摘要
目的:研究同型半胱氨酸(Hcy)对小鼠成神经瘤细胞(N2a)毒性作用并探讨其对p ERK1/2蛋白表达的影响。方法:培养的N2a细胞加入不同浓度Hcy,随机分为4组:分别为正常对照组及Hcy低、Hcy中、Hcy高浓度组,根据预实验结果确定以上4组Hcy干预浓度分别为0、30、300和1 000μmol/L。作用72 h后,用酶标仪测定细胞培养液中乳酸脱氢酶(LDH)活性,MTT法检测细胞增殖情况,蛋白质免疫印迹法检测N2a细胞p ERK1/2蛋白表达水平。结果:与正常对照组相比,Hcy低、中、高浓度组细胞培养液中LDH活性均显著升高,差异具有统计学意义(P值分别为0.013、0.008和0.011,P<0.05)。低、中、高浓度Hcy作用后,MTT检测细胞增殖活力下降,OD值降低,与对照组比较,差异具有统计学意义(P值分别为0.01、0.006和0.009,P<0.05)。各Hcy干预组p ERK1/2蛋白表达量降低,且中、高浓度Hcy组p ERK1/2蛋白表达量与对照组相比差异具有统计学意义(P值分别为0.038、0.007,P<0.05)。结论:Hcy能抑制磷酸化的ERK1/2蛋白表达,从而对N2a产生毒性作用,进而抑制N2a增殖,提示降低血清中Hcy水平可以对神经细胞产生保护作用。
Objective： To explore the neurotoxicity ofhomocysteine （Hcy） on mouse neuroblastoma cells （N2a） and its effect on pERK 1/2 protein level. Methods ： Cultured N2a cells were divided into four groups according to the concentrations of homocysteine ： control group （ Hcy 0 μmol/L）, low-Hcy （ 30 μmol/L）, moderate-Hcy （ 300 μmol/L）, high-Hcy （ 1 000 μmol/L） groups. After 72 h Hcy treatment, the toxicity of N2a cells was detected by LDH （laeate dehydrogenase） assay kit, cell proliferation ability was detected by MTT methods, and the protein expression of pERK1/2 was detected by western blot. Results： The LDH activity in Hcy-treated groups was increased compared with control group （P〈0.05）. The cells proliferation ability was decreased after Hey treatment, and OD values were reduced compared with control group （P〈0.05）. The protein expression of pERK1/2 decreased after Hcy treatment, and significant differences among the moderate, high Hey dose groups and control group were found （P〈0.05）. Conclusion： The toxic effect of Hey on N2a cells may be caused by reduced ERK1/2 protein phosphorylation expression. It suggests that a low serum Hcy level may have a protective effect on neural cells.
出处
《天津医科大学学报》
2015年第1期6-8,17,共4页
Journal of Tianjin Medical University
基金
国家自然科学基金资助项目(81373003)
中国博士后科学基金资助项目(2014M550148)
