Woo K I, Lee J
Department of Ophthalmology, Hanil General Hospital, Seoul, Korea.
Korean J Ophthalmol. 1995 Jun;9(1):19-25. doi: 10.3341/kjo.1995.9.1.19.
This study was conducted to investigate the effect of ascorbic acid on oxidative injury of cultured porcine retinal pigment epithelial (RPE) cells induced by t-butylhydroperoxide. The porcine RPE cells were cultured in Dulbecco's modified Eagle's medium and the culture medium was replaced with one containing 0.01 mM to 5 mM ascorbic acid and/or 0.2 mM t-butylhydroperoxide. After 2 hours incubation, the test medium was replaced with the control medium. The number of cells was counted with a Coulter counter after a 2-day incubation period. The medium was pretreated with 900 U/ml and the previous procedure was repeated to eliminate the toxic effects of hydrogen peroxide induced by ascorbic acid. Not only t-butylhydroperoxide (p < 0.01) but also ascorbic acid (p < 0.01) were found to have dose-dependent cytotoxicity on RPE cells. The cytotoxicity was more significant when both agents were added to the culture media. In the presence of catalase, the cytotoxicity of ascorbic acid became insignificant (p < 0.05). The cytotoxicity of t-butylhydroperoxide decreased when 1 mM and 5 mM of ascorbic acid was added to the culture media with catalase pretreatment (p = 0.0277). These results indicate that ascorbic acid was toxic to RPE cells in our culture model but this cytotoxicity was not detected in the presence of catalase. With catalase pretreatment, ascorbic acid in relatively high concentration provided protection against oxidative injury of t-butylhydroperoxide.
本研究旨在探讨抗坏血酸对叔丁基过氧化氢诱导的猪视网膜色素上皮（RPE）细胞氧化损伤的影响。将猪RPE细胞培养于杜尔贝科改良伊格尔培养基中，然后用含有0.01 mM至5 mM抗坏血酸和/或0.2 mM叔丁基过氧化氢的培养基替换。孵育2小时后，将测试培养基替换为对照培养基。孵育2天后，用库尔特计数器对细胞进行计数。培养基用900 U/ml进行预处理，并重复上述步骤以消除抗坏血酸诱导的过氧化氢的毒性作用。结果发现，不仅叔丁基过氧化氢（p < 0.01），而且抗坏血酸（p < 0.01）对RPE细胞均具有剂量依赖性细胞毒性。当两种试剂都添加到培养基中时，细胞毒性更为显著。在过氧化氢酶存在的情况下，抗坏血酸的细胞毒性变得不显著（p < 0.05）。在过氧化氢酶预处理的培养基中添加1 mM和5 mM抗坏血酸时，叔丁基过氧化氢的细胞毒性降低（p = 0.0277）。这些结果表明，在我们的培养模型中，抗坏血酸对RPE细胞有毒性，但在过氧化氢酶存在的情况下未检测到这种细胞毒性。经过氧化氢酶预处理后，相对高浓度的抗坏血酸可提供对叔丁基过氧化氢氧化损伤的保护作用。
