FREE SHIPPING over $40
OVER 10000 FIVE-STAR REVIEWS
5% OFF ALL SUBSCRIPTIONS
100% HAPPINESS GUARANTEE
Analyze Diet
0
Journal of veterinary internal medicine2022; 36(2); 770-777; doi: 10.1111/jvim.16362

Effect of early or late blood sampling on thyrotropin releasing hormone stimulation test results in horses.

Abstract: Diagnosis of pituitary pars intermedia dysfunction (PPID) using the thyrotropin-releasing hormone (TRH) stimulation test requires blood collection 10 minutes after TRH injection; it is unknown if small differences in timing affect test results. Objective: To determine whether early or late sampling results in a significant (≥10%) difference in plasma adrenocorticotropic hormone (ACTH) concentration compared to standard 10-minute sampling. Methods: Twenty-four healthy adult horses with unknown PPID status. Methods: In this prospective study, subjects underwent a single TRH stimulation test, with blood collected exactly 9 minutes (early), 10 minutes (standard), and 11 minutes (late) after injection. ACTH was measured by chemiluminescent immunoassay. Two aliquots of the 10-minute plasma sample were analyzed separately to assess intra-assay variability. Data were reported descriptively and bias was calculated using Bland-Altman plots. Significance was set at P = .05. Results: Minor variability was observed between the paired 10-minute sample aliquots (range, 0%-6%; median 3%). Overall variability of early or late samples compared to the corresponding paired (average) 10-minute standard concentration ranged from 0% to 92% (median 10%). Seventy-five percent of horses (18/24) tested had at least 1 early or late reading that differed by ≥10% from its corresponding 10-minute standard concentration, and 21% of horses (5/24) would have a different interpretation of testing result with either early or late sampling. Incidence of ≥10% variability was independent of PPID status (P = .59). Conclusions: Precise timing of sample collection is critical to ensure accurate assessment of PPID status given the observation of significant variability associated with minor alterations in timing of sample collection.
© 2022 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals LLC. on behalf of the American College of Veterinary Internal Medicine.
Get Full Text
Publication Date: 2022-01-20 PubMed ID: 35049089PubMed Central: PMC8965261DOI: 10.1111/jvim.16362Google Scholar: Lookup
The Equine Research Bank provides access to a large database of publicly available scientific literature. Inclusion in the Research Bank does not imply endorsement of study methods or findings by Mad Barn.
LinkedInCopy
  • Journal Article

Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

This research studied the impact of small timing differences in blood sampling during thyrotropin-releasing hormone (TRH) stimulation test on the diagnosis of pituitary pars intermedia dysfunction (PPID) in horses. The findings suggest that precision in blood sampling timing is essential since minor departures from standard timing can significantly influence the plasma adrenocorticotropic hormone (ACTH) concentration and thus, interpretation of test results.

Objective and Methodology

  • The researchers aimed to ascertain if early or late blood sampling led to significant differences (≥10%) in plasma ACTH concentration compared to the 10-minute standard blood collection after TRH injection which is a clinical norm for diagnosing PPID in horses.
  • Twenty-four healthy adult horses of unknown PPID status were included in this study. They underwent a single TRH stimulation test where blood was collected exactly at 9 minutes (early), 10 minutes (standard), and 11 minutes (late) after the TRH injection. The ACTH levels in the blood were then measured using chemiluminescent immunoassay.
  • This prospective study also analyzed two aliquots of the 10-minute plasma sample separately to determine intra-assay variability. The data bias was addressed using statistical methods with set significance at P = .05.

Results

  • The research observed minor variability (0%-6%, median 3%) between the paired 10-minute sample aliquots, showing reliable intra-assay consistency.
  • The variability in early or late samples compared to the standard 10-minute blood sample ranged from 0% to 92% (median 10%). It implies that minor changes in the timing of blood collection could lead to high variability in ACTH concentration measured.
  • Out of the horses tested, 75% had at least one early or late reading differing by ≥10% from its corresponding 10-minute standard concentration. It would significantly alter the interpretation of test results in 21% of horses.
  • The researchers concluded that ≥10% variability was independent of the horse’s PPID status since the statistical correlation was insignificant (P = .59).

Conclusion

  • The research concluded that precise timing of blood sample collection is crucial for an accurate assessment of PPID status in horses.
  • Even minor alterations in the timing of sample collection led to significant variability in ACTH concentration, thereby possibly affecting the test result interpretation and diagnostic accuracy for PPID.

Cite This Article

APA
Thane K, Uricchio C, Frank N. (2022). Effect of early or late blood sampling on thyrotropin releasing hormone stimulation test results in horses. J Vet Intern Med, 36(2), 770-777. https://doi.org/10.1111/jvim.16362

Publication

Journal of veterinary internal medicine
ISSN: 1939-1676
NlmUniqueID: 8708660
Country: United States
Language: English
Volume: 36
Issue: 2
Pages: 770-777

Researcher Affiliations

Thane, Kristen
  • Tufts Cummings School of Veterinary Medicine, Department of Comparative Pathobiology, North Grafton, Massachusetts, USA.
Uricchio, Cassandra
  • University of Massachusetts, Department of Veterinary and Animal Sciences, Amherst, Massachusetts, USA.
Frank, Nicholas
  • Tufts Cummings School of Veterinary Medicine, Department of Comparative Pathobiology, North Grafton, Massachusetts, USA.

MeSH Terms

  • Animals
  • Horse Diseases
  • Horses
  • Pituitary Diseases / diagnosis
  • Pituitary Diseases / veterinary
  • Pituitary Gland, Intermediate
  • Prospective Studies
  • Thyrotropin-Releasing Hormone

Grant Funding

  • PR0481 / Boehringer Ingelheim Vetmedica, Inc

Conflict of Interest Statement

Dr Frank consults on study design and the diagnosis and treatment of pituitary pars intermedia dysfunction in horses for Boehringer Ingelheim, which provided funds to support this study.

References

This article includes 38 references
  1. Spelta CW. Equine pituitary pars intermedia dysfunction: current perspectives on diagnosis and management.. Vet Med (Auckl) 2015;6:293-300.
    pmc: PMC6067528pubmed: 30101114doi: 10.2147/vmrr.s74191google scholar: lookup
  2. Schott HC 2nd. Pituitary pars intermedia dysfunction: equine Cushing's disease.. Vet Clin North Am Equine Pract 2002 Aug;18(2):237-70.
    pubmed: 15635907doi: 10.1016/s0749-0739(02)00018-4google scholar: lookup
  3. McGowan TW, Pinchbeck GP, McGowan CM. Prevalence, risk factors and clinical signs predictive for equine pituitary pars intermedia dysfunction in aged horses.. Equine Vet J 2013 Jan;45(1):74-9.
    pubmed: 22594955doi: 10.1111/j.2042-3306.2012.00578.xgoogle scholar: lookup
  4. Horn R, Bamford NJ, Afonso T, Sutherland M, Buckerfield J, Tan RHH, Secombe CJ, Stewart AJ, Bertin FR. Factors associated with survival, laminitis and insulin dysregulation in horses diagnosed with equine pituitary pars intermedia dysfunction.. Equine Vet J 2019 Jul;51(4):440-445.
    pubmed: 30417404doi: 10.1111/evj.13041google scholar: lookup
  5. Frank N, Andrews FM, Sommardahl CS, Eiler H, Rohrbach BW, Donnell RL. Evaluation of the combined dexamethasone suppression/ thyrotropin-releasing hormone stimulation test for detection of pars intermedia pituitary adenomas in horses.. J Vet Intern Med 2006 Jul-Aug;20(4):987-93.
    pubmed: 16955827doi: 10.1892/0891-6640(2006)20[987:eotcdt]2.0.co;2google scholar: lookup
  6. Goodale L, Frank N, Hermida P, D'Oench S. Evaluation of a thyrotropin-releasing hormone solution stored at room temperature for pituitary pars intermedia dysfunction testing in horses.. Am J Vet Res 2015 May;76(5):437-44.
    pubmed: 25909376doi: 10.2460/ajvr.76.5.437google scholar: lookup
  7. Hodge E, Kowalski A, Torcivia C, Lindborg S, Stefanovski D, Hart K, Frank N, van Eps A. Effect of thyrotropin-releasing hormone stimulation testing on the oral sugar test in horses when performed as a combined protocol.. J Vet Intern Med 2019 Sep;33(5):2272-2279.
    pmc: PMC6766522pubmed: 31432575doi: 10.1111/jvim.15601google scholar: lookup
  8. Horn R, Bertin FR. Evaluation of combined testing to simultaneously diagnose pituitary pars intermedia dysfunction and insulin dysregulation in horses.. J Vet Intern Med 2019 Sep;33(5):2249-2256.
    pmc: PMC6766519pubmed: 31498947doi: 10.1111/jvim.15617google scholar: lookup
  9. Beech J, Boston R, Lindborg S. Comparison of cortisol and ACTH responses after administration of thyrotropin releasing hormone in normal horses and those with pituitary pars intermedia dysfunction.. J Vet Intern Med 2011 Nov-Dec;25(6):1431-8.
    pubmed: 22092639doi: 10.1111/j.1939-1676.2011.00810.xgoogle scholar: lookup
  10. Horn R, Stewart AJ, Jackson KV, Dryburgh EL, Medina-Torres CE, Bertin FR. Clinical implications of using adrenocorticotropic hormone diagnostic cutoffs or reference intervals to diagnose pituitary pars intermedia dysfunction in mature horses.. J Vet Intern Med 2021 Jan;35(1):560-570.
    pmc: PMC7848300pubmed: 33368633doi: 10.1111/jvim.16017google scholar: lookup
  11. Schottn HC, Andrews FM, Durham AE. Recommendations for the Diagnosis and Treatment of Pituitary Pars Intermedia Dysfunction (PPID). .
  12. McFarlane D, Beech J, Cribb A. Alpha-melanocyte stimulating hormone release in response to thyrotropin releasing hormone in healthy horses, horses with pituitary pars intermedia dysfunction and equine pars intermedia explants.. Domest Anim Endocrinol 2006 May;30(4):276-88.
    pubmed: 16115743doi: 10.1016/j.domaniend.2005.07.005google scholar: lookup
  13. Beech J, Boston R, Lindborg S, Russell GE. Adrenocorticotropin concentration following administration of thyrotropin-releasing hormone in healthy horses and those with pituitary pars intermedia dysfunction and pituitary gland hyperplasia.. J Am Vet Med Assoc 2007 Aug 1;231(3):417-26.
    pubmed: 17669045doi: 10.2460/javma.231.3.417google scholar: lookup
  14. Beech J, McFarlane D, Lindborg S, Sojka JE, Boston RC. α-Melanocyte--stimulating hormone and adrenocorticotropin concentrations in response to thyrotropin-releasing hormone and comparison with adrenocorticotropin concentration after domperidone administration in healthy horses and horses with pituitary pars intermedia dysfunction.. J Am Vet Med Assoc 2011 May 15;238(10):1305-15.
    pubmed: 21568777doi: 10.2460/javma.238.10.1305google scholar: lookup
  15. Funk RA, Stewart AJ, Wooldridge AA, Kwessi E, Kemppainen RJ, Behrend EN, Zhong Q, Johnson AK. Seasonal changes in plasma adrenocorticotropic hormone and α-melanocyte-stimulating hormone in response to thyrotropin-releasing hormone in normal, aged horses.. J Vet Intern Med 2011 May-Jun;25(3):579-85.
    pubmed: 21457320doi: 10.1111/j.1939-1676.2011.0712.xgoogle scholar: lookup
  16. Haffner JC, Neal DL, Hoffman RM, Grubbs ST. Plasma adrenocorticotropic hormone concentration in horses decreases after freezing for 60 days.. J Vet Diagn Invest 2019 Nov;31(6):856-858.
    pmc: PMC6900716pubmed: 31646941doi: 10.1177/1040638719882374google scholar: lookup
  17. Perkins GA, Lamb S, Erb HN, Schanbacher B, Nydam DV, Divers TJ. Plasma adrenocorticotropin (ACTH) concentrations and clinical response in horses treated for equine Cushing's disease with cyproheptadine or pergolide.. Equine Vet J 2002 Nov;34(7):679-85.
    pubmed: 12455838doi: 10.2746/042516402776250333google scholar: lookup
  18. Rendle DI, Duz M, Beech J, Parkin T, Durham AE. Investigation of single and paired measurements of adrenocorticotropic hormone for the diagnosis of pituitary pars intermedia dysfunction in horses.. J Vet Intern Med 2015 Jan;29(1):355-61.
    pmc: PMC4858100pubmed: 25312676doi: 10.1111/jvim.12489google scholar: lookup
  19. Rendle DI. Laboratory diagnosis of the endocrine causes of laminitis. Livestock 2017;22(4):216‐220.
    doi: 10.12968/live.2017.22.4.216google scholar: lookup
  20. Beech J, Boston RC, McFarlane D, Lindborg S. Evaluation of plasma ACTH, alpha-melanocyte-stimulating hormone, and insulin concentrations during various photoperiods in clinically normal horses and ponies and those with pituitary pars intermedia dysfunction.. J Am Vet Med Assoc 2009 Sep 15;235(6):715-22.
    pubmed: 19751169doi: 10.2460/javma.235.6.715google scholar: lookup
  21. Rendle DI, Litchfield E, Heller J, Hughes KJ. Investigation of rhythms of secretion and repeatability of plasma adrenocorticotropic hormone concentrations in healthy horses and horses with pituitary pars intermedia dysfunction.. Equine Vet J 2014 Jan;46(1):113-7.
    pubmed: 23742059doi: 10.1111/evj.12114google scholar: lookup
  22. Shepard KN, Haffner JC, Neal DL, Grubbs ST, Pearce GL. Effect of delayed plasma centrifugation on equine adrenocorticotropic hormone concentration.. J Vet Diagn Invest 2019 Jul;31(4):585-587.
    pmc: PMC6857008pubmed: 31246158doi: 10.1177/1040638719860877google scholar: lookup
  23. Rendle DI, Litchfield E, Gough S, Cowling A, Hughes KJ. The effects of sample handling and N-phenylmaleimide on concentration of adrenocorticotrophic hormone in equine plasma.. Equine Vet J 2015 Sep;47(5):587-91.
    pubmed: 24980684doi: 10.1111/evj.12319google scholar: lookup
  24. Irvine KL, Burt K, Hill AJ, Shaw S, Papasouliotis K. Initial analytic quality assessment and method comparison of an immunoassay for adrenocorticotropic hormone measurement in equine samples.. Vet Clin Pathol 2016 Mar;45(1):154-63.
    pubmed: 26756538doi: 10.1111/vcp.12326google scholar: lookup
  25. Banse HE, Schultz N, McCue M, Geor R, McFarlane D. Comparison of two methods for measurement of equine adrenocorticotropin.. J Vet Diagn Invest 2018 Mar;30(2):233-237.
    pmc: PMC6505866pubmed: 29284383doi: 10.1177/1040638717752216google scholar: lookup
  26. McGilvray TA, Knowles EJ, Harris PA, Menzies-Gow NJ. Comparison of immunofluorescence and chemiluminescence assays for measuring ACTH in equine plasma.. Equine Vet J 2020 Sep;52(5):709-714.
    pubmed: 31955443doi: 10.1111/evj.13227google scholar: lookup
  27. Knowles EJ, Moreton-Clack MC, Shaw S, Harris PA, Elliott J, Menzies-Gow NJ. Plasma adrenocorticotropic hormone (ACTH) concentrations in ponies measured by two different assays suggests seasonal cross-reactivity or interference.. Equine Vet J 2018 Sep;50(5):672-677.
    pubmed: 29247453doi: 10.1111/evj.12797google scholar: lookup
  28. Diez de Castro E, Lopez I, Cortes B, Pineda C, Garfia B, Aguilera-Tejero E. Influence of feeding status, time of the day, and season on baseline adrenocorticotropic hormone and the response to thyrotropin releasing hormone-stimulation test in healthy horses.. Domest Anim Endocrinol 2014 Jul;48:77-83.
    pubmed: 24906932doi: 10.1016/j.domaniend.2014.02.004google scholar: lookup
  29. Restifo MM, Frank N, Hermida P, Sanchez-Londoño A. Effects of withholding feed on thyrotropin-releasing hormone stimulation test results and effects of combined testing on oral sugar test and thyrotropin-releasing hormone stimulation test results in horses.. Am J Vet Res 2016 Jul;77(7):738-48.
    pubmed: 27347827doi: 10.2460/ajvr.77.7.738google scholar: lookup
  30. Copas VE, Durham AE. Circannual variation in plasma adrenocorticotropic hormone concentrations in the UK in normal horses and ponies, and those with pituitary pars intermedia dysfunction.. Equine Vet J 2012 Jul;44(4):440-3.
    pubmed: 21848531doi: 10.1111/j.2042-3306.2011.00444.xgoogle scholar: lookup
  31. Donaldson MT, McDonnell SM, Schanbacher BJ, Lamb SV, McFarlane D, Beech J. Variation in plasma adrenocorticotropic hormone concentration and dexamethasone suppression test results with season, age, and sex in healthy ponies and horses.. J Vet Intern Med 2005 Mar-Apr;19(2):217-22.
    pubmed: 15822567doi: 10.1892/0891-6640(2005)19<217:vipahc>2.0.co;2google scholar: lookup
  32. Frank N, Elliott SB, Chameroy KA, Tóth F, Chumbler NS, McClamroch R. Association of season and pasture grazing with blood hormone and metabolite concentrations in horses with presumed pituitary pars intermedia dysfunction.. J Vet Intern Med 2010 Sep-Oct;24(5):1167-75.
    pubmed: 20666984doi: 10.1111/j.1939-1676.2010.0547.xgoogle scholar: lookup
  33. Jacob SI, Geor RJ, Weber PSD, Harris PA, McCue ME. Effect of dietary carbohydrates and time of year on ACTH and cortisol concentrations in adult and aged horses.. Domest Anim Endocrinol 2018 Apr;63:15-22.
    pubmed: 29195115doi: 10.1016/j.domaniend.2017.10.005google scholar: lookup
  34. McFarlane D. Diagnostic Testing for Equine Endocrine Diseases: Confirmation Versus Confusion.. Vet Clin North Am Equine Pract 2019 Aug;35(2):327-338.
    pubmed: 31076223doi: 10.1016/j.cveq.2019.03.005google scholar: lookup
  35. Secombe CJ, Tan RHH, Perara DI, Byrne DP, Watts SP, Wearn JG. The Effect of Geographic Location on Circannual Adrenocorticotropic Hormone Plasma Concentrations in Horses in Australia.. J Vet Intern Med 2017 Sep;31(5):1533-1540.
    pmc: PMC5598888pubmed: 28661009doi: 10.1111/jvim.14782google scholar: lookup
  36. Prutton JS, Kass PH, Watson JL, Pusterla N. Pre-analytical stability of adrenocorticotrophic hormone from healthy horses in whole blood, plasma and frozen plasma samples.. Vet J 2015 Apr;204(1):123-4.
    pubmed: 25744807doi: 10.1016/j.tvjl.2015.02.010google scholar: lookup
  37. Kam YN, McKenzie K, Coyle M, Bertin FR. Repeatability of a thyrotropin-releasing hormone stimulation test for diagnosis of pituitary pars intermedia dysfunction in mature horses.. J Vet Intern Med 2021 Nov;35(6):2885-2890.
    pmc: PMC8692209pubmed: 34642962doi: 10.1111/jvim.16281google scholar: lookup
  38. Frank N. Endocrine Disorders of the Equine Athlete.. Vet Clin North Am Equine Pract 2018 Aug;34(2):299-312.
    pubmed: 29857965doi: 10.1016/j.cveq.2018.04.003google scholar: lookup

Citations

This article has been cited 2 times.
  1. Kirkwood NC, Hughes KJ, Stewart AJ. Prospective Case Series of Clinical Signs and Adrenocorticotrophin (ACTH) Concentrations in Seven Horses Transitioning to Pituitary Pars Intermedia Dysfunction (PPID). Vet Sci 2022 Oct 17;9(10).
    doi: 10.3390/vetsci9100572pubmed: 36288186google scholar: lookup
  2. Kirkwood NC, Hughes KJ, Stewart AJ. Pituitary Pars Intermedia Dysfunction (PPID) in Horses. Vet Sci 2022 Oct 10;9(10).
    doi: 10.3390/vetsci9100556pubmed: 36288169google scholar: lookup
Subscribe to our newsletter
Join 99,658 horse owners like you who receive our email updates on horse health and nutrition.