Plasma concentration of serum amyloid A and lipopolysaccharide binding protein in horses with laminitis resulted from hindgut acidosis.

Many studies have shown a strong correlation between Hindgut Acidosis (HGA) and the occurrence of laminitis in horses; therefore, the early diagnosis of HGA is essential. In this study, we investigated changes in the plasma concentrations of lipopolysaccharide-binding protein (LBP) and serum amyloid A (SAA) as inflammatory markers in horses with laminitis. Sixteen healthy male Arabian horses that had cecal cannulation without visible laminitis or general symptoms were randomly divided into two groups. The horses were fed two different diets in a forage-to-concentrate ratio. Blood samples were collected on Days 1, 10, and 20. The primary objective of this study was to analyze plasma levels of LBP and SAA. Cecal specimens were obtained from each equine subject on three designated days: days 1, 10, and 20. The second objective was to assess the levels of pH and volatile fatty acids (VFA) in the samples. Throughout the study period, horses fed a high-concentrate diet exhibited a significantly elevated average lameness grade on days 10 and 20 compared to the initial stage (P < 0.001). On day 20, a significant increase in the concentration of SAA was observed in horses fed a high-concentrate diet, in contrast to the initial stage of the study. LBP levels in the plasma were significantly elevated on days 10 and 20 in horses fed a high-concentrate diet. Based on our findings, it is recommended that the evaluation of plasma LBP concentrations is more effective than SAA for the early identification of HGA in horses fed a high-grain diet.

Sodium-glucose transport protein 2 inhibitor use in the management of insulin dysregulation in ponies and horses.

Laminitis is a common and painful condition of the equine foot and approximately 90% of cases are associated with insulin dysregulation (ID) that is a central feature of the common endocrine disorder equine metabolic syndrome (EMS) and occurs in a subset of animals with pituitary pars intermedia dysfunction. Additional features of EMS include obesity, altered circulating concentrations of adipokines (particularly adiponectin and leptin) and hypertriglyceridaemia. Obesity, ID, hypoadiponectinaemia, hyperleptinaemia and an altered plasma lipid profile are also features of human metabolic syndrome (HMS) alongside hyperglycaemia. Sodium-glucose cotransporter 2 inhibitors (SGLT2i) are a novel class of oral hypoglycaemic agents used in combination with lifestyle changes in the management of HMS. SGLT2 receptors are responsible for 90% of the renal glucose reabsorption that occurs in the proximal convoluted tubule. Thus, these drugs increase urinary glucose excretion by suppressing glucose reabsorption from the glomerular filtrate resulting in urinary calorie loss with consequent weight loss and improvements in ID, hyperglycemia, hypoadiponectinaemia and hyperleptinaemia. There are no licenced veterinary drugs available for treating ID and preventing insulin-associated laminitis in horses. Thus, the use of SGLT2i for the control of equine hyperinsulinaemia with the goal of improving recovery from associated active laminitis or preventing future laminitis has recently been advocated. There are a small number of published studies reporting the use of the SGLT2i canagliflozin, ertugliflozin and velagliflozin to aid the management of equine ID. However, the doses used are largely extrapolated from human studies with limited consideration of species-specific variations. In addition, there is limited evaluation of the fundamental differences between ID in horses and humans, particularly the fact that most horses with ID remain hyperinsulinaemic but normoglycaemic such that increased urinary loss of glucose may not explain the beneficial effects of these drugs. Further study of the potential deleterious effects of treatment-associated hypertriglyceridaemia is required together with the effect of SGLT2i therapy on circulating concentrations of adipokines in horses.

The Detection of Thoracolumbar Spine Injuries in Horses with Chronic Laminitis Using a Novel Clinical-Assessment Protocol and Ultrasonographic Examination.

Postural adaptation is a prominent feature in horses affected by laminitis. Laminitis induces intense pain, especially in the forelimbs, prompting affected horses to assume a caudally displaced trunk posture, resulting in the hyperflexion of the thoracolumbar spine. This study assessed the nature and prevalence of thoracolumbar injuries in horses with chronic laminitis compared to horses without it. Sixty horses were used (thirty laminitic and thirty non-laminitic) of different athletic purposes and ages (2-20 years). The experimental protocol entailed a single assessment of horses' thoracolumbar spines, utilizing physical examination by MACCTORE, a scoring system developed specifically for this study. Additional evaluations included the Grimace Equine Pain Scale (HGS) and ultrasound exams. Statistical tests were used to compare values (Mann-Whitney or t-test) and lesions prevalences (Fisher) between groups (p < 0.05). The results showed a higher pain manifestation (HGS and heart rate, p < 0.0001) and thoracolumbar-spine-injury levels in chronic laminitis horses, both in MACCTORE clinical examinations (11.7 ± 4.8 vs. 4.2 ± 3.3, p < 0.0001) and general ultrasonographic indices (39.6 ± 12.0 vs. 20.7 ± 7.1, p < 0.0001), including specific examination approaches for various spinal elements. Horses with laminitis presented with a 14-fold higher prevalence of ultrasound-relevant lesions in the thoracolumbar spine (CI: 4.4 to 50.6, p < 0.0001) compared to controls. These findings constitute new evidence of an association between chronic laminitis and the presence of thoracolumbar spine injuries in horses, which may be confirmed by more sophisticated study designs.

Radiological measurements of the feet of normal Straight Egyptian Arabian horses in Qatar.

The Arabian horse has been identified as carrying a risk locus for equine metabolic syndrome, predisposing this breed to development of laminitis. Radigraphy of the equine foot is widely considered the main diagnostic imaging technique for evaluation of the laminitic horse. Knowledge of 'normal' breed values allows assessment of the degree and severity of radiological changes associated with laminitis. The objective of this study was to investigate the normal values for radiological measurements of the feet of the Straight Egyptian Arabian horse in Qatar. The design was a clinical prospective study. Radiographs of the fore and hind feet of 10 clinically normal adult Straight Egyptian Arabian horses were taken. On the lateromedial views, 17 measurements were taken (13 distances and four angles). On the horizontal dorsopalmar/plantar views, two measurements were taken. On the dorsal 45 degree, proximo-palmarodistal oblique projections, four measurements were taken. Normal reference ranges were reported for radiological measurements of the feet of the Straight Egyptian Arabian horse. Several variables showed significant differences between fore and hind feet, including hoof angle, distal wall thickness, and two proximal inner layer measurements (p < 0.05). In addition, the Straight Egyptian Arabian horse was found to have a number of measurements which varied from previously published reports. The results reported within provide a useful reference for normal radiographic measurements of the Straight Egyptian Arabian horse with relevance for laminitis.

Investigation of glucagon-like peptide-1 response to six oral carbohydrates in ponies.

Glucagon-like peptide-1 (GLP-1), the principal incretin in horses, may play a role in the pathophysiology of insulin dysregulation (ID). This study aimed to describe its concentration in response to three preserved forages and four dynamic tests for ID in ponies. Twelve adult ponies of mixed ID status were given a meal of hay, soaked hay or haylage, an in-feed oral glucose test (OGT), oral sugar test (OST), an oral test using a proprietary breakfast cereal (WEET) or a combined glucose-insulin tolerance test (CGIT) weekly in a randomised cross-over study. Glucose, insulin and GLP-1 concentrations were measured before and following each intervention. Ponies were designated ID or non-ID and insulin resistant (IR) or non-IR according to OGT and CGIT results, respectively. All interventions apart from the CGIT provoked a GLP-1 response within 30 min. The OGT and WEET interventions, (containing the greatest dose of non-structural carbohydrate, 1.06 and 1 g/kg BW, respectively), resulted in a greater area under the curve (AUC) for GLP-1 compared to all other interventions (P < 0.001). No difference in GLP-1 response was detected according to ID or IR status, despite there being strong positive correlations (rs [95 % CI]) between GLP-1 and insulin concentrations measured at individual time points (0.67 [0.62 - 0.71]; P < 0.001) and as AUC (0.66 [0.49-0.79], P < 0.001). These data do not support of the use of GLP-1 as an adjunctive diagnostic test for ID or IR, as defined by conventional intravenous or oral dynamic tests.

Exploring the impact of high-energy diets on cattle: Insights into subacute rumen acidosis, insulin resistance, and hoof health.

Cattle lameness remains a significant concern, causing economic losses and compromising animal welfare. Claw horn lesions have been identified as a major cause of lameness in dairy cows, but their correlation with high-energy diets and ruminal acidosis remains unclear. Hence, the primary objective of this study was to assess the effects of a high-starch diet and a conventional diet on the rumen environment, acute-phase proteins, and metabolic alterations, with a particular focus on insulin resistance and the consequent implications for the histology of the hooves in Holstein steers. A total of 16 animals were divided into the high-starch (HS; 37% starch) and conventional (CON; 16.8% starch) groups. Glucose tolerance tests (GTT), blood analyses, rumen fluid analyses, and histological evaluations of the hoof tissue were conducted over a 102-d experimental period. The HS group showed a lower ruminal pH than the CON group, and with values indicating SARA. The plasma glucose and IGF-1 concentrations were higher in the HS group, suggesting an anabolic state. Both groups exhibited an increase in the insulin area under the curve (AUC) after the GTT on d 102. Histological analysis of the hooves showed a reduction in the length and width of the epidermal lamella in both groups. We found a significant negative correlation between the insulin AUC and the length and width of the epidermal lamella. Because both groups were similarly affected, the hypothesis that histological alterations were caused by the experimental diets still needs confirmation. Additionally, the development of SARA was not essential for the observed histological changes in the hoof. Further studies are warranted to thoroughly investigate the role of insulin and IGF-1 imbalances in claw health.

Neutrophil extracellular traps and active myeloperoxidase concentrate in lamellar tissue of equids with naturally occurring laminitis.

Laminitis is a pathology of the equine digit ultimately leading to a failure of the dermo-epidermal interface. Neutrophil activation is recognized as a major factor in SIRS-associated laminitis and has recently been described in induced endocrinopathic laminitis evidenced by the presence of myeloperoxidase (MPO). Neutrophil extracellular traps (NET) are released with neutrophil activation. This study aimed to investigate the presence and activity of MPO and NET in the lamellar tissue of equids presented with naturally occurring laminitis. Samples of lamellar tissue of five horses and five donkeys presented with laminitis, as well as eight control horses without laminitis, were collected. Lamellar tissue extracts were submitted to ELISA and specific immuno-extraction followed by enzymatic detection (SIEFED) assays to confirm the presence and activity of both MPO and NET. Lamellar sections were also immunohistopathologically stained for MPO and NET. Analysis of lamellar tissue extracts revealed that laminitis cases had significantly higher levels of total MPO concentration, MPO activity, and NET-bound MPO activity in comparison to control horses. Moreover, a strong correlation was identified between the activity of NET-bound MPO and the total MPO activity, which suggests that MPO activity partly originates from NET-bound MPO. Immunohistochemical staining showed that MPO and NET labelling in laminitis cases was moderate to marked, primarily in the epidermis and in inflammatory infiltrates containing neutrophils, while labelling in control horses was minimal. This article constitutes the first indication of the presence and activity of NET-bound MPO in the lamellar tissue of horses and donkeys with naturally occurring laminitis. Targeting these substances may provide new treatment possibilities for this debilitating disease.

Retrospective assessment of the use of extended-release cabergoline in the management of equine pituitary pars intermedia dysfunction.

Introduction: Dopaminergic agonists are accepted as the most effective treatment for pituitary pars intermedia dysfunction. However, some horses are refractory to daily oral pergolide, the recommended registered treatment. Extended-release cabergoline (ERC) injection may offer an alternative. The objective of this retrospective case series was to describe clinical and endocrinological responses to ERC. Methods: Medical records of horses treated with weekly intramuscular injections of ERC (5 mg/mL, BOVA Aus) at either 0.01 mg/kg (high dose, HD) (n = 10) or 0.005 mg/kg (low dose, LD) (n = 30) were reviewed. Short-term ACTH responses were assessed at 5-8 days using a Wilcoxon signed ranked test. Longer-term ACTH responses (30 to 365 days) were assessed using generalised estimating equations. Results: Five to eight days after the first dose of LDERC, median adrenocorticotropic hormone (ACTH) concentration was lower (p = 0.001), changing from 153 pg/mL (IQR: 78, 331) to 57 pg/mL (IQR: 30, 102). With HDERC, median ACTH concentration was also 153 pg/mL (IQR: 96, 185) before and then 56 pg/mL (IQR: 29, 86) after 5-8 days of treatment (p = 0.047). Over 12 months of treatment, ACTH concentration ranged from 14 to >1,250 pg/mL (median: 51 pg/mL) in horses treated with LDERC and 20 to 472 pg/mL (median: 50 pg/mL) in horses treated with HDERC. Measurements remained above the seasonal reference range in 39.3 and 52.3% of horses treated with LDERC and HDERC, respectively. Clinical improvement was reported by owners in 78.3 and 100% of horses treated with LDERC and HDERC, respectively. Partial, self-limiting inappetence was reported in 30.0% of LDERC and 60% HDERC cases. Seven horses exhibited lethargy (5 LDERC, 2 HDERC). Insulin concentrations measured 30 days post-ERC treatment were no different from baseline. Discussion: Clinical and endocrinological responses were consistent with results of previous reports of oral pergolide treatment. Weekly injection of ERC may be an effective alternative to pergolide; the 0.005 mg/kg dose appeared to be as effective, with less risk of inappetence, than the 0.01 mg/kg dose that has been reported previously.

Effect of phenylbutazone on insulin secretion in horses with insulin dysregulation.

BACKGROUND: Phenylbutazone is often prescribed to manage pain caused by hyperinsulinemia-associated laminitis, but in diabetic people nonsteroidal anti-inflammatory drugs increase insulin secretion and pancreatic activity. HYPOTHESIS/OBJECTIVES: Investigate the effect of phenylbutazone administration on insulin secretion in horses. It was hypothesized that phenylbutazone will increase insulin secretion in horses with insulin dysregulation (ID). ANIMALS: Sixteen light breed horses, including 7 with ID. METHODS: Randomized cross-over study design. Horses underwent an oral glucose test (OGT) after 9 days of treatment with phenylbutazone (4.4 mg/kg IV q24h) or placebo (5 mL 0.9% saline). After a 10-day washout period, horses received the alternative treatment, and a second OGT was performed. Insulin and glucose responses were compared between groups (ID or controls) and treatments using paired t test and analyses of variance with P < .05 considered significant. RESULTS: In horses with ID, phenylbutazone treatment significantly decreased glucose concentration (P = .02), glucose area under the curve (2429 ± 501.5 vs 2847 ± 486.1 mmol/L × min, P = .02), insulin concentration (P = .03) and insulin area under the curve (17 710 ± 6676 vs 22 930 ± 8788 µIU/mL × min, P = .03) in response to an OGT. No significant effect was detected in control horses. CONCLUSION AND CLINICAL IMPORTANCE: Phenylbutazone administration in horses with ID decreases glucose and insulin concentrations in response to an OGT warranting further investigation of a therapeutic potential of phenylbutazone in the management of hyperinsulinemia-associated laminitis beyond analgesia.

Metabolic and Endocrine Insights in Donkeys.

Donkey medicine is gaining attention due to their increased use as companion animals, in shows, asinotherapy, etc. The increasing demand and unique aspects call for specialized care, requiring new information (physiology, infectious disorders, pharmacology, etc.). Since obesity is common in this species, hyperlipemia, metabolic syndrome and insulin dysregulation (ID) are common disorders in donkeys, in some cases with high mortality, either directly (multiorgan dysfunction) or indirectly due to poor quality of life (chronic laminitis). Donkeys have long-life expectancy and are often afflicted with pituitary pars intermedia dysfunction (PPID), a neurodegenerative and endocrine disease. Hyperlipemia is diagnosed based on high plasma triglyceride concentration in association with clinical findings and laboratory abnormalities from affected tissues (liver, kidney and pancreas). The measurement of resting serum insulin and plasma ACTH concentrations is the first step in ID and PPID diagnosis. In donkeys with clinical signs of ID (obesity or recurrent laminitis) or PPID (hypertrichosis, regional adiposity, laminitis and weight loss), where these hormones are in the normal or non-diagnostic range (donkey-specific cut-off values and reference ranges need to be established), dynamic tests are recommended (oral sugar test or thyrotropin-releasing hormone, respectively). Equine treatment protocols apply to donkeys, although pharmacological studies for most drugs, except pergolide, are lacking.

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.

Clinical implications of imprecise sampling time for 10- and 30-min thyrotropin-releasing hormone stimulation tests in horses.

BACKGROUND: The thyrotropin-releasing hormone (TRH) stimulation test is used to diagnose pituitary pars intermedia dysfunction (PPID) using 10- or 30-min protocols. Imprecise sampling time for the 10-min protocol can lead to misdiagnoses. OBJECTIVES: To determine the effect of imprecise sampling time for the 30-min protocol of the TRH stimulation test. STUDY DESIGN: In vivo experiment. METHODS: Plasma immunoreactive adrenocorticotropin (ACTH) concentrations were measured 9, 10, 11, 29, 30 and 31 min after intravenous administration of 1 mg of TRH in 15 control and 12 PPID horses. Differences in ACTH concentrations between sampling times, variability in ACTH concentrations between protocols, and diagnostic classification of PPID were assessed using Friedman's test, Bland-Altman plots, and Fisher's exact test, respectively, with 95% confidence intervals reported and significance set at p < 0.05. RESULTS: Imprecise sampling time resulted in variable ACTH concentrations, but significant differences in absolute ACTH concentrations were not detected for imprecise sampling within each protocol or between protocols. Imprecise sampling changed PPID diagnostic classification for 3/27 (11 [4-28] %) horses for both protocols. Using the 30-min protocol as a reference, 1/12 (8 [1-35] %) horses returned a negative test result and 5/12 (42 [19-68] %) horses returned equivocal test results that would be considered positive in practice due to the presence of supportive clinical signs. MAIN LIMITATIONS: Limited sample size and inter-horse variability reduced the ability to detect small but potentially relevant differences. CONCLUSIONS: Overall, the impact of imprecise sampling was not significantly different between the 10- and 30-min TRH stimulation test protocols. However, diagnostic classification for PPID would have varied between the 10- and 30-min protocols in this population, if clinical signs had been ignored. Precise timing during TRH stimulation tests and contextual interpretation of ACTH concentrations remain fundamental for the diagnosis of PPID.

Factors associated with insulin responses to oral sugars in a mixed-breed cohort of ponies.

BACKGROUND: Serum insulin concentration at 60 min (InsulinT60) during an oral sugar test (OST) indicates future laminitis risk and insulin dysregulation (ID). Associations between InsulinT60 and physical and owner-reported variables may help clinicians select individuals for ID testing. Associations between InsulinT60 and other metabolic markers may help elucidate ID pathophysiology. OBJECTIVES: To describe associations between (A) season, physically-apparent and owner-reported factors and binary InsulinT60 interpretation (initial models) and (B) variables included in the initial models, other metabolic markers and continuous InsulinT60 (full models). STUDY DESIGN: Prospective longitudinal. METHODS: Non-laminitic ponies were examined and OSTs (0.3 mL/kg Karo syrup) performed every 6 months (autumn and spring) for ≤4 years. Factors associated with InsulinT60 were determined using mixed-effects models with physical, owner-reported, season and serum/plasma markers as fixed effects and pony and premises identifiers as random effects. Autumn and spring data were analysed separately for full models. RESULTS: One thousand seven hundred and sixty-three OSTs from 367 ponies were included. High-risk InsulinT60 (>153 µIU/mL) was independently associated with (odds ratio, 95% confidence interval [CI]): age (1.36, 1.16-1.60), body condition score (BCS) (2.38, 1.21-4.69), and bulging supraorbital fatpads (7.25, 2.1-24.98). However, the initial models provided little explanatory power (Nakagawa R2 = 0.1-0.27). LoginsulinT60 was independently associated with (effect estimate, 95% CI): age (0.02, 0.01-0.04), Welsh/Welsh X breed (0.22, 0.05, 0.39), sex (gelding = -0.2, -0.34 to 0.06), BCS (0.16, 0.08-0.23), plasma adiponectin (-0.02, -0.02 to 0.01) and basal insulin (0.01, 0.01-0.01) in spring, and: age (0.03, 0.02-0.04), BCS (0.17, 0.08-0.26), bulging supraorbital fatpads (0.37, 0.2-0.54), turnout score (0.05, 0.02-0.09), plasma adiponectin (-0.01, -0.02 to 0.01), ACTH (per 10 pg/mL) (0.01, 0.00-0.01), triglycerides (0.28, 0.07-0.49) and InsulinT0 (0.01, 0.01-0.01) in autumn. MAIN LIMITATIONS: Only non-laminitic ponies in one region were included. CONCLUSIONS: Owner-reported and physically-apparent data were limited InsulinT60 predictors. InsulinT60 is a complex trait, independently associated with numerous variables, some with seasonal interactions.

Intra-articular triamcinolone acetonide injection results in increases in systemic insulin and glucose concentrations in horses without insulin dysregulation.

BACKGROUND: Corticosteroids are a commonly used, inexpensive intra-articular treatment for osteoarthritis which may increase the risk for laminitis in horses due, in part, to hyperinsulinaemia. Humans with metabolic syndrome experience increases in insulin and glucose concentrations post-injection, but responses in horses are unknown. OBJECTIVES: To determine the effect of a single intra-articular (IA) dose of triamcinolone acetate (TA) on blood insulin and glucose concentrations. STUDY DESIGN: Before-after study. METHODS: Ten horses with normal insulin regulation as assessed by an oral sugar test received 18 mg of TA into one middle carpal joint. Insulin and glucose concentrations were evaluated at baseline and 4, 6, 8, 24, 48, and 72 h following IA corticosteroid injection. Differences from baseline were evaluated using a repeated measures ANOVA with Dunnett's multiple comparison testing or a Friedman test with Dunn's correction (significant at p < 0.05). RESULTS: Mean ± SD blood insulin concentration post IA TA injection was increased at 6 h (15.8 ± 3.1 µIU/mL, p = 0.01), 24 h (23 ± 5.8 µIU/mL, p ≤ 0.001), and 48 h (29 ± 13 µIU/mL, p ≤ 0.01) compared to baseline (10 ± 12.3 µIU/mL), with the peak at 48 h. Median ± 95% CI blood glucose concentration post IA TA injection was increased at 6 h (112.7 ± 20.3 mg/dL, p = 0.006), 8 h (112.9 ± 21.4 mg/dL, p = 0.004), 24 h (122.6 ± 14.6, p ≤ 0.0001), and 48 h (123.5 ± 15.4 mg/dL, p ≤ 0.0001) compared to baseline (89.2 ± 6.6 mg/dL), with the peak at 48 h. MAIN LIMITATIONS: Only horses with normal insulin regulation were evaluated. CONCLUSIONS: Blood insulin and glucose concentrations modestly increased for 48 h following IA TA.

Associations between feeding and glucagon-like peptide-2 in healthy ponies.

BACKGROUND: Gastrointestinal peptides, such as glucagon-like peptide-2 (GLP-2), could play a direct role in the development of equine hyperinsulinaemia. OBJECTIVES: To describe the secretory pattern of endogenous GLP-2 over 24 h in healthy ponies and determine whether oral administration of a synthetic GLP-2 peptide increases blood glucose or insulin responses to feeding. STUDY DESIGN: A cohort study followed by a randomised, controlled, cross-over study. METHODS: In the cohort study, blood samples were collected every 2 h for 24 h in seven healthy ponies and plasma [GLP-2] was measured. In the cross-over study, 75 µg/kg bodyweight of synthetic GLP-2, or carrier only, was orally administered to 10 ponies twice daily for 10 days. The area under the curve (AUC0-3h ) of post-prandial blood glucose and insulin were determined before and after each treatment. RESULTS: Endogenous [GLP-2] ranged from <0.55 to 1.95 ± 0.29 [CI 0.27] ng/mL with similar peak concentrations in response to meals containing 88-180 g of non-structural carbohydrate, that were ~4-fold higher (P < 0.001) than the overnight nadir. After GLP-2 treatment peak plasma [GLP-2] increased from 1.1 [0.63-1.37] ng/mL to 1.54 [1.1-2.31] ng/mL (28.6%; P = 0.002), and AUC0-3h was larger (P = 0.01) than before treatment. The peptide decreased (7%; P = 0.003) peak blood glucose responses to feeding from 5.33 ± 0.45 mmol/L to 5.0 ± 0.21 mmol/L, but not AUC0-3h (P = 0.07). There was no effect on insulin secretion. MAIN LIMITATIONS: The study only included healthy ponies and administration of a single dose of GLP-2. CONCLUSIONS: The diurnal pattern of GLP-2 secretion in ponies was similar to other species with no apparent effect of daylight. Although GLP-2 treatment did not increase post-prandial glucose or insulin responses to eating, studies using alternative dosing strategies for GLP-2 are required.

Effect of 5'-adenosine monophosphate-activated protein kinase agonists on insulin and glucose dynamics in experimentally induced insulin dysregulation in horses.

BACKGROUND: 5'-adenosine monophosphate-activated protein kinase (AMPK) agonists, particularly resveratrol (RES), have not been extensively evaluated for their effect on insulin dysregulation (ID) in horses. OBJECTIVES: Evaluate the effects of treatment with RES (10 mg/kg PO q12h), metformin (MET; 30 mg/kg PO q12h), and aspirin (ASP; 20 mg/kg PO q24h) on experimentally induced ID. ANIMALS: Thirty-three healthy, adult, light-breed horses. METHODS: Unblinded, placebo-controlled, experimental trial evaluating effects of AMPK agonists (RES, MET, and ASP) on experimentally induced ID. Horses were randomly assigned to a treatment group (RES, MET/ASP, RES/ASP, RES/MET/ASP, or placebo [CON]) after induction of ID with dexamethasone (0.08 mg/kg PO q24h for 7 days). Frequently sampled insulin-modified IV glucose tolerance tests (FSIGTT) and oral sugar tests (OST) were performed at baseline, 7 days after ID, and ID plus 7 days of treatment. Minimal model and OST variables were compared between (1-way ANOVA) and within (1-way ANOVA for repeated measures) groups over time to determine effects of treatment on ID. RESULTS: Administration of dexamethasone for 14 days resulted in significantly altered insulin and glucose dynamics (SI, DI, basal [glucose], and [insulin]) and produced clinical signs of laminitis in 5 out of 33 (15%) of horses included in the study. Combination therapy with RES, MET, and ASP did not significantly improve insulin and glucose dynamics in horses with experimentally induced ID. CONCLUSIONS AND CLINICAL IMPORTANCE: Metabolic testing before glucocorticoid administration should be considered in horses with clinical signs of metabolic syndrome.

Prolonged hyperinsulinemia increases the production of inflammatory cytokines in equine digital lamellae but not in striated muscle.

Hyperinsulinemia is the key feature of equine metabolic syndrome (EMS) which leads to debilitating sequelae. Hyperinsulinemia-associated laminitis (HAL) is one of the major sequelae of EMS, although the pathophysiological mechanisms are not well elucidated. Using an equine model, we hypothesized that expression of inflammatory markers would be increased in digital lamellae and striated muscle following prolonged hyperinsulinemia. Healthy Standardbred horses (5.4 ± 1.9 years) were alternately assigned to a prolonged euglycemic-hyperinsulinemic clamp (pEHC) or control group (n = 4 per group). Following a 48 h pEHC or a 48 h infusion of a balanced electrolyte solution (controls), biopsies were collected from digital lamellar tissue, skeletal muscle and cardiac muscle were obtained. All hyperinsulinemic horses developed laminitis regardless of previous health status at enrollment. Protein expression was quantified via Western blotting. A significant (P < 0.05) upregulation of the protein expression of heat shock protein 90 (HSP90), alpha 2 macroglobulin (A2M) and fibrinogen (α, ß isoforms), as well as inflammatory cytokines including interleukin-1ß were detected in digital lamellae following prolonged hyperinsulinemia. In contrast, protein expression of cytokines and acute phase proteins in heart and skeletal muscle was unchanged following hyperinsulinemia. Upregulation of inflammatory cytokines and acute phase proteins in digital lamellae during prolonged hyperinsulinemia may reveal potential biomarkers and novel therapeutic targets for equine endocrinopathic laminitis. Further, the lack of increase of inflammatory proteins and acute phase proteins in striated muscle following prolonged hyperinsulinemia may highlight potential anti-inflammatory and cardioprotective mechanisms in these insulin-sensitive tissues.

Insulin, but not adiponectin, is detectable in equine saliva using an automated, commercial assay.

BACKGROUND: The hormones insulin and adiponectin are commonly measured in equids because of their involvement in endocrinopathic laminitis. These are currently measured in serum/plasma, although jugular venipuncture can cause stress in some animals and may be impossible in needle-shy individuals. However, both hormones can be measured in saliva in other species. OBJECTIVES: To determine whether [insulin] and [total adiponectin] are detectable in equine saliva using automated assays and whether saliva collection is associated with changes in stress indicators. Additionally, the correlation between serum and salivary [insulin] was investigated. STUDY DESIGN: In vivo experiment. METHODS: Paired blood and saliva samples were collected from eight adult ponies at multiple time-points (n = 45 paired samples). [Insulin] and [total adiponectin] were measured using automated assays validated for equine serum/plasma. Blink rates and heart rates were determined, using video recordings and a wearable heart rate monitor respectively, to assess the effects of sample collection on stress indicators compared with a control situation without a stressful stimulus. RESULTS: [Total adiponectin] was undetectable in saliva. However, salivary [insulin] was measurable with acceptable inter-assay (1.3 ± 0.9%) and intra-assay (1.1 ± 0.6%) variability. Blink and heart rates during saliva collection did not differ significantly from those in the control setting. Serum and salivary [insulin] were not significantly correlated. MAIN LIMITATIONS: Small sample size comprising native UK ponies; potential sampling bias as animals were recruited to the study partly based on their behaviour during blood sampling; saliva collected from unfasted animals. CONCLUSIONS: Insulin is measurable in equine saliva using an automated assay currently available in the UK, but further validation and the determination of specific diagnostic thresholds are required. Saliva collection was not associated with changes in stress indicators. Further research is therefore needed to determine the potential of equine saliva as a non-invasive alternative to blood for insulin determination.

Radiological features of arterial channels in the equine third phalanx measured using a novel customized software represent changes of laminitis.

OBJECTIVE: To identify and measure radiolucencies at the solear margin of the distal phalanx in radiographs of healthy and laminitic hooves. SAMPLE: Clinical records and dorsoproximal-palmarodistal radiographs of equine forelimbs with radiological diagnoses of either laminitis (n = 40, L) or navicular syndrome (n = 40, NS). METHODS: Outlines of the radiolucent structures at the solar margin were drawn in ImageJ, and a customized novel plugin "Arteries Analyzer/ImageJ" was used for measurements. The diverging radiolucencies outside the terminal arc of the distal phalanx were differentiated as arterial channels (open at the solear margin) and ellipses (closed at the solear margin). Comparisons between L and NS, between distal phalanges with and without ellipses, and of arterial channels and ellipses in areas were compared using Wilcoxon and the Mann-Whitney U tests, respectively. The reliability and repeatability of the method were tested using Friedman's test. RESULTS: Fewer arterial channels but more ellipses were identified in L than in NS. In phalanges with ellipses (n = 47), the number of ellipses and the number of arterial channels were negatively correlated (PCC -0.181, P = .224). The number of ellipses correlated positively with the severity of laminitis (PCC 0.495, P < .001; n = 80) and with the degree of rotation of the distal phalanx (PCC 0.392, P < .001; n = 80). CLINICAL RELEVANCE: The software tool successfully measured arterial channels and ellipses outlined by the evaluators. Results indicate that healthy arteries develop into pathological ellipses in laminitic feet. This may be used to complement the interpretation of radiographs and support clinical decision-making.

Radiographic analysis of the dorsal hoof wall thickness in clinically normal draft horses.

OBJECTIVE: To evaluate the radiographic thickness of the dorsal hoof wall in normal draft horse feet. ANIMALS: 33 adult draft horses with no history of laminitis, no clinically obvious lameness, and visibly unremarkable front feet were included. METHODS: This was a prospective, descriptive study of clinically normal draft horses' front feet. Lateromedial radiographs were acquired of the front feet. A ratio of the dorsal hoof wall thickness to the length of the distal phalanx (DHWP3 ratio) was calculated. RESULTS: The dorsal hoof wall thickness to length of the distal phalanx was calculated as 0.33 ± 0.03 (range of 0.28 to 0.39) in this population of draft horses. CLINICAL RELEVANCE: With very few exceptions, the heterogeneous population of draft horses evaluated in this study had a DHWP3 ratio greater than previously published values in lighter breeds (< 0.30).