Matrix metalloproteinases and their inhibitors as biomarkers for metal toxicity in vitro.

Exposure to nickel and chromium, and their compounds, has been associated with adverse health effects. These metals are two human carcinogens whose pathogenesis involves active extracellular matrix degradation and remodelling. In this work we have compared the effects of in vitro exposure to nickel and chromium of a keratinocyte cell line (HaCat). The modulation of matrix metalloproteinase genes was used as biomarker of chemical damage. Confluent cells were constantly exposed to subtoxic chromium and nickel concentrations (10(-5) and 10(-7)M) up to 72 h. Total RNA was extracted and specific matrix metalloproteinase, and inhibitor, gene expression was analyzed by RT-PCR. Moreover, cell cycle alterations were evaluated by flow cytometry. Nickel and chromium showed different results, with an upregulation of MMP-2 mRNA production in nickel-treated cells while chromium exposure down-regulated MMP-2 mRNA production. This result could be correlated to the precocious (6h) over-expression of tissue inhibitor-1 (TIMP-1) mRNA in chromium-treated cells. Cell cycle analysis showed and increase of cells with 4N DNA. These results could be explained as a survival response of cells that escape metal induced apoptosis through the anti-apoptotic effects of TIMP-1. These cells that encompass the genotoxic insult may have a selective proliferation advantage, and therefore represent the precursor pool from which degenerating variants may emerge. To study if the chemical damage was reversible, subconfluent cells were stimulated only for 24 h, then the medium was replaced without metal. Cells were able to recover from nickel exposure, showing only weak alterations in specific mRNA expression and cell cycle alteration respect to control. Chromium-induced damage was irreversible. Our results demonstrated that there is an association between metal toxicity and expression of MMPs and their inhibitors. These biomarkers could be potentially useful to elaborate a prediction model of chemical toxicity.