion of cells in G0/G1 stage（P=0.035） and apoptosis cells were decreased（P=0.006）. Expressions of TGF-β1mRNA were decreased（P=0.003） and those of Smad7mRNA were elevated（P=0.000） in keloid fibroblasts intervened by 5-aza-2-deoxycytidine. FCM demonstrated that the proportions of cells in G0/G1 stage and apoptosis cells were increased（P=0.000，P=0.047） in keloid fibroblasts intervened by 5-aza-2-deoxycytidine. But changes in normal skin fi－broblast group were insignificant（P >0.05）. Conclusions：Methylaze inhibitors，5-aza-2-deoxycytidine may inhibit the proliferation and promote the apoptosis of keloid fibroblast as well as influence expressions of its related cytokines，however，the effects on normal skin fibroblasts are insignificant. Pathogenesis of keloid may relate with the methylation of certain genes. 5-aza-2-deoxycytidine may be a new choice in the treatment of pathological scar.