Enhanced host immune recognition of E.coli causing mastitis in CD-14 transgenic mice.

Authors: Wall, Robert; Powell, Anne; PAAPE, M - RETIRED; SOHN, E - NIH

Submitted to: Animal Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/7/2008
Publication Date: 1/9/2009
Citation: Wall, R.J., Powell, A., Sohn, E.J., Foster-Frey, J., Bannerman, D.D., and Paape, M. 2009. Enhanced host immune recognition of mastitis causing Escherchia Coli in CD-14 transgenic mice. Animal Biotechnology. 20:1-14

Interpretive Summary: Mastitis, a bacterial infection of mammary glands, cost the dairy industry almost $2 billion a year. That cost, in addition to medical expenses, includes losses in both quality and quantity of milk from infected cows. The loss in production can be attributed to damage to mammary glands, not by the invading organisms but by the over reaction of the cow’s immune system. Escherchia coli, an environmental pathogen, is responsible for a high percentage of mastitis cases. The innate immune system, the first line of defense against infection, initiates its attack by binding proteins to lipopolysaccharide (LPS), a molecule on the surface of E. coli. One of those proteins is CD-14 which resides in the membranes of neutrophils and macrophages. When bound to LPS CD-14 triggers an intercellular response resulting in the release of pro-inflammatory cytokines and interleukins that serve as chemoattractants which recruit phagocytes to deal with the invading bacteria. Excessive release of pro-inflammatory cytokines causes tissue damage. We hypothesized that by supplying a soluble form of CD-14 to compete with the membrane bound form we could lessen the chance of an over reaction by the innate immune system. To test that theory we produced transgenic mice expressing soluble CD-14 in their milk. We then infected those transgenic mice and non-transgenic controls and measured their response to infection. Most of the parameters measured: number of bacteria recovered, and various cytokines did not show the transgene to be of any advantage. However, one important parameter, the amount of edema in the infected mammary glands was clearly lowest in the transgenic animals that made the most CD-14. Those animals that made less CD-14 were less well protected. It was concluded that transgenic animals secreting a soluble form of CD-14 in their mammary glands could provide some protection against over stimulation of the innate immune response in the face of bacterial infection of mammary glands, thus having a beneficial effect on milk yield and quality.

Technical Abstract: Escherchia coli causes mastitis, an economically significant disease in dairy animals. E. coli endotoxin (lipopolysaccharide, LPS) when bound by host membrane proteins such as CD-14, causes release of pro-inflammatory cytokines recruiting neutrophils as a early innate immune response. Excessive pro-inflammatory cytokines causes tissue damage, compromising mammary function. If present soluble CD-14 (sCD-14) might out compete membrane bound CD-14, lessening severity of the inflammatory response. To test this hypothesis transgenic mice, expressing sCD14 in their milk (31 to 316 ug/ml) were evaluated. A cell culture study demonstrated in the presence of LPS, milk from transgenic mice increased secretion of cytokine IL-8 compared to milk from non-transgenic littermates (18 ± 3 vs. 35 ± 2 ng/mL, p < 0.001). To assess protection afforded by the transgene, glands were infused with E. coli. Recovery of bacteria showed no clear relationship between sCD14 concentration and the number of organisms recovered. However, there was a strong relationship between sCD14 concentration and edema (r2 = 0.999, p < 0.001), as measured by weight fluid in harvested glands. Highest expressing lines had the least edema suggesting the presence of sCD14 had an effect on reducing the inflammatory response to E. coli thus possibly protecting against gland tissue damage.