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United States Department of Agriculture

Agricultural Research Service

Research Project: IDENTIFICATION OF FACTORS ASSOCIATED WITH IMMUNE SUPPRESSION AND MASTITIS
2011 Annual Report


1a.Objectives (from AD-416)
Develop proteomic profiles of immune cells during the acute and chronic stages of mastitis and under different physiologic conditions known to be associated with immune suppression, such as parturition or nutritional imbalances. Identify proteins that are up- or down-regulated during these immunosuppressive states. Initiate studies into the function of these differentially regulated proteins.

Develop proteome profiles of mastitis-causing bacteria isolated from in vivo conditions and in bacteria grown in vitro or in lab counterparts that are not considered highly virulent. Identify proteins that are up- or down-regulated across bacteria studied in each ecological state and initiate studies into the role these differentially regulated proteins play in the establishment of chronic infection of the mammary gland.

Studies of immunomodulators to test their effectiveness at preventing disease by imposing them on a controlled reproducible mastitis challenge model. In the course of these studies we will discover and develop immunological reagents for the bovine and relevant wildlife animal species that will facilitate the discovery of innovative immunointervention strategies.


1b.Approach (from AD-416)
The approach to this research project is through experimentation discover new insights into the mechanisms of how the bovine dairy cow immune system fights diseases. Failure of the dairy cow immune system results in numerous diseases, of which mastitis is the predominate disease resulting in an economic burden to the industry. Our research will investigate the host immune system – pathogen interaction with the goal of enhancing immune function. To accomplish this goal we will study how immune system functions are affected by nutritional status. We will also study a newly described immune system function in the context of the dairy cow. In addition, we will investigate how pathogenic bacteria adapt to the in vivo environment and escape immune clearance. Finally, we will add to the large animal immunological reagent toolbox, to aid in study of immunological questions in dairy animals for the entire research community.


3.Progress Report
Mastitis is the most common disease that affects the dairy industry. Our work is designed to better understand the basic mechanisms of the immune response to this disease and how we can help the dairy farmer better protect their herds. We have determined that vitamin D affects bovine immune function. This year, we have published a paper that demonstrates the up-regulation of the vitamin D pathway in immune cells isolated from milk from cows with clinical mastitis. This is the first demonstration of association of the vitamin D pathway in immune cells responding to an infection in any species. In addition, we have published a paper demonstrating the role of vitamin D in the adaptive immune system, elucidating the complex interaction between vitamin D and immune cells. This work is some of the only vitamin D research in vivo, in any species, and represents significant progress in understanding the role of vitamin D in infectious diseases. We also have demonstrated that intramammary vitamin D administration at the time of infection reduces the severity of disease. This work is the subject of a provisional patent and we are in discussions with pharmaceutical companies regarding use as a treatment for mastitis.


4.Accomplishments
1. Determination of the role of vitamin D in an infected mammary gland. Mastitis is the biggest problem in the dairy industry, costing the industry roughly $2 billion annually. ARS researchers at Ames, IA, are working to enhance the cow’s immune system to better fight this disease. ARS researchers have shown that vitamin D influences gene expression in bovine immune cells. We have identified the specific immune cell types that express the enzyme that activates the precursor vitamin D. Immune cells of the monocyte lineage were shown to express ribonucleic acid (RNA) for the enzyme that activates the vitamin D pathway, as well as activating other genes with immunological functions. The impact is a better understanding of the effect of vitamin D, at a molecular level, will allow for determination of cellular and molecular pathways that may be targeted for therapeutic interventions and the subject of future research.

2. Reduction of mastitis severity with use of vitamin D. ARS researchers at Ames, IA, have shown that vitamin D influences gene expression in bovine immune cells in the mammary gland. The milk compartment of the mammary gland is deficient in vitamin D, even if the levels are normal in the blood. Therefore, vitamin D therapy may uniquely affect the molecular and cellular pathways in immune cells in the mammary gland and may be an important non-antibiotic option for mastitis treatment. We have infected animals with mastitis and treated the infection with vitamin D. Preliminary data shows that vitamin D treatment reduces the severity of the infection. Vitamin D is a simple and natural immune stimulator that in combination with current antibiotics could become an effective therapy for mastitis. In addition, vitamin D’s ability to stimulate the immune system could reduce the time and amount of antibiotics needed to treat mastitis. This combination therapy may cure mastitis infections that are currently resistant to antibiotic treatment alone. The result of a reduction of antibiotic use would be reduced antibiotic residues that may get into the food supply, and an increase in consumer confidence and international trading opportunities.


Review Publications
Lippolis, J.D., Reinhardt, T.A. 2010. Utility, limitations, and promise of proteomics in animal science. Veterinary Immunology and Immunopathology. 138(4):241-251.

Nelson, C.D., Reinhardt, T.A., Beitz, D.C., Lippolis, J.D. 2010. In vivo activation of the intracrine vitamin D pathway in innate immune cells and mammary tissue during a bacterial infection. PLoS One. 5(11):e15469. Available: http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0015469.

Nelson, C.D., Nonnecke, B.J., Reinhardt, T.A., Waters, W.R., Beitz, D.C., Lippolis, J.D. 2011. Regulation of Mycobacterium-specific mononuclear cell responses by 25-hydroxyvitamin D3. PLoS One. 6(6):e21674. Available: http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0021674.

Last Modified: 4/16/2014
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