Methylation of selected CpG islands involved in the transcription of myeloperoxidase and superoxide dismutase 2 in neutrophils of periparturient and mid-lactation cows.

It is generally accepted that intracellular killing of microorganisms by production of reactive oxygen species (ROS) in the phagosome of the neutrophil is an important arm of innate defense. High-producing dairy cows are prone to periparturient metabolic and infectious diseases. Both myeloperoxidase (MPO) activity and ROS production decrease the day of parturition. Several studies have demonstrated changes in the expression of genes involved in, for example, metabolism and defense in the circulating neutrophil during peripartum. In this study, we wanted to further characterize the periparturient neutrophil in terms of its oxidative killing capacity by analyzing the oxidative burst at 3 levels. First, the ROS phenotype was evaluated using chemiluminescence. The cows (sampled within 24 h after parturition and at 135 d in milk) showed a significantly slower production of ROS at parturition. Both primiparous (n = 13) and multiparous (n = 12) cows were included in this study, but parity did not affect the kinetics of ROS production. Second, the expression of 11 genes involved in ROS production was measured in the same cows: cytochrome b-245 α and β chain (CYBA, CYBB; coding for membrane-bound constituents of NADPH oxidase); neutrophil cytosolic factors 1, 2, and 4 (NCF1, NCF2, and NCF4); Rac family small GTPase 1 and 2 (RAC1 and RAC2; coding for regulatory proteins of NADPH oxidase); superoxide dismutase 2 (SOD2); catalase (CAT); myeloperoxidase (MPO; coding for enzymes involved in metabolizing downstream ROS); and spleen-associated tyrosine kinase (SYK; involved in signaling). During peripartum, a shift in expression in the oxidative killing pathway was observed, characterized by a downregulation of MPO and a simultaneous upregulation of the genes coding for NADPH oxidase. Third, as total DNA methylation is known to change during pregnancy, we investigated whether the observed differences were due to different methylation patterns. Promotor regions initiate transcription of particular genes; therefore, we analyzed the methylation status in annotated CpG islands of MPO and SOD2, 2 genes with a significant difference in expression between both lactation stages. The differences in methylation of these CpG islands were nonsignificant. High-throughput techniques may be necessary to obtain more detailed information on the total DNA methylation dynamics in bovine neutrophils and increase our understanding of how gene expression is controlled in neutrophils.