Diagnostic Testing For Equine Endocrine Diseases
Diagnostic Testing For Equine Endocrine Diseases
Diagnostic Testing For Equine Endocrine Diseases
KEYWORDS
Equine pituitary pars intermedia dysfunction Equine metabolic syndrome ACTH
Insulin
KEY POINTS
No diagnostic tests should be interpreted without consideration of signalment, history,
and specific clinical signs of disease.
Measurement of hormones, both baseline or after stimulation, is affected by many other
factors, including concurrent illness, season, thriftiness, color, breed, and diet.
Any diagnostic test is only as good as the assay measuring the hormone.
Reference intervals are only valid for a specific method and assay. Breed-specific refer-
ence intervals are needed.
Although once a fairly obscure field, equine endocrinology now holds great interest to
the horse-owning public. The 2 common endocrine syndromes, pituitary pars interme-
dia dysfunction (PPID) and equine metabolic syndrome (EMS), are each frequently
featured in horse industry news outlets. In addition, the availability of highly marketed
pharmaceuticals has contributed to heightened awareness of these conditions by vet-
erinarians and clients alike. As awareness of equine endocrine diseases has grown,
more owners are requesting their horses be tested, often in the absence of convincing
clinical signs supportive of disease. Unfortunately, the current diagnostic tests are
poor at predicting endocrine disease when only minimal or nonspecific clinical signs
are present.
Equine endocrinology is a field that has undergone numerous transformations, with
its share of incorrect conclusions along the way. For example, historically it was
Disclosure Statement: Dr D. McFarlane has no conflict of interest. She has participated in the
Equine Endocrinology Group that provides recommendations on diagnostic testing for PPID
that is distributed by Boehringer Ingelheim (BI) in marketing handouts and in the Equine Endo-
crine Summit (a small group scientific meeting), both of which have been supported by BI.
Department of Physiological Sciences, Center for Veterinary Health Sciences, Oklahoma State
University, 264 McElroy Hall, CVHS-OSU, Stillwater, OK 74078, USA
E-mail address: diannem@okstate.edu
thought that hypothyroidism was a common equine disease and the mechanism
behind obesity-associated laminitis in horses. Many horses were prescribed lifelong
treatment with levothyroxine as a result. Later, the role of dysfunctional thyroid glands
in obesity-related laminitis was challenged, and the concept that hyperinsulinemia
was responsible for inciting endocrinopathic laminitis was introduced.1 Along with a
new understanding of the mechanism of endocrinopathic laminitis came a new
name for this condition, equine metabolic syndrome (EMS).2 The definition of EMS
has evolved since the term was first introduced in 2002, and changes in diagnostic
and treatment approaches have followed.2
The understanding of equine PPID has also undergone many shifts in the past
several decades. Once thought to be an uncommon condition, PPID affects 20% to
30% of the geriatric horse population.3–5 Clinical signs attributed to PPID have often
been based on poorly defined cases, and as a result, are inaccurate. Originally called
“equine Cushing disease,” the condition was thought to be similar to human and
canine Cushing disease. Pituitary-dependent hyperadrenocorticism was considered
to be the mechanistic cause, despite an inability to demonstrate high-serum cortisol
concentrations or enlarged adrenal glands in most equine cases.6–8 These miscon-
ceptions pervaded the veterinary literature for several decades.
Over the years, many diagnostic tests have been suggested that ultimately were
proven to be poor discriminators of disease. Problems from “interference” due to
physiologic endocrine activity, from overgeneralizing data collected in a small number
of nonrepresentative animals, poor case definition, to a lack of a gold standard,
plagued validation studies.
This article is not intended to serve as an exhaustive review of equine endocrine dis-
ease or testing strategies. Several comprehensive reviews have been recently pub-
lished.9–12 Rather, it discusses areas in the field of equine endocrine disease
diagnosis that need further consideration due to common misconceptions or misuse
of the current recommendations. In addition, suggestions for future directions for
enhanced diagnostic capabilities in equine endocrinology are included.
ASSAY MATTERS
It is important that the diagnostic tests used to identify cases have been appropriately
validated and that guidelines for interpretation of results be broadly applicable and ac-
curate. A well-accepted but often overlooked fact is that reference intervals are only
valid for the specific assay and methodology for which they were established. There
are numerous reports comparing the performance of radioimmunoassays, enzyme-
linked immunosorbent assays (ELISAs), chemiluminescent immunoassay, and immu-
nofluorescent assays for measuring adrenocorticotropic hormone (ACTH), insulin, and
other hormones; all show bias or lack of agreement among the assays, with a high per-
centage of discordant samples.13–18 Therefore, results among assays are not inter-
changeable, and reference intervals provided by laboratories or diagnostic
guidelines are only relevant if the samples are run using the same methodology.
When monitoring patients over time, the same method of hormone measurement
must be used if current and previous results are to be compared.
It is also important to understand the limitations and specificity of the assay used.
Most assays used to measure equine hormones are designed for use in other species,
most commonly human-specific assays. Validation for use with equine samples is
required, and details of the validation should be publicly available. Unfortunately, often
the validation process performed is not ideal. This failure to appropriately valid assays
may be the consequence of an absence of an unequivocal method (a “gold standard”)
Diagnostic Testing for Equine Endocrine Diseases 3
Many diagnostic tests for PPID have come and gone out of favor over the years.
Today, the most frequently used test in practice is endogenous plasma ACTH concen-
tration, and the test considered to be most discriminating is the thyrotropin-releasing
hormone (TRH) stimulation assay.20,21 The overnight dexamethasone suppression test
(DST), although no longer commonly used, is still considered an appropriate albeit less
sensitive diagnostic test. All of the current tests have their strengths and weaknesses;
understanding how to best use and interpret the available tests will aid in a better likeli-
hood of reaching an accurate diagnosis.
dopaminergic neuronal activity. None are able to characterize whether ACTH is of pars
intermedia or pars distalis origin.
Test mechanism
In healthy horses, administration of dexamethasone suppresses adrenal cortisol
release by inhibiting corticotrope-derived ACTH and hypothalamic corticotropin-
releasing hormone secretion. Glucocorticoids do not inhibit ACTH production by
pars intermedia melanotropes.
6 McFarlane
EMS is a list of clinical signs or pathologic conditions that, if present, increase risk of
endocrinopathic laminitis.56 Among these risk factors, insulin dysregulation is the
strongest predictor of laminitis.
Insulin-glucose regulation is complex, and different parts of the insulin-glucose reg-
ulatory pathway may be altered in horses that have insulin dysregulation. Conse-
quently, not all horses with insulin dysregulation will respond similarly to a specific
diagnostic test.
The specificity of individual diagnostic tests in assessing insulin-glucose regulation
is illustrated by a series of studies investigating metformin efficacy. Initial studies
considered the ability of metformin to improve insulin sensitivity in horses using a
frequently sampled intravenous glucose tolerance test.57 No improvement was
observed. Later work used an oral glucose test and demonstrated a highly significant
reduction in both glucose and insulin concentration.58 Pharmacokinetic studies sug-
gest metformin is not well absorbed when given orally to horses.59 However, in other
species, metformin can elicit a local gastrointestinal effect, reducing small intestinal
glucose absorption and subsequent pancreatic insulin release. A local gastrointes-
tinal effect would only be appreciated with an oral glucose challenge test, not an
intravenous glucose challenge. Although a local effect of metformin in the horse
has not been indisputably demonstrated, this explanation fits all the available equine
data.
The story of metformin in horses serves as an example of how results from the
various diagnostics tests are not interchangeable, and no one diagnostic test is ideal
for all cases. Some horses and ponies with EMS have a more profound tissue insulin
resistance, whereas others have a more exaggerated glucose-stimulated insulin
release. Both pathways lead to hyperinsulinemia and thus increased risk of endocrino-
pathic laminitis.
Baseline Testing
Equine serum insulin concentration
Measurement of serum insulin concentration as a diagnostic test for insulin resis-
tance is a noninvasive, technically easy method. However, false negative results
are extremely common.21 Guidelines suggest normal serum insulin concentration
is less than 20 mU/mL; however, for some breeds, normal serum insulin concentra-
tion is lower.60 Opinions on the best type of resting sample (fasted, fed, or feed
restricted) change frequently and continue to be debated. Current recommenda-
tions suggest withholding grain but allowing availability to hay or pasture when
testing.
Dynamic Testing
Oral sugar or glucose tests
To reduce the high false negative rate, a dynamic test is suggested. Postprandial
insulin release may be assessed using an oral sugar test or an in-feed oral glucose
test. The oral sugar test provides the horse a sugar bolus in the form of Karo syrup
administered via dose syringe.61 The in-feed oral glucose test provides a sugar
bolus in the form of dextrose powder in a nonglycemic feed such as chaff.62
Both evaluate glucose absorption and postprandial insulin release. Debates
regarding these tests center primarily on the optimal dose of sugar and degree
of fasting. Significant breed differences occur, suggesting multiple reference inter-
vals are needed.60
8 McFarlane
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