Otolaryngology-Head and Neck Surgery Department, Ningbo First Hospital, Ningbo 315000, China.
J Zhejiang Univ Sci B. 2012 Dec;13(12):997-1005. doi: 10.1631/jzus.B1200055.
To evaluate the effects of mitomycin on the growth of human dermal fibroblast and immortalized human keratinocyte line (HaCat cell), particularly the effect of mitomycin on intracellular messenger RNA (mRNA) synthesis of collagen and growth factors of fibroblast.
The normal dermal fibroblast and HaCat cell were cultured in vitro. Cell cultures were exposed to 0.4 and 0.04 mg/ml of mitomycin solution, and serum-free culture medium was used as control. The cellular morphology change, growth characteristics, cell proliferation, and apoptosis were observed at different intervals. For the fibroblasts, the mRNA expression changes of transforming growth factor (TGF)-β1, basic fibroblast growth factor (bFGF), procollagen I, and III were detected by reverse transcription polymerase chain reaction (RT-PCR).
The cultured normal human skin fibroblast and HaCat cell grew exponentially. A 5-min exposure to mitomycin at either 0.4 or 0.04 mg/ml caused marked dose-dependent cell proliferation inhibition on both fibroblasts and HaCat cells. Cell morphology changed, cell density decreased, and the growth curves were without an exponential phase. The fibroblast proliferated on the 5th day after the 5-min exposure of mitomycin at 0.04 mg/ml. Meanwhile, 5-min application of mitomycin at either 0.04 or 0.4 mg/ml induced fibroblast apoptosis but not necrosis. The apoptosis rate of the fibroblast increased with a higher concentration of mytomycin (p<0.05). A 5-min exposure to mitomycin at 0.4 mg/ml resulted in a marked decrease in the mRNA production of TGF-β1, procollagen I and III, and a marked increase in the mRNA production of bFGF.
Mitomycin can inhibit fibroblast proliferation, induce fibroblast apoptosis, and regulate intracellular protein expression on mRNA levels. In addition, mitomycin can inhibit HaCat cell proliferation, so epithelial cell needs more protecting to avoid mitomycin's side effect when it is applied clinically.
评价丝裂霉素对人真皮成纤维细胞和永生化人角质形成细胞（HaCat 细胞）生长的影响，特别是丝裂霉素对成纤维细胞细胞内信使 RNA（mRNA）合成胶原和生长因子的影响。
体外培养正常真皮成纤维细胞和 HaCat 细胞。将细胞培养物暴露于 0.4 和 0.04mg/ml 的丝裂霉素溶液中，以无血清培养液作为对照。在不同时间间隔观察细胞形态变化、生长特征、细胞增殖和凋亡。对于成纤维细胞，通过逆转录聚合酶链反应（RT-PCR）检测转化生长因子（TGF）-β1、碱性成纤维细胞生长因子（bFGF）、I 型和 III 型前胶原的 mRNA 表达变化。
培养的正常人皮肤成纤维细胞和 HaCat 细胞呈指数生长。0.4 或 0.04mg/ml 的丝裂霉素作用 5min 可明显抑制成纤维细胞和 HaCat 细胞的增殖，呈剂量依赖性。细胞形态发生变化，细胞密度降低，生长曲线无指数期。在丝裂霉素 0.04mg/ml 作用 5min 后第 5 天，成纤维细胞开始增殖。同时，0.04 或 0.4mg/ml 的丝裂霉素作用 5min 诱导成纤维细胞凋亡而不导致坏死。成纤维细胞的凋亡率随丝裂霉素浓度的增加而增加（p<0.05）。0.4mg/ml 的丝裂霉素作用 5min 可明显降低 TGF-β1、I 型和 III 型前胶原的 mRNA 产生，明显增加 bFGF 的 mRNA 产生。
丝裂霉素可抑制成纤维细胞增殖，诱导成纤维细胞凋亡，并调节细胞内蛋白表达的 mRNA 水平。此外，丝裂霉素可抑制 HaCat 细胞增殖，因此在临床上应用时需要更多保护上皮细胞以避免丝裂霉素的副作用。
