脑源性神经营养因子对丙烯酰胺致NB-1细胞损伤的保护作用  

Study on effects of brain-derived neurotrophic factor in acrylamide-induced nerve damage in vitro

作　　者：肖经纬[1] 孙彦彦[1] 王秀会[1] 陈宵[1] 张斌[1] 李斌[1] 

机构地区：[1]中国疾病预防控制中心职业卫生与中毒控制所,北京100050

出　　处：《毒理学杂志》2014年第4期259-264,共6页Journal of Toxicology

基　　金：国家自然科学基金(30901219;81273110)

摘　　要：目的在体外实验条件下探讨脑源性神经营养因子(brain-derived neurotrophic factor,BDNF)对丙烯酰胺(acrylamide,ACR)所致神经损伤的干预效应。方法 MTT法检测不同剂量ACR(0、10、20、40、60、80和100μg/ml)作用时NB-1细胞的相对存活率并计算半数抑制浓度(IC50);根据ACR细胞毒性测试结果设对照组、ACR染毒组(60μg/ml ACR)、BDNF干预组:BDNF1(50 ng/ml BDNF+60μg/ml ACR)、BDNF2(75 ng/ml BDNF+60μg/ml ACR)、BDNF3(100 ng/ml BDNF+60μg/ml ACR);光学显微镜观察神经细胞形态变化,MTT法检测细胞相对存活率,流式细胞术检测胞内游离钙离子浓度;Western-blot检测突触素I(synapsin I)蛋白表达。结果 ACR对NB-1细胞的IC50为69.8μg/ml(48 h)。50和75 ng/ml BDNF对ACR所致神经损伤具有一定的干预作用,与60μg/ml ACR单独染毒组比较,神经细胞形态异常减轻,细胞相对存活率增加,胞内钙离子浓度降低,synapsin I蛋白含量增加(P<0.05);BDNF剂量增加到一定水平(100 ng/ml),干预效果减弱,细胞相对存活率、胞内钙离子浓度及synapsin I蛋白表达与60μg/ml ACR单独染毒组差异无统计学意义(P>0.05)。结论适当剂量的BDNF在体外条件下对ACR所致神经损伤具有一定的保护作用,可能是通过对抗胞内钙超载,上调synapsin I蛋白表达,维持突触结构完整来实现的。Objective The present study was designed to investigate the effects of brain-derived neurotrophic factor（BDNF） on acrylamide（ACR）-induced nerve damage in vitro.Methods NB-1 cells were treated with graded concentrations of ACR（0,10,20,40,60,80 and 100 μg/ml） for 48 h.Relative survival rate was measured by MTT and IC50 was calculated.According to the results of cytotoxicity of ACR on NB-1 cells,three groups were set,which were control group,60 μg/ml ACR treatment group and BDNF intervention groups：BDNF1（50 ng/ml BDNF ＋60 μg/ml ACR）,BDNF2（75 ng/ml BDNF ＋60 μg/ml ACR）,BDNF3（100 ng/ml BDNF ＋60 μg/ml ACR）.Morphological changes of cells were observed by light microscopy.Relative survival rate was measured by MTT.Protein level of synapsin I was measured by western blotting.[Ca2 ＋]i was measured by flow cytometry.Results ACR had toxic effect on NB-1 cells with IC50 of 69.8 μg/ml（48 h）.Appropriate dose ranges of BDNF（50-75 ng/ml） had protective effects on NB-1 cells damaged by ACR.Relative survival rate and synapsin I protein levels of cells treated with 50 ng/ml or 75 ng/ml BDNF combined with 60 μg/ml ACR were higher than that of cells treated with 60 μg/ml ACR alone（P 〈 0.05）,while [Ca2 ＋]I was lower than that of 60 μg/ml ACR treatment group（P 〈 0.05）.But that protective effect was reduced when the dose of BDNF was equal to 100 ng/ml,which was showed as relative survival rate,synapsin I protein level and [Ca2 ＋]I of cells treated with 50 ng/ml or 75 ng/ml BDNF combined with 60 μg/ml ACR had no significant difference with cells treated with 60 μg/ml ACR only（P 〉 0.05）.Conclusion ACR had toxic effect on matured NB-1 cells.BDNF of appropriate levels attenuated ACR-induced nerve damage in vitro.Possible mechanisms were against intracellular calcium overload,raising synapsin I protein expression and maintaining synaptic structure.

关 键 词：丙烯酰胺 脑源性神经营养因子 突触素I 胞内游离钙 

分 类 号：R135.1[医药卫生—劳动卫生]