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目的从三邻甲苯磷酸酯(TOCP)暴露鸡的脊髓组织中筛选可能与调控微丝解聚作用相关的差异表达蛋白,为探讨有机磷化合物诱发的迟发性神经毒性(OPIDN)作用机制提供靶蛋白依据。方法 42只罗曼鹤母鸡随机分成1000 mg/kg TOCP组、预先给予40 mg/kg苯甲基磺酰氟(PMSF)后再投1000 mg/kg TOCP的干预组和生理盐水对照组,每组14只。染毒第5和20天,每组分别处死4只鸡,低温环境下分离脊髓,提取总蛋白。利用双向电泳结合质谱分析技术,筛选和鉴定可能与调控微丝解聚作用相关的差异表达蛋白。结果 TOCP组鸡于染毒第7日前后出现进行性共济失调和肌无力等OPIDN典型症状,起病从下肢远端部分开始且病变程度随时间逐渐加重直至全瘫,而其他组鸡在实验观察期间未见OPIDN症状。TOCP组鸡于暴露第5天,分别与对照组和PMSF前干预组比较,其脊髓组织肌动蛋白解聚因子Cofilin-1b分别下调3.4倍和2.8倍,且有统计学意义(差异表达<0.5或差异表达>2),而PMSF前干预组与对照组比较,鸡脊髓组织Cofilin-1b的表达差异无统计学意义。在TOCP暴露第20天,TOCP组鸡脊髓组织Cofilin-1b表达与其他两组比较尽管有下降趋势,但无显著性变化。结论 TOCP暴露能导致鸡脊髓神经组织Cofilin-1b表达在早期显著下调,且该蛋白表达下调可能与微丝骨架结构紊乱及其OPIDN诱发机制有关。
OBJECTIVE: To screen differentially expressed proteins that may be involved in depolymerization of regulatory micro-filaments in the spinal cord tissue of chickens exposed with tri-ortho-tolyl phosphate (TOCP), and to provide targets for exploring the mechanism of OPIDN induced by organophosphorus compounds Protein basis. Methods Twenty-four roman hen chickens were randomly divided into 1000 mg / kg TOCP group, pretreatment with PMSF 40 mg / kg and then TOCP 1000 mg / kg intervention group and saline control group, each group 14 only. On the 5th and 20th day of exposure, four chickens were sacrificed in each group. The spinal cord was separated under low temperature and total protein was extracted. Two-dimensional electrophoresis combined with mass spectrometry was used to screen and identify differentially expressed proteins that may be involved in the regulation of microfilament depolymerization. Results The typical symptoms of OPIDN such as progressive ataxia and myasthenia were found in the TOCP group before and after the 7th day of exposure. The onset began from the distal part of the lower extremity and the severity of the disease gradually aggravated with time until full paralysis, while in the other groups, No OPIDN symptoms were observed during the observation period. On the fifth day after exposure, the TOCP group rats were respectively 3.4 and 2.8 fold lower than those in the control group and the pre-PMSF intervention group (P <0.05). The differences were statistically significant (difference <0.5 Or difference expression> 2). However, there was no significant difference in the expression of Cofilin-1b between the pre-PMSF intervention group and the control group. At day 20 of TOCP exposure, Cofilin-1b expression in chicken spinal cord tissue in TOCP group showed no significant change compared with the other two groups despite the downward trend. Conclusion Exposure of TOCP can lead to significant down-regulation of Cofilin-1b expression in chicken spinal cord nerve tissue at early stage. The down-regulation of the expression of Cofilin-1b may be related to the disorder of actin cytoskeleton and the mechanism of OPIDN induction.