锰致神经毒性效应与其潜在效应标志物-Parkin的关系研究

Manganese was widely used in steelmaking, electric welding, gasoline anti-knock and stainless steel. Over exposure to manganese lead neurotoxicity, called manganism, with the symptoms similar to Parkinson disease, and it is irreversible if the motor dysfunction of the patient occurred. The mechanism of manganism is not clear, and there is no available early effect-biomarker for the neurotoxicity. Research results showed that the oxidative stress, mitochondrial dysfunction induced by Mn and other mechanism play main roles in the lesions of dopaminergic and gabaergic system. Parkin, the product of PARK2, alleviated the lesion of dopaminergic system caused by oxidative stress and mitochondrial dysfunction. Our study results demonstrated that the PARK2 expression decreased both in the leucocyte and the buccal cells of the Mn-exposure workers, the same tendency with decreased PARK2 expression in the striatum of the Mn-exposed rats. Nevertheless，it is necessary to have assays to prove Parkin mitigate the Mn-induced neurons lesion, and to have experiments to support the hypothesis that the Decreased PARK2 expression can work as the effect-biomarker of Mn-induced neurotoxicity. This program designed studies on the relationship of Parkin and the Mn-induced neurotoxicity by occupational exposure investigation, animals experiment and primary neuron culture experiments. It is promising that these studies will explain some of the mechanisms of manganism, and demonstrate that decreased PARK2 expression in the Mn exposure persons could work as the effect-biomarker of neurotoxicity. It is practical for protection the Mn-exposed people from neurotoxicity.

锰在炼钢、电焊、汽油抗爆剂及不锈钢制品中均得到广泛使用。接触锰过多主要引起中枢神经损害，出现类似帕金森样症状，这种损害在病人出现运动系统症状以后则不可逆。锰引起神经损害的机制尚不完全清楚，也没有合适效应生物标志物。前人研究证实锰可通过氧化应激、线粒体损伤等多种机制对多巴胺能神经和GABA能神经引起损害。而PARK2基因的表达产物Parkin对多巴胺能神经具有保护作用。我们的前期研究发现，人群锰暴露引起血液和唾液中PARK2表达降低，与染锰大鼠纹状体中PARK2表达降低一致。但是，Parkin是否在锰致神经损害中发挥保护作用及PARK2的表达降低能否作为锰致神经毒性的效应标志物都需要验证。本项目拟从整体动物、细胞水平和锰（接触）中毒人群三个层面研究锰致神经损害与Parkin之间的关系，探索Parkin在此过程中的作用。试图找到较为合适的锰神经毒性效应标志物。该研究对预防锰神经危害有实际意义。

该研究项目通过从人群流行病学、细胞培养和整体动物试验三个层面研究锰致神经损害与PARK2表达之间的关系。在人群流行病学研究方面，用石墨炉原子吸收测定红细胞、血浆及唾液的锰和铁含量，用实时荧光PCR测定PARK2, DMT1, Tf, Hepcidin, TfR的表达。在细胞培养研究方面，对细胞株进行染锰之后测定PARK2的表达及多巴胺（DA）水平的变化，对大鼠纹状体细胞培养进行干扰PARK2的表达，测定干扰前后PARK2的表达及DA水平的变化。在整体动物水平，大鼠染锰之后测定PARK2的表达。人群流行病学研究发现PARK2表达在暴露于锰的工人的血液中显着降低，并且与血浆锰，红细胞锰，唾液锰和累积锰暴露呈负相关。 这一发现表明，PARK2可以作为锰暴露的生物标志。职业接触锰人群，红细胞和血浆中DMT1, Tf, Hepcidin 三个基因的表达水平均与红细胞和血浆中的锰浓度呈负相关。而TfR的表达水平与红细胞和血浆中的锰浓度无相关关系。我们用具有神细胞功能的细胞珠及大鼠原代培养神经细胞均证实锰对神细胞的作用过程与PARK2表达相关，说明PARK2在锰致神经系统危害的过程中参与了相关过程，PARK2的表达水平降低，可以作为锰接触的生物标志，同时染锰之后DA水平下降。用整体动物实验研究证实，锰引起神经系统危害，及所导致的神经行为学改变与PARK2的表达和功能相关，该作用过程极有可能是锰所引起神经系统危害的作用机制之一。从人群流行病学、细胞培养和整体动物试验三个层面研究锰致神经损害与PARK2表达之间的关系，探索出PARK2降低可作为锰所引起神经系统危害的作用机制之一和锰神经毒性生物标志，这对今后预防锰神经危害有实际意义。