Arnal Nathalie, de Alaniz María J Tacconi, Marra Carlos Alberto
INIBIOLP, CCT La Plata, CONICET-UNLP, Cátedra de Bioquímica y Biología Molecular, Facultad de Ciencias Médicas, Universidad Nacional de La Plata, La Plata, Argentina.
Biochim Biophys Acta. 2012 Jul;1820(7):931-9. doi: 10.1016/j.bbagen.2012.03.007. Epub 2012 Mar 17.
Copper (Cu) is an essential trace metal used as a catalytic cofactor for many enzymes. However, it can have nocive effects when it participates in the Fenton reaction, producing reactive oxygen species (ROS). Excess Cu is present in the plasma of patients with diseases in which cell survival is crucial. In order to investigate the effect of Cu overload on the induction of cellular damage we chose two human cell lines derived from liver (HepG2) and lung (A-549) as representative cells exposed to exogenous (polluted air) and/or endogenous (systemic) Cu overload.
We studied ROS production using thiobarbituric acid reactive substances (TBARS) and fluorimetric measurements with dichlorofluorescein, cell viability by the trypan dye exclusion test, the methyltetrazolium (MTT) and lactate dehydrogenase leakage (LDH) assays, various cytotoxic indexes, and caspasa-3 and calpain-dependent activation as the main signals involved in the apoptosis pathway.
Cu overload induces cell death by a differential activation of calpains (m- and μ-) and caspase-3, and modifies various proliferative indexes in a cell-type and concentration-dependent manner. The involvement of these two protease systems and the response of the two main Cu homoestatic proteins ceruloplasmin and metallothioneins are specific to each cell type. We demonstrated that Cu can trigger cell death by activation of specific protease systems and modify various proliferative indexes in a cell-type and concentration-dependent manner.
These findings contribute to understanding the diverse effects of Cu overload on the pathogenesis of human diseases like cancer, cirrhosis and degenerative disorders.
铜（Cu）是一种必需的微量元素，用作许多酶的催化辅因子。然而，当它参与芬顿反应产生活性氧（ROS）时，可能会产生有害影响。在细胞存活至关重要的疾病患者血浆中存在过量的铜。为了研究铜过载对细胞损伤诱导的影响，我们选择了两种源自肝脏（HepG2）和肺（A - 549）的人类细胞系，作为暴露于外源性（污染空气）和/或内源性（全身性）铜过载的代表性细胞。
我们使用硫代巴比妥酸反应性物质（TBARS）和二氯荧光素荧光测量法研究ROS的产生，通过台盼蓝染料排斥试验、甲基噻唑基四唑（MTT）和乳酸脱氢酶泄漏（LDH）测定法研究细胞活力，各种细胞毒性指标，以及作为凋亡途径中主要信号的caspasa - 3和钙蛋白酶依赖性激活。
铜过载通过钙蛋白酶（m - 和μ - ）和caspase - 3的差异激活诱导细胞死亡，并以细胞类型和浓度依赖性方式改变各种增殖指标。这两种蛋白酶系统的参与以及两种主要的铜稳态蛋白铜蓝蛋白和金属硫蛋白的反应对每种细胞类型都是特异性的。我们证明铜可以通过激活特定的蛋白酶系统触发细胞死亡，并以细胞类型和浓度依赖性方式改变各种增殖指标。
这些发现有助于理解铜过载对人类疾病如癌症、肝硬化和退行性疾病发病机制的多种影响。
