Differential phenotype of immune cells in blood and milk following pegylated granulocyte colony-stimulating factor therapy during a chronic Staphylococcus aureus infection in lactating Holsteins

摘要

Neutrophils are principal host innate immune cellresponders to mastitis infections. Thus, therapies havebeen developed that target neutrophil expansion. Thisincludes the neutrophil-stimulating cytokine granulocytecolony-stimulating factor (gCSF). PegylatedgCSF (PEG-gCSF; Imrestor, Elanco Animal Health,Greenfield, IN) has been shown to reduce the naturalincidence of mastitis in periparturient cows in commercialsettings and reduce severity of disease againstexperimental mastitis challenge. Pegylated gCSF stimulatesneutrophil expansion but also induces changes inmonocyte and lymphocyte circulating numbers, surfaceprotein expression changes, or both. We hypothesizedthat PEG-gCSF modulates surface expression of monocytesand neutrophils and facilitates their migration tothe mammary gland. We challenged 8 mid-lactationHolsteins with approximately 150 cfu of Staphylococcusaureus (Newbould 305) in a single quarter via intramammaryinfusion. All animals developed chronicinfections as assessed by bacteria counts and somaticcell counts (SCC). Ten to 16 wk postchallenge, 4 of theanimals were treated with 2 subcutaneous injections ofPEG-gCSF 7 d apart. Complete blood counts, SCC,bacterial counts, milk yield, feed intake, neutrophils extracellulartrap analysis, and flow cytometric analysesof milk and blood samples were performed at indicatedtime points for 14 d after the first PEG-gCSF injection.The PEG-gCSF-treated cows had significantly increasednumbers of blood neutrophils and lymphocytescompared with control cows. Flow cytometric analysesrevealed increased surface expression of myeloperoxidase(MPO) on neutrophils and macrophages in milkbut not in blood of treated cows. Neutrophils isolatedfrom blood of PEG-gCSF-treated cows had decreasedsurface expression of CD62L (L-selectin) in blood, consistentwith cell activation. Surprisingly, CD62L cellsurface expression was increased on neutrophils andmacrophages sourced from milk from treated animalscompared with cells isolated from controls. The PEGgCSF-treated cows did not clear the S. aureus infection,nor did they significantly differ in SCC from controls.These findings provide evidence that PEG-gCSFtherapy modifies cell surface expression of neutrophilsand monocytes. However, although surface MPO+cells accumulate in the mammary gland, the lack ofbacterial control from these milk-derived cells suggestsan incomplete role for PEG-gCSF treatment againstchronic S. aureus infection and possibly chronic mammaryinfections in general.