RESUMO
The outcome of sepsis occurs due to influence of environmental and genetic factors besides genes variants whose expression support its outcome or not. Oxidative stress is associated to the pathogenicity of sepsis, occurring when there is a reactive species overproduction associated with inflammation. The aim of this study was to characterize the cellular redox status of human peripheral blood mononuclear cells (PBMCs) with either -9Ala (AA) or -9Val (VV) SOD2 genotypes and evaluate their response to oxidative stress induced by lipopolysaccharide (LPS). The PBMCs were isolated from the blood of 30 healthy human volunteers (15 volunteers for each allele) and the following assays were performed: antioxidant enzyme activities (superoxide dismutase; catalase; glutathione peroxidase), total radical-trapping antioxidant parameter, non-enzymatic antioxidant capacity (total antioxidant reactivity), and quantification of conjugated dienes (lipid peroxidation). At basal conditions (i.e., not stimulated by LPS), cells from 47C allele carriers showed higher activities of CAT and SOD, as well as higher TAR compared to 47T allele. However, when 47CC cells were challenged with LPS, we observed a higher shift toward a pro-oxidant state compared to 47TT cells. The CAT activity and lipid peroxidation were increased in cells with both alleles, but SOD activity increased significantly only in 47TT cells. These results demonstrate that SOD2 polymorphisms are associated with different cellular redox environments at both basal and LPS-stimulated states, and identification of this polymorphism may be important for a better understanding of pro-inflammatory conditions.
Assuntos
Leucócitos Mononucleares/enzimologia , Lipopolissacarídeos/farmacologia , Polimorfismo de Nucleotídeo Único , Superóxido Dismutase/genética , Adulto , Substituição de Aminoácidos , Catalase , Células Cultivadas , Feminino , Radicais Livres/metabolismo , Glutationa Peroxidase/metabolismo , Heterozigoto , Humanos , Líquido Intracelular/enzimologia , Leucócitos Mononucleares/imunologia , Peroxidação de Lipídeos , Masculino , Nitritos/metabolismo , Estresse Oxidativo , Fator de Necrose Tumoral alfa/metabolismo , Adulto JovemRESUMO
A task-force to resolve 26 pending forensic caseworks was carried out. We tested four different protocols to extract DNA from molar and pre-molar teeth from 26 cadavers with post-mortem intervals from 2 months to 12 years. We compared the amount of DNA and DNA profiles with the time elapsed between death and laboratory procedures. Molar or pre-molar teeth were removed from the corpses, cleaned, and DNA was extracted using 2 or 12h of incubation on lysis buffer and filtered using concentration column or precipitated with isopropanol. DNA profiles were obtained using PowerPlex16™ System PCR Amplification Kit, AmpFlSTR(®) Yfiler™ and/or mtDNA sequencing. Complete DNA profiles comparison and statistical evaluation allowed unambiguous identification of the 26 victims. No significant differences were observed in the amount of DNA obtained with the distinct incubation times. The use of concentration column resulted in an increased amount of DNA when compared to isopropanol. However, the lower concentration of DNA obtained with isopropanol seemed to have been compensated by the higher purity. No significant differences in the number of amplified loci were found. A non-significant tendency was found between the amount of total DNA recovered and the time elapsed between death and laboratory procedures. The increase of post-mortem time did not interfere in the analysed autosomal loci. In conclusion, molar and pre-molar teeth were shown to be good candidates to obtain satisfactory DNA profiles, suggesting the high potential of tooth samples as source for DNA typing independently of the decomposed corpse's time or laboratory procedures.