Andy E. Durham’s research while affiliated with Texas Equine Hospital and other places

The measurement of the blood insulin concentration, and comparison to cut-offs, is essential in diagnosing insulin dysregulation, a common equine endocrinopathy. However, different insulin assays provide disparate results. We aimed to ease comparison between assays by compiling original and published data into a web app to convert insulin measurements from one assay to another. Data were available for ADVIA Centaur insulin chemiluminescent immunoassay (CLIA), Beckman Coulter insulin radioimmunoassay (RIA), Immulite 1000 CLIA, Immulite 2000 CLIA, Immulite 2000 XPi CLIA, Mercodia equine insulin enzyme-linked immunosorbent assay (ELISA), and Millipore porcine insulin RIA. Linear models were fitted for 13 assay pairs using non-decreasing splines, and integrated into this app. Assay comparisons including data from several studies showed a lower performance. This indicates technical variation between laboratories, which has not been described before, but is relevant when diagnostic measurements and cut-offs are provided by different laboratories. Nevertheless, the models’ overall high performance (median r2 = 0.94; range 0.57–1.00) supports their use to interpret results from diagnostic insulin measurements when the reference assay is unavailable, and to compare values obtained from different assays.

Background High concentrations of adrenocorticotropic hormone (ACTH) are used as an indicator of pituitary pars intermedia dysfunction (PPID), but other factors that may influence ACTH need to be understood, if diagnostic reference ranges for ACTH are to be used with confidence. Insulin dysregulation (ID) could be one such factor, as insulin affects pituitary hormones in other species. Objectives To test the hypothesis that a relationship exists between high insulin and high ACTH in aged (>15‐year‐old) animals with no clinical signs of PPID. Study design A cohort study. Methods Thirteen horses and eleven ponies (17–25 years‐old; mares and geldings) were clinically examined for signs of PPID in the spring (November 2020) and autumn (April 2021). On the same day, blood samples were taken before and 2 h after an oral glucose test (OGT). Concentrations of insulin, glucose, ACTH and cortisol were measured. Results There was no association between ACTH and cortisol. However, there was a positive linear correlation between ACTH and post‐OGT (insulin in the autumn (r = 0.427, p = 0.04). Two horses and six ponies had ACTH above the cut‐off value for PPID diagnosis, and of these eight animals, six also had insulin concentrations above the cut‐off value for ID. Main limitations The cohort was small and thyrotropin‐releasing hormone (TRH) stimulation tests were not performed. Conclusions In autumn, high ACTH was associated with ID, when no clinical signs of PPID were present. Because ACTH is used in PPID diagnosis, further work is required to understand this interaction.

Background A glycemic challenge test is used for the diagnosis of insulin dysregulation (ID) in horses and ponies. Different forms of the test exist where the administrative route and dose of glucose vary, which makes interpretation of results challenging. Hypothesis/Objectives To evaluate the palatability of, and blood glucose and insulin responses to, carbohydrate pellets fed as an oral glucose test (OGT), and to establish the diagnostic threshold for ID when using the pellets. Animals University and privately‐owned horses and ponies (n = 157) comprised of 31 breeds and both sexes. Methods Multicenter cohort study. A custom‐produced glycemic pellet was offered for free intake at 0.5 g/kg BW soluble carbohydrate and serum insulin and blood glucose concentrations measured before and after (60, 120, and 180 minutes) the pellets were offered. Pellet acceptance and intake time (those that finished within 10 minutes) were determined to assess palatability. Results The pellets were palatable to 132/157 animals, and ponies found the pellets more (P = .004) palatable than horses. The median intake time (4 [3‐6] minutes) was positively correlated with acceptance grade (r = .51; P < .0001). Consumption of the pellets elicited peak blood glucose (6.6 [5.8‐7.8] mmol/L) and serum insulin (40.5 [19‐99.8] μIU/mL) responses at 120 minutes. At 120 minutes the optimal cut‐off was 83 μIU/mL (95% CI: 70‐99 μIU/mL) for the IMMULITE 2000XPi assay. Conclusions and Clinical Importance The pellets were palatable and a suitable, novel carbohydrate source for the OGT.

Background Outbreaks of liver disease in horses are common but the etiology of most remains unknown. Forage mycotoxins have been suspected to be a cause. Objectives To examine the association between outbreaks of liver disease and the presence of mycotoxins in forage stored on the same premises. Animals Premises were identified where ≥4 horses were contemporaneously affected by liver disease, and a control group was formed from premises where ≥4 horses had been examined and found to have no evidence of liver disease. Methods Forage was collected from 29 case and 12 control premises. The forage was analyzed for mycotoxin content using a liquid chromatography/mass spectrometry method, targeting 54 mycotoxins. The presence and distribution of mycotoxins between case and control samples was compared. Results Mycotoxins were found in 23/29 (79%) case samples and 10/12 (83%) control samples (P > .99; relative risk, 0.93; 95% confidence interval [CI], 0.64‐1.75). Median (interquartile range [IQR]) total mycotoxin concentration was similar in case and control samples (85.8 μg/kg [1.6‐268] vs. 315 μg/kg [6.3‐860]; P = .16). Ten mycotoxins were found exclusively in case premises comprising fumonisin B1, 15‐acetyldeoxynivalenol, deoxynivalenol, zearalenone, aflatoxins B1 and G1, methylergonovine, nivalenol, verruculogen, and wortmannin. The median (IQR) concentration of fumonisin B1 was significantly higher in case versus control samples (0 μg/kg [0‐81.7] vs. 0 μg/kg [0‐0]; P = .04). Conclusions and Clinical Importance Several mycotoxins with known hepatotoxic potential were found, alone or in combination, exclusively at case premises, consistent with the hypothesis that forage‐associated mycotoxicosis may be a cause of outbreaks of liver disease in horses in the United Kingdom.

Use of plasma adrenocorticotropic hormone (ACTH) concentrations for the diagnosis of PPID in equids requires awareness of other factors that might influence ACTH concentrations, including breed and time of year. This study was designed to investigate effects of breed on plasma ACTH concentrations and potential interactions between breed and time of year. Ten breeds were selected from a laboratory database containing data on plasma ACTH concentrations. Breeds represented were Arabians, Cobs, Connemara ponies, donkeys, Irish Sports horses, New Forest ponies, Shetland ponies, Thoroughbreds, Warmbloods and Welsh breeds. Multivariable analysis was used to determine the effects of month and breed on plasma ACTH concentrations. A circannual pattern of ACTH concentration was documented, with a nadir in April and a peak in September. Arabian horses and donkeys had significantly higher ACTH concentrations than some other breeds, primarily from May to November, whereas in Shetland ponies and Welsh breeds, relatively higher ACTH concentrations only occurred from July to November. The annual increase in plasma ACTH appeared to commence in late April/early May in response to increasing daylight and decreased again promptly after the late September equinox as daylength fell below 12 h per day. Inter-breed differences in plasma ACTH were substantial and indicated that breed and time of year should be considered when interpreting plasma ACTH concentrations in equids suspected for PPID.

Thyrotropin releasing hormone (TRH) stimulation testing is often used to support a diagnosis of pituitary pars intermedia dysfunction (PPID) in horses although it is unclear whether or not repeat TRH stimulation testing post-treatment is a valid means of assessing response to medical therapy. Laboratory submissions from 64 suspected equine PPID cases were examined including the initial pre-treatment TRH stimulation test and a follow up test within 100 days of starting medical therapy with pergolide. In a subset of cases, further follow-up tests were examined beyond 100 days of starting treatment. Results from tests conducted between 1 July and 30 November 30 were excluded. Significant improvements were seen in both the baseline and TRH-stimulated adrenocorticotrophic hormone (ACTH) concentrations within 100 days with no further improvements seen in the subset of cases examined thereafter. Although 88% (n=56/64) of all cases showed a decreased response to TRH post-treatment, only 24% (n=9/38) of horses with positive pre-treatment TRH stimulation tests normalised following treatment, with a further 34% (n=13/38) improving into an equivocal test outcome category. Most commonly (42%; n=16/38), horses with positive pre-treatment TRH stimulation tests remained positive following treatment, although 75% (n=12/16) of these showed a numerically lower post-treatment response to TRH. These results will help inform practitioners of expected changes in TRH stimulation test results when assessing response of horses with PPID to medical therapy with pergolide.