INVESTIGATION OF BIOMECHANICAL AND GENETIC RISK FACTORS FOR HOCK OSTEOCHONDROSIS

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: TERMINATED
• Funding Source: HATCH
• Reporting Frequency: Annual
• Accession No.: 1008733
• Project No.: ILLU-888-349
• Project Start Date: Dec 8, 2015
• Project End Date: Sep 30, 2017

Project Director
McCoy, AN.

Recipient Organization
UNIVERSITY OF ILLINOIS
2001 S. Lincoln Ave.
URBANA,IL 61801

Performing Department
Veterinary Research & Extension

Non Technical Summary
Osteochondrosis (OC) is a common manifestation of developmental orthopedic disease in young horses that is influenced by both genetic and environmental risk factors. Some OC lesions may heal spontaneously, but those present after 8 months of age are typically permanent and nearly always require surgical intervention. While OC can affect individuals of any breed, Standardbreds have a particularly high prevalence of tarsal lesions (10-26%). Differences in tarsal OC lesion prevalence and distribution between Standardbred pacers and trotters have been reported, but it is unknown if these are due to differences in genetic risk, biomechanical forces related to gait, or a combination of the two. We hypothesize that Standardbred pacers and trotters share genetic risk factors for OC, but that biomechanical differences in their natural gait patterns influence which early lesions heal and which become permanent. In Objective 1 we will prospectively follow the development of tarsal OC lesions in a cohort of Standardbred pacer and trotter foals to determine if gait preference has an impact on lesion healing/persistence. In Objective 2 we will determine if previously validated genetic risk factors are shared between pacers and trotters and determine if these risk alleles are associated with development and/or persistence of OC lesions in our experimental cohort. Prospective evaluation of differences in OC lesion formation between pacer and trotter foals will provide insight into the role that biomechanical forces related to gait may play in the development of disease. Further, this approach will allow us to determine if there are differences in genetic risk factors not just between pacers and trotters, but between horses who spontaneously heal their OC lesions and those who go on to develop permanent lesions. Defining the roles of these putative risk factors will both improve our understanding of the pathophysiology of OC and facilitate early interventions for at-risk horses to help reduce risk of clinical disease.

Animal Health Component

100%

Research Effort Categories

Basic

50%

Applied

50%

Developmental

(N/A)

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
311	3810	1080	50%
311	3810	1050	50%

Knowledge Area
311 - Animal Diseases;

Subject Of Investigation
3810 - Horses, ponies, and mules;

Field Of Science
1080 - Genetics; 1050 - Developmental biology;

Keywords

hock osteochondrosis

young horses

orthopedic disease

tarsal lesions

Goals / Objectives
We hypothesize that Standardbred pacers and trotters share genetic risk factors for osteochondrosis (OC), but that biomechanical differences in their natural gait patterns influence which early lesions heal and which become permanent. In Objective 1 we will prospectively follow the development of tarsal OC lesions in a cohort of Standardbred pacer and trotter foals to determine if gait preference has an impact on lesion healing/persistence. In Objective 2 we will determine if previously validated genetic risk factors are shared between pacers and trotters and determine if these risk alleles are associated with development and/or persistence of OC lesions in our experimental cohort. Prospective evaluation of differences in OC lesion formation between pacer and trotter foals will provide insight into the role that biomechanical forces related to gait may play in the development of disease. Further, this approach will allow us to determine if there are differences in modifying genetic risk factors not just between pacers and trotters, but between horses who spontaneously heal their OC lesions and those who go on to develop permanent lesions. Defining the roles of these putative risk factors will both improve our understanding of the pathophysiology of OC and is an important step towards our long-term goal of developing a predictive model that will allow for estimation of an individual horses' risk for the development of OC. Improved risk assessment will facilitate management changes and early disease intervention in at-risk individuals and allow for informed breeding decisions in high-risk breeds/pedigrees.

Project Methods
In Objective 1, 86 sire-matched Standardbred foals (n = 43 pacer; n = 43 trotter), raised under similar management conditions on one of three breeding farms in the central United States, will be enrolled in this study at 60 days of age. Four standard radiographic views - lateral, cranio-caudal, dorsolateral-palmaromedial oblique, and dorsomedial-palmarolateral oblique - of both tarsi will be taken using a portable digital x-ray unit beginning within one week of turning 60 days of age, and then at 60 day intervals until the foal reaches 8 months of age (i.e. 60 ± 7, 120 ± 7, 180 ± 7, 240 ± 7). Foals will also be video monitored in their normal paddock/pasture turnout at least 1 day (8-12 hours) per week. The video will subsequently be evaluated for the following activities: lying down, standing/nursing, walking, trotting and/or pacing, and moving at a faster gait (canter and/or gallop). A standardized activity log will be maintained for each foal to record the number of minutes (rounded to 30 second intervals) spent on each activity for a minimum of two consecutive hours each week.Radiographs will be randomized and blindly evaluated for the presence or absence of OC lesions. They will then be assembled serially for each foal to determine where lesions developed and whether they healed spontaneously or became permanent. An OC lesion will be considered permanent if it is seen on radiographs of a foal ≥8 months of age. Differences in presence (yes/no), location (DIRT, MM, lateral trochlear ridge of the talus [LTR], medial trochlear ridge of the talus [MTR]) and progression (healed/permanent) of OC lesions between pacers and trotters will be determined using Chi-squared analysis. Survival analysis (logrank statistic) will be used to determine if there is a difference in lesion development and healing between pacers and trotters over time. Overall activity will be assessed as percent time spent moving (combined time at the walk, trot/pace and canter/gallop divided by total observation time) during all observation periods prior to each set of radiographs (i.e. each 60 day period will be considered separately). Pacing will be evaluated in all foals as a categorical variable (observed/not observed). For those foals observed to pace, it will further be evaluated as a percentage of their total activity. Logistic regression models will be constructed to test the relationship between overall activity (in each 60 day period) and pacing with categorical outcome variables: 1) presence/absence of any OC lesion; 2) presence/absence of specific OC lesions (DIRT, MM, LTR, MTR); and 3) whether an observed OC lesion healed or became permanent. Sex and sire will also be included as predictor variables.For Objective 2, DNA will be isolated from either hair root or whole blood samples from all enrolled foals. Genotyping of 338 SNPs will be completed using a previously validated Sequenom genotyping assay. 98 of these SNPs are ancestry informative markers (AIMs) included to help control for population structure, while 240 are SNPs selected from chromosomal regions of interest associated with OC. Evaluation of additional risk alleles (not included in the Sequenom assay) is ongoing. We will genotype these risk alleles in our study cohort via restriction fragment length polymorphism (RFLP) or Sanger sequencing, as appropriate. Genotyping data will be analyzed using mixed model association analysis as implemented in the GEMMA software. Outcomes for which models will be created are 1) development of any OC lesion during the course of the study; and 2) development of a permanent OC lesion.

Progress 12/08/15 to 09/30/17

Outputs
Target Audience:Osteochondrosis (OC) is a commonly diagnosed developmental orthopedic disease in the horse. It is a complex disease, with interactions of genetics and environment (e.g. diet, exercise) determining expression and severity of lesions. Manifestations of disease can vary from mild to severe, and there is evidence that many lesions heal spontaneously. However, "permanent" OC lesions (present after 8 months of age) nearly always require surgical intervention to prevent ongoing joint damage. If left untreated, OC can potentially lead to severe degenerative joint disease and can be career- or even life-threatening. Although surgery is considered "curative" and treated horses can go on to perform as well as do their unaffected contemporaries, the cost of surgery and loss of training days represent a significant economic burden to the horse industry. Reduction in incidence, and ultimately, prevention of OC is an as-yet unattained goal of the equine industry, and thus this disease represents both an economically important entity and an animal health and welfare issue requiring ongoing research efforts. Identification of genetic risk factors is an important step towards identifying high-risk individuals for early intervention and making informed breeding decisions. Thus, this research will have direct impact on horse owners, breeders, and trainers, as well as veterinarians.This project is a multi-year study and is ongoing; USDA Hatch funds only supported the first year of the project. Changes/Problems:Unexpected foal loss during the first year of the project resulted in the addition of a second large breeding farm in the second year of the project. Depending on foal loss in this second year, a third year may be required to enroll appropriate numbers, and this has been funded through non-Hatch sources. What opportunities for training and professional development has the project provided?To date, this project has provided three veterinary students with summer research experiences under the mentorship of the PI (twoin Summer 2016, onein Summer 2017). Additionally, sevenundergraduate animal science students interested in a career in veterinary medicine have been involved in the field observation portion of the study; two of these students (involved with the 2016 foals) startedtheir first year of veterinary school in Fall 2017. Two veterinary interns have also completed small research projects with the preliminary data generated by this project. This project also comprisesa major portion of the MS thesis work for a graduate student/resident in veterinary clinical medicine. How have the results been disseminated to communities of interest?As this project is ongoing, dissemination of preliminary results has largely been restricted to the owners of the individuals enrolled in the study. However, the three veterinary students who worked on this project as summer research scholars each presented a poster of their work at a college research symposium and/or a national symposium for summer research scholars. What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Twenty foals born in 2016 completed the project, with radiographs and field observations through twelvemonths of age. Six foals had osteochondrosis lesions at twelvemonths of age, compared to elevenfoals at twomonths of age; this was consistent with our expectations. Genotyping has been completed (Affymetrix 670k SNP chip) for these foals, but analysis will not be performed until the entire cohort is complete. Seventy-five foals born in 2017 have been enrolled in the project and will havetheir eightmonth films completedby Jan 2018. To date, weekly field observations have revealed little difference in activity between pacers and trotters. This project is ongoing, with non-Hatch funding secured through July 2019.

Publications

Progress 12/08/15 to 09/30/16

Outputs
Target Audience:Preliminary data from the first few months of this project has been shared with academic scientists and veterinarians attending a summer scholar research day via aposter presentation by the professional veterinary studentinvolved in the project. Changes/Problems:We were only able to enroll about half of the expected number of foals in the first year's cohort due to unexpected pregnancy loss at the breeding farm we are working with. We have identified a second breeding farm from which to enroll foals for the second year cohort to overcome this issue and expect to be able to complete our cohort enrollment within the planned two year duration of the project. What opportunities for training and professional development has the project provided?Although training and professional development was not a stated goal of this study, it has provided some training opportunities. One veterinary student was involved in the project as a summer scholar and got to beinvolved in aspects of study design, protocol development, and data collection and analysis. Other veterinary students, interns, and residents have gotten a chance to practice their field radiography skills by helping out with the project. 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?The foals currently enrolled in the project will be followed out until they are 12 months of age (radiographs and field observation). Foals born in spring 2017 will enter the study protocol at 2 months of age and will receive serial radiographs and field observations similarly to the first year's cohort. Evaluation of genetic data will be performed after the complete study cohort has been identified and genotyped. Statistical analysis of field observation data will be performed after the complete study cohort has been followed out to 12 months of age.

Impacts
What was accomplished under these goals? Twenty-five Standardbred foals born in the spring of 2016 were enrolled in this project (approximately half pacers and half trotters); of these, 3 died and 2 were sold prior to reaching 8 months of age and will therefore not contribute to the final data set. The remaining foalshave been radiographed every 2 months and have undergone field observation weekly. Osteochondrosis (OC) lesions were initially suspected in approximately half of the enrolled foals, but most lesions have been observed to regress over time, as expected. Currently, approximately 20% of the foals show radiographic evidence of OC, which is consistent with our expectations for this breed. To date, field observations have not revealed significant differences in activity level between pacer and trotter foals. Whole-genome genotyping has been performed on the enrolled foals, but this data will not be evaluated until the second year of the study.

Publications