Oxidant resistance of cadmium-adapted human lung fibroblasts

Metallothionein (MT) is a metal and thiol-rich protein readily induced by cadmium (Cd) exposure. In vitro experiments have demonstrated that MT is able to serve as a scavenger of hydroxyl radicals as well as superoxide anions, albeit to a lesser extent. The role of MT as a mediator in Cd induced oxidant resistance was investigated in a nontransformed human lung fibroblast cell line (IMR-90). Cells were passaged three times either in a Cd-containing medium (8.9 microM CdCl2) or in a medium which lacked Cd. Cellular MT content, as quantitated by a modification of the heme/109Cd binding assay, increased significantly with each passage in Cd. Immunocytochemistry studies revealed that all Cd-pretreated cells contained MT and that MT was localized in both cytoplasmic and nuclear compartments. Immunolabeling was more intense in some cells compared to others. Very slight immunolabeling was observed in physiological control cells, grown in the absence of Cd, and virtually no staining was observed in Cd-adapted or non-adapted cells when primary antibody was omitted. Using the xanthine/xanthine oxidase system as a generating system for active oxygen species, we found that the magnitude of cell injury for Cd-adapted and non-adapted fibroblasts was dependent upon oxidant concentration and duration of oxidant exposure. Cd-adapted fibroblasts, which were characterized by over-expression of MT, were significantly more resistant to injury by active oxygen species and also exhibited a greater ability to scavenge extracellular hydrogen peroxide compared to cells with no previous history of Cd exposure. Experiments with aminotriazole demonstrated that catalase was not a major contributor to the additional hydrogen peroxide scavenging capacity of Cd-adapted cells. The data presented in this report are consistent with involvement of MT in protecting critical cellular targets from reactive oxygen species.