Li W, Zhao Y, Chou I N
Department of Microbiology, Boston University School of Medicine, Massachusetts 02118, USA.
Toxicol Appl Pharmacol. 1996 Jan;136(1):101-11. doi: 10.1006/taap.1996.0012.
As an essential metal for cell metabolism, Mg2+ is known to exert antagonism on Ni2+ genotoxic and other effects. This study examined the influence of Mg2+ on Ni(2+)-induced changes in microtubule (MT) assembly in vitro, cytoplasmic MT organization, cellular glutathione (GSH), and cytoskeletal and cytosolic protein sulfhydryls (PSH). As determined by a turbidity assay at 27 degrees C, Ni2+ enhanced the in vitro MT assembly in a Pipes buffer by shortening the initial lag (nucleation phase) and increasing the rate of polymerization with a higher final plateau. However, presence of 1 mM exogenous MgCl2 abolished the Ni2+ enhancing effect. Exposure of 3T3 cells to 2 mM NiCl2 for 20 hr resulted in perinuclear bundling of MTs and decreases in cytoskeletal and cytosolic PSH and cellular GSH levels. However, coincubation of cells with MgCl2 (1.25-20 mM) added to the culture medium markedly diminished the Ni2+ injury to MT organization. Under these conditions the Ni2+ interference with PSH was blocked such that both the cytoskeletal and cytosolic PSH levels returned to the range of control cells without metal treatment. Treatment of cells with Mg2+ (1.25-5 mM) for 20 hr slightly increased, while with higher Mg2+ doses (> 10 mM) decreased, cellular GSH content. Importantly, in Ni(2+)-treated cultures, addition of Mg2+ (1.25-10 mM) elevated GSH levels to > or = 200% of that in cells treated with Ni2+ alone. Furthermore, these Ni2+ and Mg2+ (1.25-10 mM) treated cells actually maintained GSH levels which were essentially unchanged from the basal level of control cells with no metal treatment. Although Mg2+ replacement of Ni2+ bound to MT proteins could be an important mechanism, cellular GSH may also be a critical factor in Mg2+ antagonism on Ni(2+)-enhanced MT assembly in view of the essential role of tubulin PSH in modulating MT assembly and, in turn, the GSH modulation of PSH.
作为细胞代谢的必需金属，已知Mg2+对Ni2+的遗传毒性及其他效应具有拮抗作用。本研究检测了Mg2+对体外镍离子（Ni2+）诱导的微管（MT）组装变化、细胞质MT组织、细胞内谷胱甘肽（GSH）以及细胞骨架和胞质蛋白巯基（PSH）的影响。通过在27℃下的比浊法测定，Ni2+通过缩短初始延迟期（成核阶段）并提高聚合速率及更高的最终平台期，增强了在Pipes缓冲液中的体外MT组装。然而，1 mM外源MgCl2的存在消除了Ni2+的增强作用。将3T3细胞暴露于2 mM NiCl2中20小时，导致MT在细胞核周围成束，并使细胞骨架和胞质PSH以及细胞内GSH水平降低。然而，将细胞与添加到培养基中的MgCl2（1.25 - 20 mM）共同孵育，显著减轻了Ni2+对MT组织的损伤。在这些条件下，Ni2+对PSH的干扰被阻断，使得细胞骨架和胞质PSH水平恢复到未进行金属处理的对照细胞的范围内。用Mg2+（1.25 - 5 mM）处理细胞20小时，细胞内GSH含量略有增加，而较高剂量的Mg2+（> 10 mM）则使其降低。重要的是，在Ni2+处理的培养物中，添加Mg2+（1.25 - 10 mM）可使GSH水平升高至单独用Ni2+处理的细胞中GSH水平的≥200%。此外，这些经Ni2+和Mg2+（1.25 - 10 mM）处理的细胞实际上维持了与未进行金属处理的对照细胞基础水平基本不变的GSH水平。尽管Mg2+替代与MT蛋白结合的Ni2+可能是一个重要机制，但鉴于微管蛋白PSH在调节MT组装中的重要作用，以及GSH对PSH的调节作用，细胞内GSH也可能是Mg2+拮抗Ni2+增强MT组装的关键因素。
