Source: UNIV OF MARYLAND submitted to NRP
THE ROLE OF KETONES ON ENERGY METABOLISM, MILK PRODUCTION, AND THE DEVELOPMENT OF PERIPARTAL DISEASE IN DAIRY COWS
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
COMPLETE
Funding Source
AFRI COMPETITIVE GRANT
Reporting Frequency
Annual
Accession No.
1030162
Grant No.
2023-67015-39360
Cumulative Award Amt.
$650,000.00
Proposal No.
2022-07989
Multistate No.
(N/A)
Project Start Date
Apr 15, 2023
Project End Date
Oct 10, 2024
Grant Year
2023
Program Code
[A1231]- Animal Health and Production and Animal Products: Improved Nutritional Performance, Growth, and Lactation of Animals
Recipient Organization
UNIV OF MARYLAND
(N/A)
COLLEGE PARK,MD 20742
Performing Department
(N/A)
Non Technical Summary
Ketosis has been indentified as a significant factor associated with disease and poor performance in dairy cows entering lactation. Put together,ketosis and its associated complications constitute a welfare concern, and represent an economic burden of hundreds of millions of dollars for dairy farmers in the United States alone. Despite decades of research and collective efforts to reduce the incidence of ketosis and its presumed negative impacts, our ability to mitigate this disorder and its consequences remainslimited.An increasingly strong body of evidence indicates that ketones can have positive effects on metabolism and the control of inflammation in rodents and humas; furthermore, ketosis cansometimes be associated with normal cow health and productivity. This apparent contradiction indicates that our understanding of ketones themselves and their roles on animal metabolism and metabolic health is currently imperfect. Our goalis to directly address our gaps in knowledge byperforming a series of complementary studies thatunequivocally establish the roles of ketones during health and disease.We anticipate that our research will revealthe specific conditions of metabolism and cow physiology in which ketones may promote normal and healthy metabolism, and separate these from those conditions that lead to negative outcomes of health and performance. This research is expected to provide new knowledge and management toolsto optimizelactation performance and welfare in dairy cows, thus promoting thesustainability of the dairy industry.
Animal Health Component
30%
Research Effort Categories
Basic
70%
Applied
30%
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
3023410101050%
3053410102050%
Knowledge Area
302 - Nutrient Utilization in Animals; 305 - Animal Physiological Processes;

Subject Of Investigation
3410 - Dairy cattle, live animal;

Field Of Science
1010 - Nutrition and metabolism; 1020 - Physiology;
Goals / Objectives
Our long-term goal is to improve the productive lifespan of dairy cows via an enhanced understanding of cow biology and metabolism. Our research will build a strong foundation for the development of novel and effective management strategies to increase production efficiency, and thus promote the sustainability of the dairy industry.With this research project, we aim to unveil the factors that predispose cows to reduced milk production performance and increased disease propensity during ketosis. Our investigation will allow the comprehensive understanding of factors leading to metabolic dysfunction and will specifically elucidate the role of ketosis in this context. Our specific goals are the following:1. Identify the metabolic, physiological, and behavioral phenotypes that distinguish and predict normal or aberrant trajectories of health and milk production during ketosis.2.Evaluate the effects of induced ketosis with or without concurrent hyperlipidemia and inflammation on cow metabolism, productivity, and health3.Evaluate the ability of ketones to improve the metabolic adaptation to energy deficit during peripartum, and thus to prevent metabolic dysfunction and its sequelae
Project Methods
To achieve or goals, we will utilizecomplementary research methodologies, leveraging the advantages of observational population-wide studies on the one hand, andthose of randomized controlled experimentation, on the other.1. Our population data will be utilized to identify early biological signatures that define whether a dairy cow will undergo either a normal or dysfunctional health trajectory during ketosis. In Aim 1 of this project,data for behavioral, physiological, and performance parameters collected by automated cow wearable and non-wearable sensors, health status determined by clinical examination, and blood samples from peripartal cows that meet the following categorical criteria will be utilized in 1) cows that remained healthy and did not develop ketosis; 2) cows diagnosed with ketosis but that displayed no clinical signs of disease; 3) cows diagnosed with clinical ketosis and clinical signs of others diseases. These categories will constitute the study groups, including primiparous and multiparous cows.2. Randomized controlled research will be based on the experimental induction of ketosis, which is set allow for a specific evaluation of the effects of ketones on cow physiology, while reducing the noise from other potentially confounding factors. In experiment 1, healthy multiparous lactating dairy cows will be blocked by stage of lactation, as early lactationor post-peak lactation and randomly assigned to either dietary-induced hyperketonemia (Calcium butyrateproduct --40% butyrate--fed at 3% of dietary DM 2x/d) or Ca-balanced control in a replicated Latin Square design. In experiment 2, healthy multiparous postpartum dairy cowswill be enrolled in a longitudinal randomized controlled trial to evaluate the acute effects of hyperlipidemia, immune-induced inflammation, and their combination, on metabolic, physiological, and production responses during ketosis. Cowswill be randomly allocated to 4 treatments as follows: 1) ketosis control (Ca-Bu at 3% of dietary DM 2X/d + i.v. saline solution injection); 2) ketosis + hyperlipidemia (Ca-Bu + i.v. tallow triglyceride emulsion 20%; 100 mL/h); 3) Ketosis + LPS (Ca-Bu + O55:B5 E. coli LPS injected at 1.5 μg/ kg of BW [94]); 4) Ketosis + hyperlipidemia + LPS (Ca-Bu + tallow emulsion + LPS). In all cases, the induction period will last 24 h. In experiment 3, the effects of feeding a ketogenic diet during the peripartum will be evaluated in lean (BCS = 3.25) and overweight (BCS ≥ 4.0) dairy cows. Animals will be enrolled on d -30 and randomly allocated to a standard prepartum diet (Control) or a prepartum ketogenic diet (Ketone; Ca-Bu at 3% of DM 2X/d) fed until parturition (21 d).Insulin sensitivity will be evluated via glucose tolerance testing, ad liver biopsies will be performed pre- and post-partum.

Progress 04/15/23 to 10/10/24

Outputs
Target Audience:Target audiences include scientific peers in the academic cimmunity, as well as dairy industry stakeholders, such as technicians, veterinarians, and dairy producers. Changes/Problems:Fortunately, we have completed the validation steps we had planned for the induction of hyperketonemia, as well as our work with immune cells for the study of their function during ketosis. We have not faced major obstacles to the execution of our plans so far. However, given the fact that I have recently trasferred to a new university, I would be interested in requesting a 1-year extension to this project. This is to account for the time lost associated with the change, and to guarantee that we can exceute our plans as intended. What opportunities for training and professional development has the project provided?Two graduate students and 8 undergraduate students have participated in the preliminary work completed thus far. Students were trained in general husbandry practices (feeding, milking, handling, weighing, etc), the collection of biological smaples (i.e., blood, liver, and milk), benchtop analysis (metabolites, hormones, and cytokines), and the statistical analysis of data. The hands-on experience has given undergrads critical experince in management and research that they can leverage as they build their future careers in science or clinical practice. The upcoming activities of this project will see the involvement of new graduate students, undergraduates, and post doctoral researchers. How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals?This is the final technical report (FTR) for the project at the University of Maryland; however, this project is being transferred to the University of Pennsylvania. The majority of our project objectives will be accomplished at this new institution. We have planned to complete the specific experimental sections and data analysis within the timeline mentioned in the previous sections. This means that we plan to complete most of the work for Aim 1 during 2024, while Aim 2 and 3 areplanned be completed during 2025.

Impacts
What was accomplished under these goals? Goal 1. This part of the work is being done in collaboration with Cornell University. At this point, we have selected the representative groups within the population of cows available (NY state farms) as follows: 1) normoketonemic and healthy cows (control); 2) hyperketonemic and healthy cows, and 3). We have identified differentiual patterns of behavior (e.g., rumination) and productivity (i,e., milk yield) for each of these groups. Our current work (set to start in September of 2024) is the analysis of blood metabolites (traditional markers and polar components of the metabolome) and inflammatory markers in the above-mentioned groups. Subsequent steps include the merging of behavioral, physiological, and metabolic markers, for the exploartion of the best predictors (single or combined) of positive and negative trajetcories of health in cows that develop hyperketonemia. We anticipate finalize this work in the spring of 2025. Goal 2. Via preliminary testing, we have thus far validated the models for the indiuction of hyperketonemia, namely intravenous infusion of ketones, and the nutritional induction of ketosis. a total of 3 preliminary experiments were completed that succesfully induced hyperketonemia at the subclinical level range. Hyperketonemia was sustained for 48 and 72 hours across these experiments in both early lactation cows, and mid lactation cows. Furthermore, we evaluated the impact of hyperketonemia on immune and productive responses during a lipopolysaccharide challenge (bolus infusion). Furthermore, we have validated assays for the evaluation of immune fucntion ex vivo (i.e., isolated neutrophils), from cells obtained of cows in whic hyperketonemia was induced. Our next step includes the evalution of concommitant hyperlipidemia in animals that are experiencing ketosis. This experiment will include the evaluation of immune responses (before and after an LPS challenge) in order to address the question of the interaction between ketosis, hiperlipidemi, and immune function. We anticipate completing the work in the spring of 2025. Goal 3. Our preliminary experiments validated the feasibility of the nutritonal induction of ketosis (nutritional ketosis) using clacium butyrate. Beyond validating the model and observing no deletreous efefcts of nutritional ketosis on cow health or productivity, no further progress has been made towards this goal yet. Experimental work towards this goal is expected to be initiated during the second half of 2025.

Publications


    Progress 04/15/23 to 04/14/24

    Outputs
    Target Audience: Nothing Reported Changes/Problems:Fortunately, we have completed the validation steps we had planned for the induction of hyperketonemia, as wellas our work with immune cells for the study of their function during ketosis. We have not faced major obstacles to the execution of our plans so far. However, given the fact that I have recently trasferred to a new university, I would be interested in requesting a 1-year extension to this project. This is to account for the time lost associated with the change, and to guarantee that we can exceute our plans as intended. What opportunities for training and professional development has the project provided?Two graduate students and 8 undergraduate students have participated in the preliminary work completed thus far. Students were trained in general husbandry practices (feeding,milking, handling, weighing, etc), the collection of biological smaples (i.e., blood, liver, and milk), benchtop analysis (metabolites, hormones, and cytokines), and the statistical analysis of data. The hands-on experience has given undergradscritical experince in management and research that they can leverage as they build their future careers in science or clinical practice. The upcoming activities of this project will see the involvement of new graduate students, undergraduates, and post doctoral researchers. How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals?We have planned to completethe specific experimental sections and data analysis withinthe timeline mentioned in the previous sections. This means that we plan to complete most of the work for Aim 1 during 2024, while Aim 2 and 3 are planned be completed during 2025.

    Impacts
    What was accomplished under these goals? Goal1. This part of the work is being done in collaboration with Cornell University. At this point, we have selected the representative groups within the population of cows available (NY state farms) as follows: 1) normoketonemic and healthy cows (control); 2) hyperketonemic and healthy cows, and 3). We have identified differentiual patterns of behavior (e.g., rumination) and productivity (i,e., milk yield) for each of these groups.Our current work (set to start in September of 2024) is the analysis of blood metabolites (traditional markers and polar components of themetabolome) and inflammatory markers in the above-mentioned groups. Subsequent steps include the merging of behavioral, physiological, and metabolic markers, for the exploartion of the best predictors (single or combined)of positive and negative trajetcories of health in cows that develop hyperketonemia.We anticipate finalize this work in the spring of 2025. Goal 2.Via preliminary testing,we have thus far validated the models for the indiuction of hyperketonemia, namely intravenous infusion of ketones, and the nutritional induction of ketosis. a total of 3 preliminary experiments were completed that succesfully induced hyperketonemia at the subclinical level range. Hyperketonemia was sustained for 48 and 72 hours across these experiments in both early lactation cows, and mid lactation cows. Furthermore, we evaluated the impact of hyperketonemia on immune and productive responses during a lipopolysaccharide challenge (bolus infusion). Furthermore, we have validated assays for the evaluation of immune fucntion ex vivo (i.e., isolated neutrophils), from cells obtained of cows in whic hyperketonemia was induced. Our next step includes the evalution of concommitant hyperlipidemia in animals that are experiencing ketosis. This experiment will include the evaluation of immune responses (before and after an LPS challenge) in order to address the question of the interaction between ketosis, hiperlipidemi, and immune function. We anticipate completing the work in the spring of 2025. Goal 3. Our preliminary experiments validated the feasibility of the nutritonal induction of ketosis (nutritional ketosis) using clacium butyrate. Beyond validating the model and observing no deletreous efefcts of nutritional ketosis on cow health or productivity, no further progress has been made towards this goal yet. Experimental work towards this goal is expected to be initiated during the second halfof 2025.

    Publications