Chang Shu-Wen, Chou San-Fang, Chuang Jia-Ling
Department of Ophthalmology, Far Eastern Memorial Hospital, Ban-Chiao, Taipei, Taiwan.
Cornea. 2008 Jul;27(6):686-92. doi: 10.1097/01.ico.0000611400.04418.79.
To study the morphologic changes and cytotoxicity in corneal fibroblasts after mitomycin C (MMC) treatment, ultraviolet B (UVB) irradiation, or in combination.
Primary porcine corneal fibroblasts, passages 3-5, were treated with MMC (0.1 or 0.2 mg/mL, ie, 0.01% or 0.02%, for 5 minutes), UVB irradiation (2, 5, or 10 mJ/cm2), or in combination. Control cells were treated with culture medium as a sham procedure. Alterations in cell morphology were documented by phase-contrast microscopy; cellular apoptosis was evaluated by Annexin V/propidium iodide staining and analyzed by flow cytometry; cytotoxicity was evaluated by lactate dehydrogenase assay; and cell growth was studied by genomic DNA quantification with the PicoGreen assay.
UV irradiation induced significant dose-dependent corneal fibroblast rounding and detachment and cytotoxicity. MMC at 0.1 or 0.2 mg/mL induced considerable cell elongation and retarded cell proliferation at similar rates. MMC treatment alone did not cause significant apoptosis or cytotoxicity. However, MMC treatment before UV irradiation potentiated UV-related cytotoxicity proportional to the UV radiation dose.
UV irradiation causes dose-dependent cytotoxicity in porcine corneal fibroblasts. MMC pretreatment potentiates UV-related cytotoxicity.
研究丝裂霉素C（MMC）处理、紫外线B（UVB）照射或二者联合处理后角膜成纤维细胞的形态学变化和细胞毒性。
用MMC（0.1或0.2mg/mL，即0.01%或0.02%，处理5分钟）、UVB照射（2、5或10mJ/cm²）或二者联合处理第3-5代原代猪角膜成纤维细胞。对照细胞用培养基处理作为假手术。通过相差显微镜记录细胞形态的改变；通过膜联蛋白V/碘化丙啶染色评估细胞凋亡并通过流式细胞术分析；通过乳酸脱氢酶测定评估细胞毒性；通过PicoGreen测定法对基因组DNA进行定量研究细胞生长。
紫外线照射诱导角膜成纤维细胞出现明显的剂量依赖性变圆和脱离以及细胞毒性。0.1或0.2mg/mL的MMC诱导相当程度的细胞伸长并以相似速率抑制细胞增殖。单独的MMC处理未引起明显的细胞凋亡或细胞毒性。然而，紫外线照射前的MMC处理增强了与紫外线相关的细胞毒性，且与紫外线辐射剂量成比例。
紫外线照射在猪角膜成纤维细胞中引起剂量依赖性细胞毒性。MMC预处理增强了与紫外线相关的细胞毒性。
