Transition Cow Health. Enhancing Immunity Through Vitamin D. Vitamin D, Calcium and Immunity. Transition Cow Health. Calcium

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1 Toll-like receptor Liver Blood Vitamin D receptor Vitamin D receptor 52nd Florida Dairy Production Conference 28 Gainesville, FL, April 6, 216 Enhancing Immunity Through Vitamin D Outcomes of Milk Check-Off Projects: Minimizing Costs of Mastitis Through Enhancing Antimicrobial Protein Production in the Udder -Awarded to Nelson and Santos Use of 1,25 Vitamin D3 (lcitriol) to Maintain Postpartum Blood lcium () and Improve Immune Function in Dairy Cows -Awarded to Santos and Nelson Corwin Nelson, Ph.D. Assistant Professor of Physiology Department of Animal Sciences Transition Cow Health Approach: Identify viable therapies that enhance immunity of transition cows. Basic Research: Understand how the immune system works and what signals control immunity of the cow. Applied Research: Test the effectiveness of immune modulating therapies that have potential for on-farm use. Transition Cow Health Our goal: Minimize incidence of transition cow diseases. Disease risk is minimized through proper nutrition and management. But, nutrition and management are not always enough. HO Vitamin D 3 Vitamin D, lcium and Immunity Vit D 3 25D 3 25D 3 1,25D 3 1,25D 3 CYP27B1 Macrophage RANTES Nitric Oxide β-defensins lcium Improved Immunity Disease Resistance

2 lcium Vitamin D3 HO Blood 1,25D3 CYP27B1 Inducible Nitiric Oxide Synthase (inos) Improved Immunity Disease Resistance Active 25D3 25D3 Toll-like receptor 25-Hydroxyvitamin D3 25(OH)D3 1,25-Dihydroxyvitamin D3 1,25(OH)2D3 Liver Relative fold increase Vit D3 [NO2-], mm 52nd Florida Dairy Production Conference Vitamin D helps activate antimicrobial defenses Vitamin D, lcium and Immunity 2 15 inos mrna P <.1, n = 6 Nitric oxide production P <.1, n = (OH)D 3 dose (ng/ml) Nelson, 21 J. Dairy Sci. Vitamin D helps activate antimicrobial defenses Hypothesis: 29 Toll-like receptor Liver 25D3 Blood 25D3 1,25D3 CYP27B1 16 Nature Reviews Immunology 3, (September 23) Monocyte DEFB3 mrna b 12 # Def transcripts/1 RPS9 Gainesville, FL, April 6, 216 Bovine β-defensins # Def transcripts/1 RPS9 Vit D b 4 6 a 31 Control a ,25D LPS LPS+1,25D Monocyte DEFB7 mrna b 4 2 ab a Control 1,25D Objective: Determine the effects of intramammary 1,25D treatment on induction of host defenses in the udder to understand mammary immunity. 14 cows Isolate Milk Somatic Cells 426 a 15 LPS LPS+1,25D Merriman, 215. J. Steroid Biochem. Mol. Biol. Vitamin D contributes to activation of antimicrobial defenses in the mammary gland. 1,25D3 Placebo Measure Gene Expression

3 52nd Florida Dairy Production Conference 3 Gainesville, FL, April 6, 216 inos/1 3 RPS9 bbd6/1 3 RPS Responses to Intramammary 1,25D Treatment inos P = bbd6 Gene Expression in Milk Somatic Cells bbd3/1 3 RPS9 bbd7/1 3 RPS bbd3 P = Hour Relative to Treatment bbd Hour Relative to Treatment P =.29 bbd4/1 3 RPS9 bbd1/1 3 RPS bbd4 P = bbd1 P = Responses to Intramammary 1,25D Treatment bbd7/1 3 RPS9 Gene Expression in Milk Somatic Cells bbd7 P = Hour Relative to Treatment Responses to Intramammary 1,25D Treatment inos/1 3 RPS9 Gene Expression in Milk Somatic Cells inos P = Hypothesis: Vitamin D contributes to activation of antimicrobial defenses in the mammary gland During mastitis Objective: Determine the effects of intramammary 1,25D treatment on induction of host defenses in subclinically infected glands. 15 cows 1,25D 3 or Placebo Isolate Milk Somatic Cells Measure Gene Expression Treatment repeated after each milking for 5 milkings (every 12 h)

4 52nd Florida Dairy Production Conference 31 Gainesville, FL, April 6, 216 inos/1 3 RPS9 bbd5/1 3 RPS Responses to Intramammary 1,25D Treatment: During Subclinical Mastitis inos bbd5 P =.13 P = bbd3/1 3 RPS9 bbd7/1 3 RPS bbd3 P = Hour from First Treatment bbd7 P = Hour from First Treatment bbd4/1 3 RPS9 bbd1/1 3 RPS DEFB bbd1 P =.39 P = Responses to Intramammary 1,25D Treatment: During Subclinical Mastitis Gene Expression in Milk Somatic Cells bbd4/1 3 RPS DEFB4 P = Responses to Intramammary 1,25D Treatment: During Subclinical Mastitis inos/1 3 RPS9 Gene Expression in Milk Somatic Cells inos P =.13 Minimizing Costs of Mastitis Through Enhancing Antimicrobial Protein Production in the Udder Conclusions: Activated vitamin D stimulates antimicrobial responses of cattle. Implications: Vitamin D requirement for immunity Therapeutic potential for vitamin D compounds in treatment of mastitis

5 52nd Florida Dairy Production Conference 32 Gainesville, FL, April 6, 216 Vitamin D, lcium and Immunity HO Vitamin D 3 25-Hydroxyvitamin D 3 25(OH)D 3 1,25-Dihydroxyvitamin D 3 1,25(OH) 2 D 3 Active Hypocalcemia lcium Improved Immunity Disease Resistance Onset of lactation causes transient decrease of calcium in circulation Hypocalcemia Circulating calcium is tighlty regulated Hypocalcemia Suppresses Immunity Immune cells require calcium to detect and eliminate pathogens Martinez et al. (212) J. Dairy Sci 95:

6 52nd Florida Dairy Production Conference Hypocalcemia Solutions to Hypocalcemia Control of sustained circulating calcium is complex Not as simple as providing more calcium Tightly controlled endocrine mechanisms Risk of metritis increases with decreased calcium Requires increases in absorption, retention, and mobilization of calcium lcium Mobilization Bone Martinez et al. (212) J. Dairy Sci 95: lcium Absorption Small intestine lcium Retention Kidney 33 Solutions to Hypocalcemia Role of Vitamin D Impaired even with good management Low 25-vitamin D 1,25-vitamin D 3 Active Inactive Gainesville, FL, April 6, 216 lcium Mobilization Active Bone lcium Absorption Small intestine 3 lcium Retention Kidney Use of 1,25 Vitamin D3 (lcitriol) to Maintain Postpartum Blood lcium () and Improve Immune Function in Dairy Cows Achilles Vieira-Neto and Jose E.P. Santos University of Florida, Gainesville, USA

7 52nd Florida Dairy Production Conference 34 Gainesville, FL, April 6, 216 Experimental Design Fifty cows fed negative DCAD prepartum Randomized to receive subcutaneous Placebo 3 μg 1,25-vitamin D Treatments given within 12 h of calving Measured responses of: feed intake, milk yield, energy balance, blood minerals, neutrophil function Effects of Post-Partum 1,25D on Minerals Total lcium Effects of Post-Partum 1,25D on Minerals Ionized lcium Total lcium Magnesium Phosphorous Effects of Post-Partum 1,25D on Neutrophils % Phagocytosis % Oxidative Burst Phagocytosis Intensity Oxidative Burst Intensity

8 52nd Florida Dairy Production Conference 35 Gainesville, FL, April 6, 216 Use of 1,25 Vitamin D3 (lcitriol) to Maintain Blood lcium () and Improve Immune Function Conclusions: 3 ug of 1,25D given immediately post-partum prevents subclinical hypocalcemia The 1,25D treatment increased neutrophil phagocytosis and oxidative burst Implications: Improved calcium and immunity status will lead to decreased transisiton cow diseases Potential adjunctive therapy to good transition cow management Serum 25(OH)D (ng/ml) Vitamin D Status of Dairy ttle Except: Whole milk-fed calves may be deficient Effect of Vitamin D supplementation on Dairy lf serum 25(OH)D Age (weeks) lves fed replacer of enhancer providing 5, 6, IU vitamin D per day lves fed pasteurized waste-milk and no sun Number of Cows Vitamin D Status of Dairy ttle Do dairy cattle get enough vitamin D? Distrubution of Serum 25(OH)D YES Cows sampled from 1 herds fed 3, IU to 5, IU per day No benefit achieved beyond 5,IU per day Serum 25(OH)D (ng/ml) Summary Intramammary 1,25D treatment boosts immune responses of the udder Post-partum subq 1,25D treatment prevents subclinical hypocalcemia and boosts neutrophil function Better knowledge of what factors contribute to a strong immune system

9 52nd Florida Dairy Production Conference 36 Gainesville, FL, April 6, 216 Acknowledments Funding: Southeast Milk Check-Off Nelson Lab: Kathryn Merriman Mercedes Kweh Michael Poindexter Jessi Powell Monika Trejos Paula Pulido Leslie Blakely Wayne Garcia Christopher Colston Santos Lab: Achilles Vieira-Neto Natalia Martinez Rafael Bisinotto Eduardo Ribeiro Fabio Lima Leandro Greco Leticia Sinedino milo Lopera I.R.P. Lima Francisco R. Lopes Jr. Roney Zimpel

10 NOTES 52nd Florida Dairy Production Conference 37 Gainesville, FL, April 6, 216

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