PURPOSE: 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.
PURPOSE: 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.
Authors: Tobias D Henning; Rakhee Gawande; Aman Khurana; Sidhartha Tavri; Lydia Mandrussow; Daniel Golovko; Andrew Horvai; Barbara Sennino; Donald McDonald; Reinhard Meier; Michael Wendland; Nikita Derugin; Thomas M Link; Heike E Daldrup-Link Journal: Mol Imaging Date: 2012-06 Impact factor: 4.488
Authors: Alexander Nedopil; Christopher Klenk; Cy Kim; Siyuan Liu; Mike Wendland; Daniel Golovko; Tibor Schuster; Barbara Sennino; Donald M McDonald; Heike E Daldrup-Link Journal: Invest Radiol Date: 2010-10 Impact factor: 6.016