摘要
目的探讨增塑剂邻苯二甲酸二丁酯对类神经细胞PC12细胞增殖能力的影响及其对PC12细胞凋亡相关基因表达的调控,为接触人群可能产生远期效应的生物学检测提供参考评价指标。方法体外培养类神经细胞株PC12细胞,培养基分别含有0,12.5,25,50 mg/L邻苯二甲酸二丁酯,作用24,48,72和96 h后,用细胞计数CCK-8法检测细胞的存活和增殖,台盼蓝染色绘制生长曲线,DNA凝胶电泳观察DNA-Ladder,RT-PCR检测p53,bcl-2,Bax等凋亡相关基因表达变化情况。结果邻苯二甲酸二丁酯在12.5 mg/L时即表现出显著的抑制PC12细胞增殖的作用(P<0.01),且随剂量增加抑制作用增强,同时出现DNA断裂阳性;凋亡相关基因均有表达,其中p53基因、Bax基因表达增加,bcl-2基因表达降低,且呈剂量相关关系(P<0.01)。结论邻苯二甲酸二丁酯暴露可抑制PC12细胞增殖并诱导PC12细胞凋亡,同时通过调节凋亡相关基因p53、Bax、bcl-2基因的表达水平而在其凋亡发生中起重要作用。推测邻苯二甲酸二丁酯长期低剂量暴露的累积效应与阿尔茨海默病以及其他神经细胞凋亡相关性疾病的发生发展可能存在相关关系。
Objective To study the effect of the plasticizer-dibutyl phthalate （DBP） on proliferation of pheochromocytoma （PC12） cells （neural-like cells） and its regulation of the expression of PC12 cell apoptosis-relating genes, and to further provide the indicators for reference and assessment in biological assays of possible long-term effects on the contact crowd. Methods PC12 cell strains were incubated in vitro in culture media with 0, 12.5, 25 and 50 mg/L DBP for 24, 48, 72 and 96 h, respectively. Cell counting kit-8 （CCK-8） was used to examine the survival and proliferation of cells, the trypan blue dye exclusion method was used to draw the growth curve, DNA gel electrophoresis was performed to observe DNA-Ladder and reverse transcriptase-polymerase chain reaction （RT-PCR） was applied to determine the changes in mRNA expressions of apoptosis-relating genes like p53, bcl-2 and Bax. Results The CCK-8 assay revealed that DBP evidently inhibited the proliferation of PC12 cells at the dose of 12.5 mg/L （P〈0.01）, its inhibitory effect increased with the increase in the DBP dose and DNA fragmentation （positive） occurred simultaneously. DNA gel electrophoresis showed that the apoptosis of PC12 cells could be induced by DBP （12.5-50 mg/L, 48 h）. And RT-PCR showed that DBP promoted mRNA expressions of p53 and Bax and inhibited bcl-2 mRNA expression; the expression of apoptosis-relating genes was correlated with the DBP dose （P〈0.01）. Conclusion DBP exposure may inhibit the proliferation of PC12 cells and induce PC12 cell apoptosis; meanwhile, through regulating the mRNA expression levels ofapoptosis-relating genes like p53, Bax and bcl-2, it plays an important role in the development of PC12 cell apoptosis. It can be predicted that the cumulative effects of long-term low-dose DBP might be correlated with the occurrence and development of Alzheimer＇s disease （AD） and other neural cell apoptosis-relating diseases.
出处
《中华神经医学杂志》
CAS
CSCD
2007年第9期903-908,共6页
Chinese Journal of Neuromedicine
基金
"十五"国家科技攻关课题(2003BA869C)
重庆市科委自然科学基金(2007BB7006)
