Using genomic approaches to detect the underlying molecular mechanisms for catastrophic fracture in Thoroughbred horses

Presenter Esther Palomino Lago
Authors Esther Palomino-Lago (1), Arabella Baird(2), Androniki Psifdi(1), Lucy Davison(1) and Debbie Guest(1)
Affiliations 1.Department of Clinical Sciences and Services, The Royal Veterinary College, Hawkshead Lane, North Mymms, Hatfield, Herts, AL9 7TA, UK., 2. Animal Health Trust, Lanwades Park, Kentford, Newmarket, CB8 7UU, UK.
Presentation Type Talk


Catastrophic fractures in horses are the most common type of injury to end a racehorse’s career and are the main cause of euthanasia in the UK. Fractures have a negative economic and welfare impact. Bone fracture is a complex disease and is caused by environmental and genetic factors. A genome wide association study (GWAS) showed significant genetic variation for catastrophic fracture risk on chromosomes 9, 18, 22 and 31. Chromosome 18 (ECA18) contains the most strongly associated region, which is highly conserved across species and includes 11 known genes that are involved in bone formation or fracture . We developed a genome wide polygenic risk score (PRS) to score a bank of 44 Thoroughbred cell samples for their relative genetic risk of catastrophic fracture. The PRS allowed us to generate equine induced pluripotent stem cells (iPSCs) based on their relative risk. We have differentiated iPSCs from the highest and lowest genetic risk horses into osteoblast-like cells and applied RNA-sequencing (RNA-seq) to analyze and compare the transcriptome expression between groups. RNA-seq analysis revealed that there are 112 differentially expressed genes (DEGs) between samples at high and low genetic risk of fracture. Gene ontology analysis associated the DEGs in numerous biological processes such as adhesion, development, morphogenesis and differentiation. Additionally, the DEGs were associated with ECM (extracellular matrix), collagen chain trimerization or ERK (extracellular signal-regulated kinase) pathways among others. Of the 112 DEGs, 27 are unannotated, 37 have published roles in bone formation or fracture and 48 have no known role in bone formation of fracture. To identify genetic variants significantly associated with the risk of fracture we performed Whole Genome Sequencing (WGS) on 14 samples, 7 cases and 7 controls. Variants were filtered based on their segregation between cases and controls, consequence (mis-sense or within 5 Kb upstream of the transcriptional start site), region of interest in the genome (ECA18 or DEGs), published role in bone formation or fracture and minor allele frequency (MAF) across breeds. 500 variants were then selected for genotyping using the Agena Bioscience MassARRAY platform to identify variants that are significantly associated with catastrophic fracture. Identification of causative genetic variants would provide a significant step forward to develop a DNA test to identify and manage high risk horses.