Survival of rat retinal ganglion cell and growth of its processes in relation to the promoting effect of soluble mixed crystallin: An in vitro culture experiment

Aim: To observe the effects of blood serum and crystalline on retinal ganglion cell (RGCs) in different culture medium in order to probe into its promoting effect on survival and regeneration of RGCs following the crystalline lens injuring. Methods: The experiment was conducted in the Laboratory of Ophthalmology, Southwest Ophthalmology Hospital, Third Military Medical University of Chinese PLA from April to July 2004. Eight Long Evens rats were used for the extraction of dissoluble mixed crystalline; and 30 newly born 0 - 2 day Long Evens rats were used for culturing retinal ganglion cells. The RGCS were cultured with the following 3-group culture medium: 1 100 mL/L newly born calf serum + 900 mL/L DMEM/F12(newly born calf serum + DMEM/F12 group). 2 100 mL/L newly born calf serum + 100 mL/L fetal calf serum + 800 mL/L DMEM/F12(newly born serum + fetal calf serum + DMEM/F12 group). 3 100 mL/L fetal calf serum + 900 mL/L DMEM/F12 (fetal calf serum + DMEM/F12 group). Each group was divided into 6 subgroups, among which, 5 different concentrations of dissoluble mixed crystalline liquid (1, 0.5, 0.02, 0.01, 0.005 mg/L) were divided into 5 subgroups, 1 subgroup without dissoluble mixed crystalline liquid. The number of the survival cells and survival time were recorded following the cells were observed under the reverse phase microscope every day. The number of RGCs with process and length of the longest process were measured with Leica Qwin image analysis system after cultured for 2, 4, 6 days (5 cells chosen randomly per vision were measured, totally 30 visions, 150 processes). Results: 1 The survival time of the RGCs: the cultured cells in newly bone serum + DMEM/F12 group could survive for 6 to 7 days and those in the newly born calf serum + fetal calf serum + DMEM/F12 group and fetal calf serun + DMEM/F12 group could survive 7 to 8 days. 2 The number of RGCs with process and the length of the longest process: At the same cultured days (2, 4 and 6 days) and condition (the same concentration of crystallin at last), the number of RGCs in the newly calf serum + fetal calf serum + DMEM/F12 group was all larger than that in the newly born serum + DMEM/F12 group [adding 0.5 mg/L crystallin, (3.83 ± 4.53) vs (0.14 ± 0.38) per vision at day 2, (14.7 ± 9.62) vs (2.5 ± 3.89) per vision at day 4, P < 0.01; (20.13 ± 12.32) vs (5.43 ± 5.62) per vision at day 6, P < 0.05]; The length of process in the fetal calf serum + DMEM/F12 group was longer significantly than that in the newly calf born serum + fetal calf serum + DMEM/F12 group in 4 days (P < 0.01); The survival time and the length of the process in the group with mixed crystallin were superior to those in the group without crystallin. The effect of crystallin was obvious when its concentration was 0.01 mg/L. Conclusion: Interaction does not exist in the two groups after the crystallin and different culture mediums were addeds respectively. Crystallin and different culture mediums exert their effects independently. Dissoluble mixed crystallin can promote the survival and the growth of process of GRCs cultured in vivro.