Effects of dexamethasone and Mycoplasma bovis on bovine neutrophil function in vitro

It is well established that exposure either to elevated levels of glucocorticoids, or to Mycoplasma bovis (M. bovis), has a negative effect on bovine neutrophil function. The objective of this research was to determine whether in vitro treatment of bovine neutrophils by M. bovis strains (n=4) and glucocorticoids would additively impair phagocyte function. Twenty, healthy, dairy cows were enrolled. Whole blood was collected from all cows for neutrophil isolation. Phagocytosis and the generation of superoxide anion (O2(-)) were tested in vitro by incubation of neutrophils with FITC labeled Escherichia coli (E. coli) and cytochrome c after treatment. Treatments included: NM1-4D (neutrophils treated with dexamethasone and exposed to one of the four M. bovis strains); NM1-4 (neutrophils exposed to one of the four M. bovis strains only); ND (neutrophils treated with dexamethasone only); and N (non-treated control neutrophils). The overall percentages of neutrophils phagocytizing E. coli were: 32%, 51%, 37%, and 53% ± 5.25% for treatments NM1-4D, NM1-4, ND, and N, respectively. The overall statistically transformed means of phagocytized E. coli per neutrophil were: 1.37, 1.72, 1.33, and 1.67 ± 0.057 for treatments NM1-4D, NM1-4, ND, and N, respectively. The overall statistically transformed means of neutrophil O2(-) production were: 8.60, 11.91, 9.01, and 12.21 ± 0.21 nmol/10(6) for treatments NM1-4D, NM1-4, ND, and N, respectively. Exposure of neutrophils to M. bovis plus dexamethasone had an additive effect on generation of reactive oxygen species (p=0.0057), but not on the percentage of neutrophils phagocytizing E. coli (p=0.0817) or number of E. coli phagocytized per neutrophil (p=0.2946). Only one of the four M. bovis strains had a negative effect on neutrophil phagocytic function. Dexamethasone treatment consistently decreased neutrophil function as indicated by decreased percentage of neutrophils phagocytizing E. coli, decreased number of E. coli phagocytized per neutrophil, and decreased neutrophil O2(-) production, compared to controls (p<0.0001). Results suggested a synergistic effect of in vitro incubation of glucocorticoids and M. bovis on reduction of bovine neutrophil function as measured by generation of reactive oxygen species. These findings may explain in part the interaction between stressful events and outbreak of Mycoplasma bovis associated bovine disease.