Abstract
Methotrexate (MTX) is a folic acid antagonist used in high doses as an anti-cancer treatment and in low doses for the treatment of some autoimmune diseases. MTX use has been linked to oxidative imbalance, which may cause multi-organ toxicities that can be attenuated by antioxidant supplementation. Despite the oxidative effect of MTX, the influence of antioxidant gene polymorphisms on MTX toxicity is not well studied. Therefore, we analyzed here whether a genetic imbalance of the manganese-dependent superoxide dismutase (SOD2) gene could have some impact on the MTX cytotoxic response. An in vitro study using human peripheral blood mononuclear cells (PBMCs) obtained from carriers with different Ala16Val-SOD2 genotypes (AA, VV and AV) was carried out, and the effect on cell viability and proliferation was analyzed, as well as the effect on oxidative, inflammatory and apoptotic markers. AA-PBMCs that present higher SOD2 efficiencies were more resistance to high MTX doses (10 and 100 µM) than were the VV and AV genotypes. Both lipoperoxidation and ROS levels increased significantly in PBMCs exposed to MTX independent of Ala16Val-SOD2 genotypes, whereas increased protein carbonylation was observed only in PBMCs from V allele carriers. The AA-PBMCs exposed to MTX showed decreasing SOD2 activity, but a concomitant up regulation of the SOD2 gene was observed. A significant increase in glutathione peroxidase (GPX) levels was observed in all PBMCs exposed to MTX. However, this effect was more intense in AA-PBMCs. Caspase-8 and -3 levels were increased in cells exposed to MTX, but the modulation of these genes, as well as that of the Bax and Bcl-2 genes involved in the apoptosis pathway, presented a modulation that was dependent on the SOD2 genotype. MTX at a concentration of 10 µM also increased inflammatory cytokines (IL-1β, IL-6, TNFα and Igγ) and decreased the level of IL-10 anti-inflammatory cytokine, independent of SOD2 genetic background. The results suggest that potential pharmacogenetic effect on the cytotoxic response to MTX due differential redox status of cells carriers different SOD2 genotypes.
Highlights
Methotrexate (MTX) is a drug that has been used since the 1950 s to treat a broad number of morbidities such as cancer and autoimmune diseases
peripheral blood mononuclear cells (PBMCs) from carriers with the V allele (VV and AV) exhibited decreased viability when exposed to MTX at 1, 10 and 100 mM concentrations, whereas AA-PBMC viability was not affected by these treatments
The present study, confirmed that the cytotoxic effect of MTX, a commonly used anti-inflammatory, antiproliferative, and immunosuppressive drug, on human PBMCs involves an acute imbalance of cell oxidative and inflammatory metabolism and triggers apoptosis [14,10,11,25]
Summary
Methotrexate (MTX) is a drug that has been used since the 1950 s to treat a broad number of morbidities such as cancer and autoimmune diseases. MTX has anti-inflammatory and/or immunosuppressive effects [2] related to the induction of lymphocyte apoptosis through oxidative stress and increasing caspase-3 levels [3,4]. For this reason, it is the firstline therapy for the treatment of moderate to severe psoriasis and psoriatic arthritis all over the world [5]. Investigations suggest that oxidative stress caused by MTX involves decreasing in some antioxidant enzymes as glutathione peroxidase, glutathione reductase, catalase and superoxide dismutase, increasing of lipoperoxidation and reactive oxygen species (ROS) levels, as well as apoptosis induction [4,10]
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