三邻甲苯磷酸酯染毒鸡大脑组织中迟发性神经毒性相关蛋白的筛选  

Screening the proteins of organophosphoms ester-induced delayed neurotoxicity in the cerebral tissue of hens exposed to tri-ortho-cresyl phosphate

作　　者：张彦宁[1] 朴丰源[1] 吕飞燕[1] 吕虹[1] 

机构地区：[1]大连医科大学劳动卫生与环境卫生学教研室,116044

出　　处：《中华劳动卫生职业病杂志》2011年第5期364-366,共3页Chinese Journal of Industrial Hygiene and Occupational Diseases

基　　金：国家自然科学基金资助项目（30771819）;辽宁省教育厅项目（2008S073）

摘　　要：目的从三邻甲苯磷酸酯（TOCP）染毒鸡大脑组织中筛选迟发性神经毒性（OPIDN）相关差异表达蛋白，为进～步探讨OPIDN机制提供靶标蛋白。方法成年岁曼鹤母鸡32只随机分成1000mg／kgTOCP组（染毒组）、先给40mg／kgPMSF后24h再给予1000mg／kgTOCP组（PMSF干预组）、给予40mg／kgPMSF组（PMSF组）和给予生理盐水的对照组，每组8只。一次性染毒后第5天处死动物，低温环境下分离大脑，匀浆，高速离心提取总蛋白。通过双向凝胶电泳获得完整的全蛋自质图谱，运用图像分析软件（hnageMaster2D）对银离子染色的电泳图谱进行分析，并对所选取的蛋白点进行质谱鉴定。结果染毒组、PMSF干预组和PMSF组鸡大脑组织总蛋白中，分别检测到1185、1294和1063个蛋白点，对照组检测到1332个蛋白点。与对照组比较，染毒组、PMSF干预组和PMSF组匹配率分别为78．32％、79．56％、80．93％。与对照组比较，染毒组差异表达蛋白点有235个，其中，上调点有158个，下调点有77个。根据PMSF的作用特点，在TOCP组中选择与对照组比较差异明显且与PMSF干预组无明显差异的蛋白匹配点，共获得有102个。其中差异表达4倍以上且与PMSF组二次匹配差异无统计学意义的蛋白点有13个。对这些蛋白点做进一步质谱分析和鉴定，获得7个蛋白：homer-1b、Destrin蛋白、热休克蛋白70、真核翻译起始冈子、蛋白酶体α1亚基、乳酸脱氢酶B、代谢性谷氨酰胺合成酶。结论TOCP染毒鸡大脑组织中有112个筹异表达蛋白点，可能与OPIDN诱发有关，其中13个差异表达蛋白点可能与OPIDN诱发有密切的关联性。质谱鉴定出7个可能与OPIDN机制关联的蛋白。Objective To screen the proteins-with differential expression levels in the cerebral tissue of hens exposed to tri-ortho-eresyl phosphate （TOCP）,and to provide target proteins for studying the mechanism of organophosphoms ester-indueed delayed neurotoxicity （OPIDN）. Methods Thirty two adult Roman hens were randomly divided into four groups： TOCP group was exposed to 1000 mg/kg TOCP, PMSF group was exposed to 40 mg/kg PMSF, PMSF plus TOCP group was exposed to 40 mg/kg PMSF and after 24 h exposed to 1000 mg/kg TOCP, control group was exposed to normal saline. All hens exposed to chemicals by gastro-intestine for 5 days were sacrificed, and the cerebral tissue were disseeted and homogenized in ice hath. Total proteins extracted from the cerebral tissue were separated by isoeleetrie foeusing as the first dimension and SDS-PAGE as the second dimension. The 2-DE maps were visualized after silver staining and analyzed by Image Master 2D software. At last ,the expressed protein spots were identified by Mass spectrometry. Results From total proteins in TOCP group, the PMSF plus TOCP group and PMSF group, 1185, 1294 and 1063 spots were deteeted, respectively. One thousand three hundred thirty two spots from total proteins in control group were detected. The match rates of protein spots in TOCP group, the PMSF plus TOCP group and PMSF group were 78.32 %, 79.56 % and 80.93%, respectively. There were 235 protein spots with differential expression levels between TOCP group and control group, which included 158 up regulation spots and 77 down regulation spots. According to the PMSF features, there were 102 spots with differential expression levels between TOCP group and control group and without differential expression levels between TOCP group and PMSF plus TOCP group, among them there were 13 spots with 4 fold differential expression levels between TOCP group and control group and without differential expression levels between TOCP group and PMSF group. Seven protein spots （homer-lb, Destrin, heat shock protein 70

关 键 词：磷酸酯类 苯甲磺酰基氟化物 有机磷化合物 电泳 凝胶 双向 

分 类 号：R[医药卫生]