Chong Weelic, Jiménez Jessica, McIIvin Matthew, Saito Mak A, Kwakye Gunnar F
Neuroscience Department, Oberlin College, 119 Woodland Street, Oberlin, OH, 44074, USA.
Marine Chemistry and Geochemistry Department, Woods Hole Oceanographic Institution, Woods Hole, MA, USA.
Neurotox Res. 2017 Aug;32(2):231-246. doi: 10.1007/s12640-017-9725-x. Epub 2017 Mar 28.
This study examined the role of alpha-synuclein in regulating cadmium (Cd)-induced neurotoxicity using the N27 dopaminergic neuronal model of Parkinson's disease (PD) that stably expresses wild-type human α-synuclein (α-Syn) or empty vector (Vec) control. We report that α-Syn significantly increased Cd-induced cytotoxicity as compared to Vec control cells upon 24 h exposure. To explore the cellular mechanisms, we examined oxidative stress, caspase activation, and Cd uptake and intracellular accumulation. Expression of α-Syn coupled with Cd-induced cytotoxicity increased oxidative stress. Inductively coupled plasma-mass spectrometry (ICP-MS) revealed an increase in Cd uptake and intracellular accumulation in α-Syn-expressing cells upon Cd exposure. Analysis of the mitochondrial mediated apoptotic pathway showed greater activation of caspase-9 and caspase-3 in α-Syn cells. To functionally evaluate the role of metal transporters in the altered Cd phenotype, we examined Cd toxicity in the presence of nontoxic levels of divalent manganese Mn(II) and iron Fe(II). Co-treatment with Fe(II) or Mn(II) did not significantly attenuate Cd-induced cytotoxicity. We report that Cd exposure decreased the divalent metal transporter 1 and Akt protein levels in the α-Syn-expressing cells without altering native PKCδ protein levels in both Vec control and α-Syn lines. In addition, we show decreased basal metallothionein-3 protein expression in α-Syn-expressing cells. Co-treatment with N-acetyl-L-cysteine was sufficient to attenuate and abolish the α-Syn × Cd-induced cytotoxicity. Collectively, these results demonstrate that α-Syn exhibits neurotoxic properties upon acute Cd exposure to cause cell death by causing oxidative stress, increasing Cd uptake, altering caspase-9 and caspase-3 activation, and diminishing the neuroprotective effect of Akt in a dopaminergic neuronal model of PD.
本研究使用稳定表达野生型人α-突触核蛋白(α-Syn)或空载体(Vec)对照的帕金森病(PD)N27多巴胺能神经元模型,研究了α-突触核蛋白在调节镉(Cd)诱导的神经毒性中的作用。我们报告,与Vec对照细胞相比,暴露24小时后,α-Syn显著增加了Cd诱导的细胞毒性。为了探究细胞机制,我们检测了氧化应激、半胱天冬酶激活以及Cd摄取和细胞内积累情况。α-Syn的表达与Cd诱导的细胞毒性增加了氧化应激。电感耦合等离子体质谱(ICP-MS)显示,Cd暴露后,表达α-Syn的细胞中Cd摄取和细胞内积累增加。线粒体介导的凋亡途径分析显示,α-Syn细胞中半胱天冬酶-9和半胱天冬酶-3的激活程度更高。为了从功能上评估金属转运蛋白在改变的Cd表型中的作用,我们检测了在无毒水平的二价锰Mn(II)和铁Fe(II)存在下的Cd毒性。与Fe(II)或Mn(II)共同处理并没有显著减弱Cd诱导的细胞毒性。我们报告,Cd暴露降低了表达α-Syn的细胞中二价金属转运蛋白1和Akt蛋白水平,而在Vec对照和α-Syn细胞系中均未改变天然PKCδ蛋白水平。此外,我们发现表达α-Syn的细胞中基础金属硫蛋白-3蛋白表达降低。与N-乙酰-L-半胱氨酸共同处理足以减弱并消除α-Syn×Cd诱导的细胞毒性。总的来说,这些结果表明,在急性Cd暴露时,α-Syn表现出神经毒性特性,通过引起氧化应激、增加Cd摄取、改变半胱天冬酶-9和半胱天冬酶-3的激活以及削弱Akt在PD多巴胺能神经元模型中的神经保护作用来导致细胞死亡。