The retina of Wistar rats within 1-3 days of birth were dissociated into a retinal ceil suspension using 0.05% trypsin digestion. The cell suspension was incubated in Dulbecco＇s modified Eagle＇s medium for 24 hours,...The retina of Wistar rats within 1-3 days of birth were dissociated into a retinal ceil suspension using 0.05% trypsin digestion. The cell suspension was incubated in Dulbecco＇s modified Eagle＇s medium for 24 hours, followed by neurobasal medium for 5-7 days. Nissl staining showed that 79.86% of primary cultured retinal cells were positive and immunocytochemical staining showed that the purity of anti-neurofilament heavy chain antibody-positive cells was 71.53%, indicating that the primary culture system of rat retinal neurons was a reliable and stable cell system with neurons as the predominant cell type. The primary cultured retinal neurons were further treated with 0, 5.5, 15, 25, and 35 mM glucose for 24, 48, and 72 hours. The thiazolyl blue tetrazolium bromide test and flow cytometry showed that with increasing glucose concentration and treatment duration, the viability of retinal neurons was reduced, and apoptosis increased. In particular, 35 mM glucose exhibited the most significant effect at 72 hours. Thus, rat retinal neurons treated with 35 mM glucose for 72 hours can be used to simulate a neuronal model of diabetic retinopathy.展开更多
基金supported by the Department of Health of Hunan Province, No. B2009-050the Science and Technology Foundation of Hunan Province, No.2012FJ4077
文摘The retina of Wistar rats within 1-3 days of birth were dissociated into a retinal ceil suspension using 0.05% trypsin digestion. The cell suspension was incubated in Dulbecco＇s modified Eagle＇s medium for 24 hours, followed by neurobasal medium for 5-7 days. Nissl staining showed that 79.86% of primary cultured retinal cells were positive and immunocytochemical staining showed that the purity of anti-neurofilament heavy chain antibody-positive cells was 71.53%, indicating that the primary culture system of rat retinal neurons was a reliable and stable cell system with neurons as the predominant cell type. The primary cultured retinal neurons were further treated with 0, 5.5, 15, 25, and 35 mM glucose for 24, 48, and 72 hours. The thiazolyl blue tetrazolium bromide test and flow cytometry showed that with increasing glucose concentration and treatment duration, the viability of retinal neurons was reduced, and apoptosis increased. In particular, 35 mM glucose exhibited the most significant effect at 72 hours. Thus, rat retinal neurons treated with 35 mM glucose for 72 hours can be used to simulate a neuronal model of diabetic retinopathy.
