Epidemiological investigation of insulin dysregulation in Shetland and Welsh ponies in Australia.

BACKGROUND: Insulin dysregulation (ID) is central to equine metabolic syndrome. There are limited epidemiological studies investigating dynamic testing of ID in ponies. OBJECTIVES: To evaluate prevalence and risk factors for ID through dynamic testing of hyperinsulinaemia (DHI) and insulin resistance (IR). STUDY DESIGN: Cross-sectional. METHODS: Sex, age, breed, height, cresty neck score (CNS), body condition score (BCS), laminitis, HMGA2:c.83G>A genotype and pituitary pars intermedia dysfunction (PPID) status were documented. Dynamic hyperinsulinaemia was diagnosed with an oral sugar test (OST) and IR with an insulin tolerance test (ITT). Owners completed surveys reporting activity, laminitis history and perception of body condition using a (1-9) visual analogue scale (VASo). Ordinal scores were converted to binary outcomes for CNS (≤2/5 or ≥3/5), BCS and VASo (≤6/9 or ≥7/9). Variables associated with insulin concentrations, glucose reduction after the ITT and laminitis were evaluated with mixed effects regression models accounting for random effects of farms. RESULTS: Among 167 ponies tested, median (range) age was 9 (4-21) years and BCS was 6 (4-8). Prevalence (95% confidence interval [CI]) of ID was 61 (53-68)%. Factors associated with insulin concentrations (estimate [95% CI]; µIU/mL) 60 min post-OST were: age (1.07 [1.02-1.11]), CNS (≥3/5, 1.52 [1.04-2.23]) and VASo (≥7/9, 1.75 [1.09-2.79]); and 90 min post-OST were: age (1.08 [1.03-1.12]), CNS (≥3/5, 1.80 [1.22-2.64]), VASo (≥7/9, 2.49 [1.52-4.08]) and sex (male, 0.64 [0.45-0.91]). Factors associated with glucose reduction after the ITT (estimate [95% CI]; %) were: age (-1.34 [-2.01 to -0.67]), sex (female, -6.21 [-11.68 to -0.74]) and VASo (≥7/9, -1.74 [-18.89 to -4.78]). Factors associated with laminitis (odds ratio [95% CI]) were DHI (4.60 [1.68-12.58]), IR (3.66 [1.26-10.61]) and PPID (11.75 [1.54-89.40]). MAIN LIMITATIONS: Single time-point sampling, laminitis definition and diet analysis. CONCLUSIONS: Ageing, being female and owner-perceived obesity were associated with ID.

Evaluation of an HMGA2 variant contribution to height and basal insulin concentrations in ponies.

BACKGROUND: The HMGA2:c.83G>A variant was identified in Welsh ponies having pleiotropic effects on height and insulin concentration. OBJECTIVE: Determine whether the HMGA2:c.83G>A variant is associated with decreased height and higher basal insulin concentrations across pony breeds. ANIMALS: Two hundred thirty-six ponies across 6 breeds. METHODS: Cross-sectional study. Ponies were genotyped for the HMGA2:c.83G>A variant and phenotyped for height and basal insulin concentrations. Stepwise regression was performed for model analysis using a linear regression model for height and mixed linear model for insulin with farm as a random effect. Coefficient of determination, pairwise comparison of the estimated marginal means and partial correlation coefficients (parcor) were calculated to assess the relationship between HMGA2 genotype and height or insulin. RESULTS: Breed and genotype accounted for 90.5% of the variation in height across breeds, and genotype explained 21% to 44% of the variation within breeds. Breed, genotype, cresty neck score, sex, age, and farm accounted for 45.5% of the variation in insulin, with genotype accounting for 7.1%. The HMGA2 A allele frequency was 62% and correlated with both height (parcor = -0.39; P < .001) and insulin (parcor = 0.22; P = .02). Pairwise comparisons found A/A ponies were >10 cm shorter than other genotypes. Compared with G/G individuals, A/A and G/A individuals had 4.3 µIU/mL (95% confidence interval [CI]: 1.8-10.5) and 2.7 µIU/mL (95% CI: 1.4-5.3) higher basal insulin concentrations, respectively. CONCLUSIONS AND CLINICAL IMPORTANCE: These data demonstrate the pleiotropic effects of the HMGA2:c.83G>A variant and its role in identifying ponies at increased risk for insulin dysregulation.

A one-health approach to identifying and mitigating the impact of endocrine disorders on human and equine athletes.

Endocrinopathies affect multiple species in ever-increasing percentages of their populations, creating an opportunity to apply one-health approaches to determining creative preventative measures and therapies in athletes. Obesity and alterations in insulin and glucose dynamics are medical concerns that play a role in whole-body health and homeostasis in both horses and humans. The role and impact of endocrine disorders on the musculoskeletal, cardiovascular, and reproductive systems are of particular interest to the athlete. Elucidation of both physiologic and pathophysiologic mechanisms involved in disease processes, starting in utero, is important for development of prevention and treatment strategies for the health and well-being of all species. This review focuses on the unrecognized effects of endocrine disorders associated with the origins of metabolic disease; inflammation at the intersection of endocrine disease and related diseases in the musculoskeletal, cardiovascular, and reproductive systems; novel interventions; and diagnostics that are informed via multiomic and one-health approaches. Readers interested in further details on specific equine performance conditions associated with endocrine disease are invited to read the companion Currents in One Health by Manfredi et al, JAVMA, February 2023.

Heritability of insidious uveitis in Appaloosa horses.

Equine recurrent uveitis (ERU) is a blinding ocular disorder among horses, and the Appaloosa horse breed is disproportionally affected by a chronic form of this intraocular inflammatory disease known as insidious uveitis. Strong breed predisposition and previous investigations suggest that there is a genetic component to the pathology of insidious uveitis among Appaloosa horses; however, no estimates of the heritability of the disease have previously been determined. This study aimed to characterize the genetic underpinning of the disease by estimating the heritability for insidious uveitis among Appaloosas. After combining two genotyping array datasets from the Illumina Equine SNP70 BeadChip and the Axiom Equine 670 K Genotyping Array, heritability was estimated for 59 affected and 83 unaffected horses using both restricted maximum likelihood (REML) and phenotype correlation - genotype correlation solvers from the linkage disequilibrium adjusted kinship software. Based on previous research, age and sex were used as covariates, and the locus responsible for the characteristic Appaloosa coat pattern (LP), previously associated with ERU risk, was included as a fixed effect ('top predictor'). Using prevalence values from 0.05 to 0.42, the heritability estimate for insidious uveitis ranged from 0.95 (SE = 0.14) to 1.74 (SE = 0.25) with LP contributing 0.16-0.33 to the estimate. This study suggests that insidious uveitis is highly heritable (REML 95% CI, h2  = 0.68-1.0) and additional loci outside of LP are contributing to the genetic risk for insidious uveitis for Appaloosas. Once identified, these other genetic factors may lead to new disease mitigation efforts in veterinary care and breeding practices.

Heritability and Genomic Architecture of Episodic Exercise-Induced Collapse in Border Collies.

An episodic nervous system disorder triggered by strenuous exercise, termed border collie collapse (BCC), exists in border collies and related breeds. The genetic basis of BCC is unknown but is believed to be a complex genetic disorder. Our goal was to estimate the heritability (h2SNP) of BCC, define its underlying genetic architecture, and identify associated genomic loci using dense whole-genome single-nucleotide polymorphism (SNP) genotyping data. Genotype data were obtained for ~440,000 SNPs from 343 border collies (168 BCC cases and 175 controls). h2SNP was calculated to be 49-61% depending on the estimated BCC prevalence. A total of 2407 SNPs across the genome accounted for nearly all the h2SNP of BCC, with an estimated 2003 SNPs of small effect, 349 SNPs of moderate effect, and 56 SNPs of large effect. Genome-wide association analyses identified significantly associated loci on chromosomes 1, 6, 11, 20, and 28, which accounted for ~5% of the total BCC h2SNP. We conclude that BCC is a moderately- to highly-heritable complex polygenetic disease resulting from contributions from hundreds to thousands of genetic variants with variable effect sizes. Understanding how much the BCC phenotype is determined by genetics and whether major gene mutations are likely to exist inform veterinarians and working/stock dog communities of the true nature of this condition.

Genome-Wide Analyses for Osteosarcoma in Leonberger Dogs Reveal the CDKN2A/B Gene Locus as a Major Risk Locus.

Dogs represent a unique spontaneous cancer model. Osteosarcoma (OSA) is the most common primary bone tumor in dogs (OMIA 001441-9615), and strongly resembles human forms of OSA. Several large- to giant-sized dog breeds, including the Leonberger, have a greatly increased risk of developing OSA. We performed genome-wide association analysis with high-density imputed SNP genotype data from 273 Leonberger cases with a median age of 8.1 [3.1-13.5] years and 365 controls older than eight years. This analysis revealed significant associations at the CDKN2A/B gene locus on canine chromosome 11, mirroring previous findings in other dog breeds, such as the greyhound, that also show an elevated risk for OSA. Heritability (h2SNP) was determined to be 20.6% (SE = 0.08; p-value = 5.7 × 10-4) based on a breed prevalence of 20%. The 2563 SNPs across the genome accounted for nearly all the h2SNP of OSA, with 2183 SNPs of small effect, 316 SNPs of moderate effect, and 64 SNPs of large effect. As with many other cancers it is likely that regulatory, non-coding variants underlie the increased risk for cancer development. Our findings confirm a complex genetic basis of OSA, moderate heritability, and the crucial role of the CDKN2A/B locus leading to strong cancer predisposition in dogs. It will ultimately be interesting to study and compare the known genetic loci associated with canine OSA in human OSA.

Evaluation of an HMGA2 variant for pleiotropic effects on height and metabolic traits in ponies.

BACKGROUND: Ponies are highly susceptible to metabolic derangements including hyperinsulinemia, insulin resistance, and adiposity. HYPOTHESIS/OBJECTIVES: Genetic loci affecting height in ponies have pleiotropic effects on metabolic pathways and increase the susceptibility to equine metabolic syndrome (EMS). ANIMALS: Two hundred ninety-four Welsh ponies and 529 horses. METHODS: Retrospective study of horses phenotyped for metabolic traits. Correlations between height and metabolic traits were assessed by Pearson's correlation coefficients. Complementary genome-wide analysis methods were used to identify a region of interest (ROI) for height and metabolic traits, determine the fraction of heritability contributed by the ROI, and identify candidate genes. RESULTS: There was an inverse relationship between height and baseline insulin (-0.26) in ponies. Genomic signature of selection and association analyses for both height and insulin identified the same ~1.3 megabase region on chromosome 6 that contained a shared ancestral haplotype between these traits. The ROI contributed ~40% of the heritability for height and ~20% of the heritability for insulin. High-mobility group AT-hook 2 was identified as a candidate gene, and Sanger sequencing detected a c.83G>A (p.G28E) variant associated with height in Shetland ponies. In our cohort of ponies, the A allele had a frequency of 0.76, was strongly correlated with height (-0.75), and was low to moderately correlated with metabolic traits including: insulin (0.32), insulin after an oral sugar test (0.25), non-esterified fatty acids (0.19), and triglyceride (0.22) concentrations. CONCLUSIONS AND CLINICAL IMPORTANCE: These data have important implications for identifying individuals at risk for EMS.

Heritability of Recurrent Exertional Rhabdomyolysis in Standardbred and Thoroughbred Racehorses Derived From SNP Genotyping Data.

Recurrent exertional rhabdomyolysis (RER) in Thoroughbred and Standardbred racehorses is characterized by episodes of muscle rigidity and cell damage that often recur upon strenuous exercise. The objective was to evaluate the importance of genetic factors in RER by obtaining an unbiased estimate of heritability in cohorts of unrelated Thoroughbred and Standardbred racehorses. Four hundred ninety-one Thoroughbred and 196 Standardbred racehorses were genotyped with the 54K or 74K SNP genotyping arrays. Heritability was calculated from genome-wide SNP data with a mixed linear and Bayesian model, utilizing the standard genetic relationship matrix (GRM). Both the mixed linear and Bayesian models estimated heritability of RER in Thoroughbreds to be approximately 0.34 and in Standardbred racehorses to be approximately 0.45 after adjusting for disease prevalence and sex. To account for potential differences in the genetic architecture of the underlying causal variants, heritability estimates were adjusted based on linkage disequilibrium weighted kinship matrix, minor allele frequency and variant effect size, yielding heritability estimates that ranged between 0.41-0.46 (Thoroughbreds) and 0.39-0.49 (Standardbreds). In conclusion, between 34-46% and 39-49% of the variance in RER susceptibility in Thoroughbred and Standardbred racehorses, respectively, can be explained by the SNPs present on these 2 genotyping arrays, indicating that RER is moderately heritable. These data provide further rationale for the investigation of genetic mutations associated with RER susceptibility.

Abnormal coagulation factor VIII transcript in a Tennessee Walking Horse colt with hemophilia A.

Hemophilia A is an X-chromosome-linked disorder caused by a deficiency in factor VIII (FVIII). Although foals have been diagnosed with hemophilia A based on deficiency in FVIII activity, causative gene mutations have not been identified. The genomic DNA and cDNA encoding FVIII of a Tennesee Walking Horse colt affected with hemophilia A and the genomic DNA of his dam and a normal unrelated horse were analyzed with no splice site or coding sequence abnormalities identified in any of the horses. Polymerase chain reactions (PCR) were then performed on hepatic cDNA from the affected colt and an unrelated normal horse, and no product was obtained for the sequence between and including exon 1 and exon 2 in the affected colt. Based on these results, suspected mutations were identified in the noncoding region of FVIII (intron 1), and genomic sequencing of intron 1 in the dam and the affected colt suggested maternal inheritance.