Mitomycin C potentiates ultraviolet-related cytotoxicity in corneal fibroblasts.
OBJECTIVE
To study the morphologic changes and cytotoxicity in corneal fibroblasts after mitomycin C (MMC) treatment, ultraviolet B (UVB) irradiation, or in combination.


METHODS
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.


RESULTS
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.


CONCLUSIONS
UV irradiation causes dose-dependent cytotoxicity in porcine corneal fibroblasts. MMC pretreatment potentiates UV-related cytotoxicity.
Chang
Shu-Wen
SW
Department of Ophthalmology, Far Eastern Memorial Hospital, Ban-Chiao, Taipei, Taiwan. swchang2007@ntu.edu.tw
Chou
San-Fang
SF
Chuang
Jia-Ling
JL
D000477:Alkylating Agents / Q000494:pharmacology*; D000818:Animals; D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects / Q000528:radiation effects; D002478:Cells, Cultured; D003131:Combined Modality Therapy; D003315:Cornea / Q000166:cytology*; D004307:Dose-Response Relationship, Radiation; D004357:Drug Synergism; D005347:Fibroblasts / Q000187:drug effects* / Q000201:enzymology / Q000528:radiation effects*; D005434:Flow Cytometry; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008858:Microscopy, Phase-Contrast; D016685:Mitomycin / Q000494:pharmacology*; D013552:Swine; D014466:Ultraviolet Rays
D000477:Alkylating Agents; D017304:Annexin A5; D016685:Mitomycin; D007770:L-Lactate Dehydrogenase