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Title: RECOMBINANT BOVINE SOLUBLE CD14 SENSITIZES THE MAMMARY GLAND TO LIPOPOLYSACCHARIDE

Author
item WANG, YAN - U MD COLLEGE PARK MD
item Zarlenga, Dante
item Paape, Max
item DAHL, GEOFFREY - U ILLINOIS URBANA IL

Submitted to: Veterinary Immunology and Immunopathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/21/2001
Publication Date: 6/10/2002
Citation: WANG, Y., ZARLENGA, D.S., PAAPE, M.J., DAHL, G.E. RECOMBINANT BOVINE SOLUBLE CD14 SENSITIZES THE MAMMARY GLAND TO LIPOPOLYSACCHARIDE. VETERINARY IMMUNOLOGY AND IMMUNOPATHOLOGY. vol. 86, pp. 115-24, 2002.

Interpretive Summary: Mastitis is the most costly disease in the dairy industry, with economic losses of approximately $1.8 billion annually in the United States (National Mastitis Council, 1996, pp 2). Mastitis results in decreased milk production, increased veterinary cost, and early culling or death of animals. Coliform mastitis is the most prevalent form of clinical mastitis, with infection by Escherichia coli being the most frequent. Approximately 80% of all intramammary infections by coliform bacteria will result in clinical mastitis, and about 10% will become peracute with the sudden onset of severe clinical symptoms of endotoxin shock. Because coliforms are present in the cow's environment, they cannot be eradicated on a practical basis. Conventional herd management practices such as pre- and post-milking teat dipping and dry cow antibiotic therapy are unable to reduce the incidence of new infections. In addition, antibiotic treatment, ,extensive fluid supplementation and metabolic support are not effective in relieving symptoms associated with clinical coliform mastitis. Scientists in the Immunology and Disease Resistance Laboratory have cloned the gene for producing a potent neutralizer of endotoxin. The expressed recombinant bovine CD14 will be used to treat cows suffering from acute septic shock. Insertion of the cloned gene into cows will result in transgenic cows resistant to infection by coliform organisms. Use of this product has the potential of saving the dairy industry $1.2 billion annually.

Technical Abstract: Standard therapies including administration of potent antibiotics, aggressive fluid resuscitation, and metabolic support, have not been successful in relieving symptoms and reducing mortality associated with acute coliform mastitis. Our laboratory has previously shown that macrophages and polymorphonuclear neutrophils in milk express CD14 on their rcell surface. We hypothesized that soluble CD14 (sCD14) is present in mil and sensitizes mammary ductal epithelial cells to low concentrations of lipopolysaccharide (LPS). This study demonstrated that: 1) a 46 kD protein in milk whey reacted with anti-ovine CD14 antibody; 2) under serum-free condition, complexes of LPS-recombinant bovine soluble CD14 (rbosCD14) induced activation of mammary ductal epithelial cells (as measured by changes in steady interlukin-8 (IL-8) mRNA level by competitive RT-PCR) at low concentrations of LPS after 6 or 24 h incubation (1 to 1000 ng/ml), whereas LPS alone did not induce activation of mammary ductal epithelial cells at the same concentrations; and 3) intramammary injection of low concentrations of LPS did not increase concentration of leukocytes in milk. In contrast, LPS-rbosCD14 complex containing the same concentration of LPS increased the concentration of leukocytes in the injected mammary gland at 12 and 24 h post injection. These results indicate that rbosCD14 sensitizes mammary epithelial cells to low concentrations of LPS in vitro and in vivo. Endogenous sCD14 in milk may be important in initiating host responses to Gram-negative bacterial infections.