| The Effect of Glucose on Anoxia-induced Cell Damage in the Cultured Human Retinal Pigment Epithelial (RPE) Cell. |
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Seung Young Yu, Dae Ho Kim, Jae Myung Kim, Hyung Woo Kwak |
1Department of Ophthalmology, Kyung Hee University Hospital, Korea. hwkwak@khmc.or.kr
2Kim Dae Ho Eye Clinic, Korea. |
| 무산소에 의한 배양 인체 망막색소상피 손상에서 Glucose의 영향 |
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유승영 ( Seung Young Yu ) , 김대호 ( Dae Ho Kim ) , 김재명 ( Jae Myung Kim ) , 곽형우 ( Hyung Woo Kwak ) |
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| Abstract |
PURPOSE
It is difficult to make the standard ischemic RPE culture model which is still not established. In ischemic model, oxygen and glucose are very important. To improve experimental model, we evaluate the effect of glucose on anoxia-induced human RPE cell damage. METHODS: Human RPE cells were incubated with 95% N2/5% CO2 and standard conditions (95% air/5% CO2) at 37degrees C, and various concentation of glucose were added to a glucose-free medium : no glucose, 0.2 mg/ml, 0.4 mg/ml, -2.0 mg/ml. On a daily basis, morphological and proliferation analysis was conducted using the inverted phase contrast microscope. The extent of cell damage was estimated through cell viability analysis. Cell viability was examined using PMS(phenazinemethosulphate)-MTS(3-(4.5 -dimethyl-thiazol-2-yl)-5-(3-caboxymethoxyphenyl)-2-(4-sulfo phenyl)-2H-tetrazolium, inner salt)assay after 24, 48, 72, and 96 hours. Single-stranded DNAs were detected with monoclonal antibody to prove cell apoptosis in cell damage. RESULTS: At slightly low glucose concentration(0.4 mg/ml), the cell viability of human RPE cells cultured at room air was maintained up to 96 hours, whereas that cultured at 95% N2 decreased after 48 hours. At the anoxic condition. the cell viability of human RPE cells cultured with the low glucose concentration (0.2 mg/ml) decreased after 24 hours, but those done with higher glucose concentrations (more than 0.8 mg/ml) were maintained up to 96 hours. In the cell death caused by the anoxic culture, apoptosis was not suggested by single-stranded DNA detection. CONCLUSIONS: Properly higher glucose concentration on culture medium improve cell viability and reduce the cell damage in human RPE culture in anoxic condition. This result helps to improve experimental ischemic model of RPE cell. |
| Key Words:
Ischemia model;Anoxia;Glucose;Retinal pigment epithelium (RPE) |
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