Centre for Cardiovascular and Metabolic Research, Hull York Medical School, University of Hull, Hull, HU6 7RX, UK.
Toxicol Sci. 2012 Jan;125(1):56-68. doi: 10.1093/toxsci/kfr268. Epub 2011 Oct 9.
Mercury is an established worldwide environmental pollutant with well-known toxicity affecting neurodevelopment in humans, but the molecular basis of cytotoxicity and the detoxification procedure are still unclear. Here we examined the involvement of the canonical transient receptor potential (TRPC) channel in the mercury-induced cytotoxicity and the potential detoxification strategy. Whole-cell and excised patches, Ca(2+) imaging, and site-directed mutagenesis were used to determine the mechanism of action of mercurial compounds on TRPC channels overexpressed in HEK293 cells, and cytotoxicity and preventive effect were investigated in cell culture models using small interfering RNA and pharmacological blockers. Mercury potently activates TRPC4 and TRPC5 channels. The extracellular cysteine residues (C(553) and C(558)) near the channel pore region of TRPC5 are the molecular targets for channel activation by mercury. The sensitivity of mercury to TRPC5 is presumed to be specific because other divalent heavy metal pollutants, such as Cd(2+), Ni(2+), and Zn(2+), had no stimulating effect, and TRPC3, TRPC6, TRPV1, and TRPM2 were resistant to mercurial compounds. The channel activity of TRPC5, as well as TRPC4, induced by mercury, was prevented by 2-aminoethoxydiphenyl borate and modified by a reducing environment. The inhibition of TRPC5 channels by specific TRPC5 pore-blocking antibody or by SKF-96365 alleviated the cytotoxicity, whereas the mercury chelator, meso-2,3-dimercaptosuccinic acid, showed nonselective prevention of cell survival. Silencing of the TRPC5 gene reduced the mercury-induced neuronal damage. These results indicate that mercurial compounds are activators for TRPC5 and TRPC4 channels. Blockade of TRPC channels could be a novel strategy for preventing mercury-induced cytotoxicity and neurodevelopment impairment.
汞是一种已被全球广泛认定的环境污染物，其毒性众所周知，会对人类的神经发育产生影响，但细胞毒性的分子基础和解毒过程仍不清楚。在这里，我们研究了经典的瞬时受体电位 (TRPC) 通道在汞诱导的细胞毒性中的作用，以及潜在的解毒策略。我们使用全细胞和膜片钳技术、钙离子成像和定点突变技术，确定了在过表达于 HEK293 细胞的 TRPC 通道中，汞化合物的作用机制，并在细胞培养模型中使用小干扰 RNA 和药理学抑制剂研究了细胞毒性和预防作用。汞能强烈激活 TRPC4 和 TRPC5 通道。TRPC5 通道孔区附近的细胞外半胱氨酸残基 (C(553) 和 C(558)) 是汞激活通道的分子靶点。汞对 TRPC5 的敏感性被认为是特异性的，因为其他二价重金属污染物，如 Cd(2+)、Ni(2+) 和 Zn(2+)，没有刺激作用，而 TRPC3、TRPC6、TRPV1 和 TRPM2 则对汞化合物有抗性。由汞诱导的 TRPC5 以及 TRPC4 通道活性可被 2-氨基乙氧基二苯硼酸盐阻止，并可被还原环境改变。特异性 TRPC5 孔阻断抗体或 SKF-96365 可抑制 TRPC5 通道，减轻细胞毒性，而汞螯合剂，meso-2,3-二巯基丁二酸则表现出非选择性的细胞存活预防作用。沉默 TRPC5 基因可减少汞诱导的神经元损伤。这些结果表明，汞化合物是 TRPC5 和 TRPC4 通道的激活剂。阻断 TRPC 通道可能是预防汞诱导的细胞毒性和神经发育损伤的一种新策略。
