苯并(a)芘对神经细胞的毒性及细胞色素P4501A1的诱导表达  

Relationship between cytotoxicity of neural cell and CYP1A1 expression induced by benzo(a)pyrene

作　　者：魏建宏[1] 张红梅[2] 牛侨[2] 聂继盛[2] 

机构地区：[1]山西医科大学汾阳学院解剖学教研室,山西汾阳032200 [2]山西医科大学劳动卫生教研室,太原030001

出　　处：《解剖学报》2012年第5期608-613,共6页Acta Anatomica Sinica

基　　金：国家自然科学基金资助项目(30800899;30471437);山西省自然科学基金资助项目(20051093)

摘　　要：目的探讨苯并(a)芘[B(a)P]对神经细胞毒性、细胞凋亡与细胞色素P4501A1(CYP1A1)诱导表达的关系。方法选用新生1～3d的SD大鼠,分离大脑皮质,进行神经元培养,在细胞培养第5天左右,选取生长良好的同批次神经细胞,以苯并(a)芘分别对神经细胞染毒,使苯并(a)芘终浓度分别为0μmol/L、10μmol/L、20μmol/L、40μmol/L。继续培养40h,应用cck8试剂盒检测神经细胞活力、Annexin V和PI双染法进行细胞凋亡的检测,应用RT-PCR法检测神经细胞CYP1A1mRNA的表达,免疫组织化学SABC法检测神经元CYP1A1蛋白的表达。结果随着苯并[a]芘浓度的增加,神经细胞活力降低,早期凋亡率逐渐增高,细胞活力中高剂量组与对照组比较差异均有统计学意义,早期凋亡率仅高剂量组与对照组比较差异有统计学意义,趋势检验表明,细胞活力降低、凋亡率增高具有剂量依赖性。而且随B(a)P剂量的增高,CYP1A1mRNA及蛋白表达增多,有剂量-反应关系,CYP1A1基因和蛋白表达与神经细胞凋亡率的相关分析表明,神经细胞凋亡率与CYP1A1mRNA表达呈正相关(r=0.831,P<0.01);与CYP1A1蛋白表达呈正相关(r=0.780,P<0.01)。结论苯并[a]芘可致神经细胞凋亡,神经细胞CYP1A1诱导表达是神经细胞损伤的关键因素。Objective To investigate the relationship between neural cell cytotoxicity, apoptosis and cytochrome P4501A1 （ CYPIA1 ） mRNA exposed to benzo （a） pyrene [ B （a） P . Methods Primary neural cells were dissociated from the cerebral cortex of 1-3 days old SD rats and cultured in a DMEM incubator at 37℃ After cultivation for 5 days, the well-grown neural cells were selected and treated with the concentrations 0, 10, 20 and 40μmol/L of B （a） P respectively. The neural cells were further cultivated for 40 hours. Neural cell vitality was detected using cck8 kit. Apoptosis rate was measured by flow cytometry using annexinV - FITC and propidium iodide （PI） staining. CYPIA1 mRNA was tested with RT-PCR technique. CYP1A1 mRNA protein expression was determined by immunohistochemistry SABC. Results With the increase of B（a） P concentration, the neural cell vitality declined in both the medium dose group and the high dose group compared with controls, and early apoptotic rate increased steadily only in the high dose group compared with controls. Trend test indicated that cell vitality declined and apoptotic rate increased in a dose dependent manner. Also, there was a dose-reaction relationship between B（a） P and CYP1A1 mRNA. The apoptotic rate of neural cell was positively correlated with CYPIA1 mRNA （r = O. 83i, P 〈 0. 01 ） and CYPIA1 protein expression （r = 0. 780, P 〈0.01 ）. Conclusion B （a） P may induce apoptosis of neural cell, and CYP1A1 induction may be the key factor to start the neural cell damage.

关 键 词：苯并(A)芘 细胞毒性 CYP1A1 AnnexinV和PI双染法 反转录-聚合酶链反应 免疫组织化学 大鼠 

分 类 号：R322[医药卫生—人体解剖和组织胚胎学]