氟对原代培养大鼠海马细胞NCAM mRNA和蛋白表达的影响

目的 探讨氟对原代大鼠海马细胞生长发育、神经细胞黏附分子（NCAM）mRNA及蛋白表达的影响。方法 原代培养大鼠海马细胞暴露于20、40和80μg/ml氟化钠24h后,观察染氟前后海马细胞生长情况,采用噻唑蓝比色试验、逆转录-聚合酶链反应和蛋白印迹技术分别检测大鼠海马细胞存活情况、NCAMmRNA和蛋白表达水平。结果与对照组相比,80μg/ml剂量组海马细胞数量减少,突起减少、变短,细胞存活率下降。40、80μg/ml剂量组NCAM mRNA水平均显著低于对照组,20μg/ml剂量组NCAM mRNA水平虽与对照组之间差异无统计学意义,但存在下降的趋势,NCAM mRNA水平与氟浓度呈现一定的剂量效应关系,且随染氟浓度升高而降低。40、80μg/ml剂量组细胞NCAM-180蛋白表达水平显著低于对照组,各染氟组细胞NCAM-140蛋白表达水平显著低于对照组,80μg/ml剂量组细胞NCAM-120蛋白表达水平显著低于对照组。结论 氟可抑制海马细胞生长和存活,并使NCAMmRNA和蛋白表达水平下降,氟对发育中海马损伤可能是其神经毒性的作用位点之一。

Effects of fluoride on neural cell adhesion molecules mRNA and protein expression levels in primary rat hippocampal neurons

OBJECTIVE: To investigate the effects of fluoride on the growth and viability, and mRNA and protein expression levels of neural cell adhesion molecules (NCAM) in primary rat hippocampal neurons. METHODS: The growth and development, the rate of cell survivor, and the mRNA and protein expression levels of NCAM were measured by MTT, RT-PCR, and Western blot respectively after the hippocampal neurons were incubated with 20, 40, and 80 microg/ml sodium fluoride for 24 hours in vitro. RESULTS: As compared with the control group, the number of cells, the length and number of neuritis, and rate of cell survivor were significantly decreased in 80 microg/ml fluoride-treated group (P < 0.05). The mRNA expression levels of NCAM in 40 and 80 microg/ml fluoride-treated groups were significantly lower than that in the control group and decreased with the increasing fluoride concentration. Compared with the control group, the mRNA expression level of NCAM in 20 microg/ml fluoride-treated group was decreased, but the difference was not statistically significant (P > 0.05). The NCAM-180 protein expression levels in 40 and 80 microg/ml fluoride-treated groups, the NCAM-140 protein expression levels in all fluoride-treated groups, and NCAM-120 protein expression level in 80 microg/ml fluoride-treated group were significantly lower than those in the control group (P < 0.05, P < 0.05, P < 0.05, respectively). CONCLUSION: Fluoride might restrain the growth and survival of rat hippocampal neurons, and decrease mRNA and protein expression levels of NCAM. The impairment of developmental hippocampus might be one of the neurotoxical target sites for fluoride toxicity.