红细胞共价交联血影蛋白作为正己烷神经毒效应替代标志物的初步人群研究

Preliminary population study on erythrocyte covalent crosslinking spectrin as a surrogate biomarker of n-hexane induced neurotoxicity

目的初步在人群中论证外周血红细胞共价交联血影蛋白(Sp)作为正己烷神经毒效应替代标志物的可行性。方法选取15名职业接触正己烷的工人作为接触组,将其中神经-肌电图异常的5人分类为慢性正己烷中毒前期接触者,其余无神经-肌电图异常的10人分类为一般接触者;选取无正己烷接触史的10名身体健康者作为对照组。采集所有研究对象肘静脉血并提取红细胞膜蛋白。聚丙烯凝胶电泳分离不同相对分子质量的膜蛋白后,分别用考马斯亮蓝和硝酸银染色分析蛋白交联情况,并用免疫印迹实验检测交联蛋白是否为Sp。比较不同组间共价交联Sp阳性率差异。结果所有膜蛋白样本电泳后经考马斯亮蓝染色均未显示高分子量交联条带,但部分蛋白样本电泳后经硝酸银染色显示交联条带,经免疫印迹实验证实交联条带为Sp。中毒前期接触者共价交联Sp阳性率(100%)高于一般接触者(10.0%)(P<0.01)。结论外周血红细胞共价交联Sp有可能作为正己烷接触人群神经毒效应替代标志物。

Objective To study preliminarily the feasibility of erythrocyte covalent crosslinking spectrin as a surrogate biomarker of n-hexane induced neurotoxicity in human population.Methods A total of 15 workers occupationally exposed to n-hexane,including 5 workers with abnormal neuro-electromyography(cases at early stage of occupational chronic n-hexane poisoning,OCHP)and 10 workers without any abnormal neuro-electromyography(cases as the general exposed),and 10 healthy workers without any exposure to n-hexane and other known hazards were studied.Their peripheral venous blood samples were taken,membrane proteins of erythrocytes were extracted and then separated with polypropylene gel electrophoresis by the molecular weight.The protein crosslinking was identified by respective dying with coomassie brilliant blue and silver nitrate,and finally the spectrin of crosslinking proteins was examined with western blot assay.Positive rate of crosslinking spectrin were compared among different groups.Results The dying with coomassie brilliant blue showed there was no high molecular weight crosslinking strip in all separated membrane protein samples,but dying with silver nitrate showed crosslinking strips in some protein samples and the crosslinking strips were testified to be spectrin by the western blot.The exposed workers with OCHP had significantly higher positive rate of crosslinking spectrin than exposed workers without abnormality(100.0%vs.10.0%,P<0.01),while no crosslinking spectrin was examined among control workers.Conclusion The erythrocyte crosslinking spectrin has the potential to be used as a surrogate biomarker of n-hexane induced neurotoxic effects among exposed population.