Submitted to: American Journal of Veterinary Research
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/10/2000
Publication Date: 8/17/2000
Interpretive Summary: Neutrophils, a type of white blood cells that ingests bacteria, are the major white blood cell in milk from cows with mastitis. However, their presence in milk is like a double edged sword. While they ingest and destroy mastitis causing pathogens, they also release potent chemicals called radicals that destroy mammary secretory tissue. This results in decreased milk production. Phosphorylation of tyrosine residues on protein inside cells is an important prerequisite for normal cell function. During the inflammatory response neutrophils also release nitric oxide and this results in nitration of tyrosine residues and blocks phosphorylation. however, scientists at the Immunology Disease Resistance Laboratory and the University of Maryland discovered that tyrosine nitration in neutrophils is associated with gland normality and low milk somatic cell counts. Tyrosine nitration can no longer be considered a pathologic result of inflammation. Indeed, nitration of tyrosine residues for some proteins in neutrophils represents a nonpathological state.
Technical Abstract: Nitration of tyrosine residues in bovine blood and mammary polymorphonuclear neutrophils (PMN) and milk proteins during acute mastitis was examined. Blood and milk were collected before and 12 h after intramammary injection of 50 æg Escherichia coli lipopolysaccharide (LPS). Nitrotyrosine residues in isolated PMN lysates and whey proteins were visualized by immunoblotting and quantitated by ELISA. Binding of the anti-bovine PMN mAb 36H10 (used as a positive control) to blood and milk PMN induced tyrosine phosphorylation and reversed tyrosine nitration after 20 s. Blood and milk PMN lysates had moderate and high levels, respectively, of nitrotyrosine before LPS injection. Nitrotyrosine levels tended to decrease in blood PMN (P < 0.2) and decreased significantly in milk PMN (P < 0.001) 12 h after LPS injection. Phosphotyrosines in blood and milk PMN increased after LPS injection. Nitrotyrosines and phosphotyrosines were present in whey proteins before injection. Tyrosine nitration of milk proteins decreased significantly in whey 12 h after LPS injection (P < 0.05). Milk protein tyrosine phosphorylation increased 12 h after LPS injection. In a second experiment, the statistical relationship between whey protein nitrotyrosine concentration and somatic cell count (SCC) was determined using composite milk from 42 cows. Nitrotyrosine concentration was negatively correlated with log10SCC (P < 0.001). Tyrosine nitration in blood and milk PMN and whey proteins was associated with gland normality and low SCC. Tyrosine nitration in bovine PMN is reversible, and may represent the normal state of many proteins in whey and PMN.