转化生长因子-β1诱导大鼠毛乳头细胞转分化为成纤维细胞的研究

Transdifferentiation of rat vibrissa dermal papilla cells into fibroblasts induced by transforming growth factor-beta 1

目的研究大鼠触须毛乳头细胞在转化生长因子-β1(TGF-β1)诱导作用下转分化为成纤维细胞的可能性,从新的角度探讨增生性瘢痕的形成机制。方法消化收集第4代毛乳头细胞,处理组以TGF-β1(10ng/mL)处理细胞4d;对照组加入正常培养基(不含胎牛血清的DMEM/F12培养液),每隔24h观察两组细胞生长形态及生长方式;分别用实时一步法RT-PCR和流式细胞仪检测细胞α平滑肌肌动蛋白(α-SMA)、波形蛋白(Vimentin)的mRNA表达和蛋白表达,Western blotting检测成纤维细胞特异蛋白(FSP1)的表达。结果流式细胞仪检测TGF-β1诱导48h、72h后α-SMA表达明显低于对照组(P<0.01);TGF-β1诱导48h、72h、96h后Vimentin表达明显高于对照组(P<0.01)。实时一步法RT-PCR检测Vimentin的mRNA表达逐渐增强,α-SMA的mRNA表达在48h后明显下降(P<0.01),但96h后表达又有所增加;Western blotting法检测FSP1表达在诱导48h后逐渐增加(P<0.01)。结论毛乳头细胞经TGF-β1诱导后,失去其典型的细胞形态及生长方式,而表现出成纤维细胞的细胞形态及生长方式;其相对特异性标记物α-SMA的表达下降,而成纤维细胞的相对特异性标记物Vimentin和FSP1表达增加。故TGF-β1可诱导大鼠毛乳头细胞转分化为成纤维细胞。

Objective To explore the possibility of transdifferentiation of dermal papilla cells into fibroblasts induced by transforming growth factor-beta 1 （TGF-β1）. Methods The fourth passage cells were digested and collected, which were treated with TGF-β1 （ 10 ng/mL） for 4 days. At 24-hour intervals a series of methods were used, including morphological observation under inverted phase contrast microscope, reverse transeriptase polymerase chain reaction （RT-PCR） to validate the mRNA expression of alpha smooth muscle actin and vimentin, flow cytometry to analyse the expression of alpha smooth muscle aetin and vimentin, and Western blotting to validate the expression of fibroblast-specific protein 1 （ FSP1 ）. Results Flow cytometry showed the expression of alpha smooth muscle actin of the treatment groups obviously lower than that of the control groups after induction for 48 h and 72 h. The expression of vimentin of the treatment groups was obviously higher than that of the control groups after induction for 48 h ,72 h and 96 h. Validated by RT-PCR, the mRNA expression of vimentin of the treatment groups gradually increased, and that of alpha smooth muscle actin decreased after induction for 48 h and 72 h but increased after induction for 96 h. Validated by Western blotting, the expression of fibroblast-specific protein 1 of the treatment groups gradually increased after induction for 48 h, 72 h and 96 h. Conclusions Induced by TGF-β1, dermal papilla cells lost its typical morphology and growth pattern and presented morphology and growth pattern of fibroblasts, expression of relative specific markers of dermal papilla cells （α-SMA） decreased, however, expression of relative specific markers of fibroblasts （Vimentin and FSP1 ） increased. So TGF-β1 could induce the transdifferentiation of rat vibrissa dermal papilla cells into fibroblasts.