title: Methotrexate-related response on human peripheral blood mononuclear cells may be modulated by the Ala16Val-SOD2 gene polymorphism.
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.
authors.affiliation: Pharmacology Graduate Program, Federal University of Santa Maria, Santa Maria, RS, Brazil.
authors.forename: Fernanda
authors.initials: F
authors.lastname: Barbisan
authors.affiliation: Biogenomic Laboratory, Federal University of Santa Maria, Santa Maria, RS, Brazil.
authors.forename: Jéssica de Rosso
authors.initials: Jde R
authors.lastname: Motta
authors.affiliation: Laboratory of Molecular Biology, University of Western Santa Catarina, UNOESC, Chapecó, SC, Brazil.
authors.forename: Alexis
authors.initials: A
authors.lastname: Trott
authors.affiliation: Pharmacology Graduate Program, Federal University of Santa Maria, Santa Maria, RS, Brazil.
authors.forename: Verônica
authors.initials: V
authors.lastname: Azzolin
authors.affiliation: Biochemical Toxicology Graduate Program, Federal University of Santa Maria, Santa Maria, RS, Brazil.
authors.forename: Eduardo Bortoluzzi
authors.initials: EB
authors.lastname: Dornelles
authors.affiliation: Biogenomic Laboratory, Federal University of Santa Maria, Santa Maria, RS, Brazil.
authors.forename: Matheus
authors.initials: M
authors.lastname: Marcon
authors.affiliation: Biochemical Toxicology Graduate Program, Federal University of Santa Maria, Santa Maria, RS, Brazil.
authors.forename: Thaís Doeler
authors.initials: TD
authors.lastname: Algarve
authors.affiliation: Biogenomic Laboratory, Federal University of Santa Maria, Santa Maria, RS, Brazil.
authors.forename: Marta Maria Medeiros Frescura
authors.initials: MM
authors.lastname: Duarte
authors.affiliation: Biogenomic Laboratory, Federal University of Santa Maria, Santa Maria, RS, Brazil.
authors.forename: Clarice Pinheiro
authors.initials: CP
authors.lastname: Mostardeiro
authors.affiliation: Biogenomic Laboratory, Federal University of Santa Maria, Santa Maria, RS, Brazil.
authors.forename: Taís Cristina
authors.initials: TC
authors.lastname: Unfer
authors.affiliation: Biochemical Toxicology Graduate Program, Federal University of Santa Maria, Santa Maria, RS, Brazil.
authors.forename: Karen Lilian
authors.initials: KL
authors.lastname: Schott
authors.affiliation: Biogenomic Laboratory, Federal University of Santa Maria, Santa Maria, RS, Brazil; Pharmacology Graduate Program, Federal University of Santa Maria, Santa Maria, RS, Brazil; Biochemical Toxicology Graduate Program, Federal University of Santa Maria, Santa Maria, RS, Brazil.
authors.forename: Ivana Beatrice Mânica
authors.initials: IB
authors.lastname: da Cruz
mesh_terms: D000975:Antioxidants / Q000378:metabolism; D020169:Caspases / Q000235:genetics / Q000378:metabolism; D016207:Cytokines / Q000096:biosynthesis; D004305:Dose-Response Relationship, Drug; D005452:Fluoresceins / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000235:genetics; D006801:Humans; D015227:Lipid Peroxidation / Q000187:drug effects / Q000235:genetics; D008727:Methotrexate / Q000494:pharmacology*; D020641:Polymorphism, Single Nucleotide; D050050:Protein Carbonylation / Q000187:drug effects / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D013482:Superoxide Dismutase / Q000235:genetics*; D000099156:Superoxide Dismutase 2
chemical_list: D000975:Antioxidants; D020169:Caspases; D016207:Cytokines; D005452:Fluoresceins; D008727:Methotrexate; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D013482:Superoxide Dismutase; D000099156:Superoxide Dismutase 2; C029569:diacetyldichlorofluorescein