N-甲基-D-天门冬氨酸和地塞米松对体外培养鼠视网膜神经节细胞凋亡作用观察

Effect of N-Methyl-D-Aspartate and Dexamethasone on Apoptosis of Rat Cultured Retinal Ganglion Cells

目的:研究N-甲基-D-天门冬氨酸(N-methyl-D-aspartate NMDA)及地塞米松对体外培养视网膜神经节细胞(RGCs)的作用。方法:出生1～3天Sprague-Dawley(SD)乳鼠做体外RGCs培养。NMDA按20～500μmol/L加入两组混合培养的RGCs中,24小时后一组用Hochest33258检测凋亡细胞。另一组加0.4%台盼蓝,计数拒染细胞,并算出细胞存活率。地塞米松按1×10^(-4)、1×10^(-5)及1×10^(-6)mol/L分别加入混合培养及纯化培养的RGCs中,对照组不加,24小时后用Hochest33258检测凋亡细胞。结果:混合培养RGCs经不同浓度NMDA处理均可见明显的凋亡细胞形态学改变。对照组则不明显。而且其中细胞的存活率与NMDA浓度呈反相关。不同浓度地塞米松处理混合培养RGCs,经Hochest33258检测均未发现有凋亡形态学改变。而纯化培养RGCs实验组及对照组均有明显凋亡细胞形态学改变。结论:NMDA对体外混合培养RGCs有诱导细胞凋亡的作用;地塞米松则对培养RGCs无诱导细胞凋亡作用。纯化的RGCs生长24小时,不加神经营养因子Neurotrophic Factors(NTFs)有细胞凋亡的形态学改变。眼科学报1999 15:65—69。

Purpose:To investigate the effect of N-methyl-D-aspartate(NMDA) and Dexametha-sone on cultured rat retinal ganglion cells (RGCS). Methods :RGCs were obtained from 1 -3 days old SD rats. In two groups, exposure of cocultured ganglion cells to N-methyl-D-aspartate (20 - 500μmol/L) lasted for 24hours. Apoptotic cells were identified by Hochest 33258 in one group; In another group,0.4%Trypan blue dye was added. Cells excluding the dye were counted, and the survival rate of cells was determined by the ratio of the excluding cells in the experiment over in the control; Cocultured RGCs and purified RGCs were exposed to Dexamethasone(1 × 10-4, 1X 10-5and 1 × 10-6 mol/L). After 24 hours, apoptotic cells were identified by Hochest33258. Result: Cocultured RGCs showed distinct morphological appearance of apoptotic cells when they were exposed to NMDA. The survival rate of cells was dose - related to the concentration of NMDA. Cocultured RGCs didn't show typical apoptotic appearance at 24 hours after exposure to Dexamethasone. But purified RGCs did so, even in the control group. Conclusion: NMDA has the effect of inducing cocultured RGCs apoptosis. Dexam-ethasone had no this effect on RGCs. Without additinal neurotrophic factors, purified RGCs may become apoptotic cells when cultured in vitro for 24 hours. Eye Science 1999; 15:65-69.