Dai Ye-Qin, Fan Wei-Xin, Wu Lei, Li Mei-Yun
Department of Dermatology, First Affiliated Hospital of Nanjing Medical University, Nanjing 210029, China.
Zhongguo Yi Xue Ke Xue Yuan Xue Bao. 2007 Apr;29(2):217-21.
To investigate the feasibility of stable transfection of human hypoxia inducible factor-1alpha (HIF-1alpha) gene into fibroblasts cells and the effects of supernatant from the transfected cell culture on hair follicle cells.
PcDNA-HIF1alpha was stably transfected into fibroblasts cells with lipofectamine 2000. Expression of HIF-1alpha was observed by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. The supernatant was obtained to detect the expression of vascular endothelial growth factor (VEGF) by ELISA. The mRNA expression of basic fibroblast growth factor (bFGF) was detected by RT-PCR. MTT was used to detect the activity of fibroblasts cells and dermal sheath cells added with supernatant.
PcDNA-HIF1alpha was successfully transfected into fibroblasts cells. HIF-1alpha could be detected by RT-PCR and Western blot. The expression of VEGF in the supernatant of cells transfected with PcDNA-HIF1alpha was detected. The mRNA expression of bFGF was significantly higher than in the control group (P < 0.01). MTT showed the activity of cells added with supernatant was enhanced (P < 0.05).
PcDNA-HIF1alpha can stably transfected into fibroblasts cells, and the expressed HIF-1alpha induces the expression of VEGF and bFGF, and the expressed VEGF enhances the activity of cells.
探讨将人缺氧诱导因子-1α（HIF-1α）基因稳定转染至成纤维细胞的可行性以及转染细胞培养上清液对毛囊细胞的影响。
用脂质体2000将PcDNA-HIF1α稳定转染至成纤维细胞。通过逆转录-聚合酶链反应（RT-PCR）和蛋白质免疫印迹法观察HIF-1α的表达。获取上清液，采用酶联免疫吸附测定法（ELISA）检测血管内皮生长因子（VEGF）的表达。用RT-PCR检测碱性成纤维细胞生长因子（bFGF）的mRNA表达。采用噻唑蓝比色法（MTT）检测添加上清液后的成纤维细胞和毛乳头细胞的活性。
PcDNA-HIF1α成功转染至成纤维细胞。通过RT-PCR和蛋白质免疫印迹法可检测到HIF-1α。检测了用PcDNA-HIF1α转染的细胞上清液中VEGF的表达。bFGF的mRNA表达明显高于对照组（P<0.01）。MTT结果显示添加上清液的细胞活性增强（P<0.05）。
PcDNA-HIF1α可稳定转染至成纤维细胞，所表达的HIF-1α诱导VEGF和bFGF表达，所表达的VEGF增强细胞活性。
