Endocrine disorders and laminitis

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1 bs_bs_banner 152 EQUINE VETERINARY EDUCATION Equine vet. Educ. (2013) 25 (3) doi: /j x Review Article Endocrine disorders and laminitis E. M. Tadros and N. Frank* Department of Large Animal Clinical Sciences, College of Veterinary Medicine, University of Tennessee, Knoxville, Tennessee, USA; Department of Clinical Sciences, Cummings School of Veterinary Medicine, Tufts University, North Grafton, Massachusetts, USA; and Division of Medicine, School of Veterinary Medicine and Science, University of Nottingham, Sutton Bonington Campus, Leicestershire, UK. *Corresponding author Keywords: horse; endocrinopathic laminitis; Cushing s; equine metabolic syndrome; pars intermedia; insulin resistance Summary Two common endocrine disorders, pituitary pars intermedia dysfunction and equine metabolic syndrome, predispose horses and ponies to laminitis and may even induce the condition. The exact mechanisms involved in endocrinopathic laminitis have not been elucidated but hyperinsulinaemia and insulin resistance are currently being investigated. Obesity and regional adiposity may also contribute to laminitis susceptibility through the release of inflammatory cytokines and adipokines. In the case of pituitary pars intermedia dysfunction, glucocorticoid excess is likely to weaken hoof structures, alter vascular dynamics within the foot and induce or exacerbate insulin resistance. This review will summarise current theories regarding the pathophysiology of endocrinopathic laminitis and provide recommendations for the diagnosis and management of these common equine endocrine disorders. Introduction The 2 endocrine disorders that we will discuss in this review are pituitary pars intermedia dysfunction (PPID), also known as equine Cushing s disease, and equine metabolic syndrome (EMS). Alternative names for EMS include insulin resistance syndrome, peripheral Cushing s syndrome and prelaminitic metabolic syndrome. Pituitary pars intermedia dysfunction is an endocrine disorder of older horses and ponies and has been reviewed in greater detail by McFarlane (2011). Most affected animals are more than 15 years of age when clinical signs of PPID are first noted and the risk of disease increases with age (Table 1). Horses with PPID suffer from hyperplasia or neoplasia of the pars intermedia, whereas pituitary-dependent hyperadrenocorticism in other species such as the dog usually develops in the pars distalis. Corticotrophs of the pars distalis and melanotrophs of the pars intermedia secrete hormones derived from the prohormone pro-opiomelanocortin (POMC). While adrenocorticotropin hormone (ACTH), beta-endorphin and beta-lipotropin are the primary products of POMC processing in the pars distalis, melanotrophs of the pars intermedia further process ACTH and release alpha melanocyte stimulating hormone (a MSH) and corticotropin-like intermediate peptide (CLIP). Mammalian pars intermedia melanotrophs are under tonic inhibition by dopaminergic periventricular neurons (Saland 2001) and loss of dopaminergic inhibition results in pars intermedia hyperplasia. Neuronal degeneration appears to result from age-related oxidative stress (McFarlane 2007; McFarlane and Holbrook 2008), but other systemic conditions such as obesity may also play a role. Obesity in human is associated with increased oxidative stress due to the production of inflammatory mediators (Fernandez-Sanchez et al. 2011), although one study evaluating oxidative stress markers in obese ponies produced equivocal results (Treiber et al. 2009). Clinical signs of PPID are attributable to higher circulating concentrations of POMC-derived hormones as well as the space-occupying effects of adenomas on surrounding tissues in advanced cases. Classical signs include delayed shedding of the winter haircoat, hypertrichosis (commonly referred to as hirsutism), skeletal muscle atrophy, polyuria and polydipsia. Pituitary pars intermedia dysfunction is associated with laminitis (Johnson et al. 2002; Donaldson et al. 2004), but a direct cause and effect relationship has not been established. Equine metabolic syndrome is both an endocrine and metabolic disorder and is described in greater detail in a consensus statement released by the American College of Veterinary Internal Medicine (Frank et al. 2010c). Key components of EMS include enhanced metabolic efficiency, increased adiposity, insulin resistance (IR) and hyperinsulinaemia. Most importantly, affected horses and ponies are predisposed to pasture-associated laminitis (Treiber et al. 2006). This syndrome is more common in ponies, Morgan horses, Paso Finos, Arabians and Warmbloods and is likely to be a heritable trait (Treiber et al. 2006). Genetically predisposed horses often express the phenotype after overfeeding so both inherent and environmental factors play a role in the development of EMS (Bailey et al. 2008; Carter et al. 2009b). Excess grain feeding or grazing on large pastures induce obesity, which is a component of EMS in most cases. Regional adiposity also develops as adipose tissues expand within the neck region and other sites throughout the body. Enlargement of the neck crest is a physical characteristic that has been used as a phenotypic marker for EMS because neck circumference and neck crest scores are negatively correlated with insulin sensitivity in horses and ponies (Frank et al. 2006; Carter et al. 2009a). Fat pads can also develop near the tailhead or within the prepuce or mammary gland regions. Many horses with EMS develop laminitis after being turned out on a new pasture or following rapid growth of grass in the spring or late summer, and obesity, regional adiposity and IR are all established risk factors for pasture-associated laminitis in ponies (Treiber et al. 2006; Carter et al. 2009c).

2 E. M. Tadros and N. Frank 153 TABLE 1: Clinical presentation of pituitary pars intermedia dysfunction and equine metabolic syndrome Pituitary pars intermedia dysfunction Pathophysiology Loss of dopaminergic inhibition leads to hyperplasia or neoplasia of the pituitary pars intermedia Clinical presentation Typically older than 15 years Hypertrichosis (hirsutism) or delayed shedding of haircoat; long hairs may be retained year round on palmar and plantar aspect of limbs, behind elbows, or under mandibles; coat may appear dull Hyperhidrosis, especially over neck and shoulders Polyuria and polydipsia Weight loss or skeletal muscle atrophy; some animals may develop a sway-backed or pot-bellied appearance Abnormal deposition of fat along the crest of the neck, above the tail head, or in the sheath/mammary region Recurrent or chronic infections such as sinusitis, skin infection, dental disease, or hoof abscesses Reproductive abnormalities including infertility and persistent lactation Chronic laminitis, often insidious in onset Therapeutic goals Improvement in clinical signs and endocrine abnormalities through pharmacological intervention Provision of supportive care including body clipping, appropriate farrier care, dentistry and nutrition Equine metabolic syndrome Pathophysiology Genetic predisposition (suspected) to obesity and insulin resistance Clinical presentation Typically 5 15 years of age when detected Obesity and regional adiposity; prominent fat pads along the crest of the neck, above the tail head or in the sheath/ mammary region; animals often described by owners as easy keepers Occasional animals may be lean with abnormal fat deposition Infertility or abnormal cyclicity in mares Chronic laminitis; acute episodes may occur after exposure to pasture that is high in nonstructural carbohydrate content Insulin resistance is a consistent finding; dynamic endocrine testing may be necessary to detect insulin resistance if resting samples are inconclusive Therapeutic goals Improvement in insulin sensitivity through weight loss, exercise and a reduction in dietary carbohydrate consumption; pharmacological intervention in some cases Appropriate farrier care to manage chronic laminitis Insulin resistance and hyperinsulinaemia are components of EMS. Hyperinsulinaemia accompanies IR in most animals and is used as a diagnostic marker for this problem (Treiber et al. 2005). The degree of IR can vary in an individual animal over time due to factors such as dietary changes (Hoffman et al. 2003), fluctuations in body condition and fitness (Freestone et al. 1992; Carter et al. 2009b), the production of stress hormones (Geor et al. 2000; Tiley et al. 2007) and by season (Bailey et al. 2008; Frank et al. 2010a). Insulin resistance is defined as failure of tissues to respond appropriately to insulin (Kahn 1978). There are several reasons why tissues become insulin resistant, including a reduction in the density of insulin receptors on cell surfaces, malfunction of insulin receptors, defective internal signalling pathways and interference with the synthesis or function of glucose transporter 4 proteins (GLUT4). Although the exact mechanism of IR in EMS remains to be fully elucidated, altered GLUT4 protein trafficking has recently been demonstrated in adipose tissue and skeletal muscle from insulin-resistant horses (Waller et al. 2011a,b). It is also important to consider the concept of converging endocrinopathies in middle aged horses (10 20 years of age). The authors hypothesise that ponies and horses with EMS are predisposed to PPID because low grade inflammation and oxidative stress associated with obesity contributes to neuronal degeneration and loss of dopaminergic inhibition. Animals with EMS should therefore be closely monitored for PPID as they enter middle age and clinical signs of both endocrinopathies are detected in some cases. One important indication that this is occurring comes from the history with owners often reporting that their obese middle-aged horse, with a history of EMS, has undergone a shift in metabolism and begun to lose condition. The same horse might have retained its winter haircoat for a few weeks longer or seemed to have aged rapidly in the last few months. These animals are concurrently affected by EMS and PPID and may be at higher risk of developing laminitis. Insulin resistance associated with EMS likely persists after PPID develops and may be exacerbated by glucocorticoid excess. Potential pathophysiological mechanisms of endocrinopathic laminitis Hyperadrenocorticism When PPID develops, melanotrophs of the pars intermedia secrete more POMC-derived hormones, including amsh, CLIP and ACTH. Higher ACTH concentrations are detected in affected animals and this hormone stimulates cortisol secretion from the adrenal glands. A state of hyperadrenocorticism is induced, although serum cortisol concentrations sometimes fall within reference range in horses with PPID and gross adrenal hyperplasia is an uncommon finding (Schott 2002; Miller et al. 2008). It is therefore assumed that hyperadrenocorticism results from loss of circadian rhythm and increased daily cumulative cortisol concentrations (Dybdal et al. 1994; Haritou et al. 2008). Cortisol concentrations can also be lower than expected in horses with high ACTH concentrations, suggesting that ACTH has lower biological activity in some PPID horses. Histological alterations in the integument associated with hyperadrenocorticism in other species include protein depletion, inhibition of fibroblast growth and reduced collagen synthesis (Johnson et al. 2002; Kahan et al. 2009). Johnson et al. (2002) described lengthening and attenuation of primary and secondary dermal lamellae in horses with glucocorticoid excess and suggested that this represents pulling apart of lamellae as structures weaken. Vascular changes may also accompany hyperadrenocorticism. Flow-mediated vasodilation of the brachial artery is reduced in

3 154 Endocrine disorders and laminitis human with Cushing s disease (Baykan et al. 2007) and hypertrophic remodelling of small resistance arteries occurs within subcutaneous tissues (Rizzoni et al. 2009). Hooves of horses with PPID may therefore be more susceptible to lamellar failure and less capable of repair after laminitis has occurred. These assumptions are based on theory rather than scientific evidence at this point and further studies are required to examine hooves from horses with PPID. Glucocorticoids inhibit the actions of insulin by disrupting post receptor signalling pathways (Buren et al. 2002; Ruzzin et al. 2005) and administration of dexamethasone has been shown to induce IR in horses (Tiley et al. 2007, 2008; Tóth et al. 2010). Interestingly, hyperinsulinaemia is detected in some horses with PPID (Reeves et al. 2001; Schott 2002), whereas others have normal insulin concentrations. One explanation for this observation is that IR only occurs with PPID when the horse is predisposed to this problem. An alternative explanation is that horses with PPID differ in the types and amounts of POMC-derived hormones secreted from the pars intermedia, so those with IR have higher ACTH-stimulated cortisol secretion. It is important to assess insulin sensitivity in horses with PPID because affected animals respond differently to their diet and are predisposed to laminitis. In a recent study, horses with PPID had higher overall insulin concentrations across a 12 month period while grazing on pasture when compared to unaffected aged horses (Frank et al. 2010b). Insulin concentrations above mu/ml have also been identified as a poor prognostic indicator for 1 2 year survival in horses with PPID (McGowan et al. 2004). It should also be noted that insulin concentrations can increase for other reasons, including systemic inflammation, stress and pain (Marik and Raghavan 2004; Frank 2009) and responses may differ in horses with PPID. One final point is that some horses with PPID suffer from laminitis whereas others do not. The authors have observed that horses with PPID and concurrent IR are more likely to suffer from laminitis than those with normal insulin sensitivity, which suggests that IR is a key determinant or marker of laminitis susceptibility. Obesity Obesity, IR, hyperinsulinaemia and laminitis are associated in equids, but it should be noted that some obese animals exhibit normal insulin sensitivity when tested (Treiber et al. 2006; Vick et al. 2007; Carter et al. 2009c). These animals may be more tolerant of obesity or require more time for IR to develop. One theory linking obesity with IR is the release of inflammatory cytokines from adipose tissues. More tumour necrosis factor alpha (TNFa) is secreted from adipose tissues as body mass index increases in human and this inflammatory cytokine inhibits insulin receptor signalling, which lowers insulin sensitivity (Hartge et al. 2007). Vick et al. (2007) detected higher blood TNFa mrna expression in obese horses, which suggests that the same mechanism contributes to obesity-associated IR in equids. Increased inflammatory cytokine production by adipose tissues may also contribute to laminitis susceptibility. Obesity also affects adipokine production by adipose tissues. Adipokines are hormones produced by adipocytes that have local (paracrine) and remote (endocrine) effects on tissues. Leptin and adiponectin are the most well known adipokines and obesity has been associated with elevated plasma leptin concentrations and lower plasma adiponectin concentrations in horses (Kearns et al. 2006). Adiponectin Insulin SENSITIVE Insulin RESISTANT ENDOTHELIAL CELL ENDOTHELIAL CELL PI3K pathway MAPK pathway [Phosphatidylinositol-3 kinase] [Mitogen-activated protein kinase] VASODILATION Nitric oxide VASOCONSTRICTION Endothelin-1 Fig 1: Theoretical relationships between insulin sensitivity and vascular tone in horses. Alterations in insulin sensitivity may determine which pathway predominates after activation by circulating insulin. enhances insulin sensitivity, so lower plasma concentrations are associated with IR (Kearns et al. 2006). Low adiponectin concentrations are also associated with impaired endothelium-dependent vasodilation in obese humans (Ritchie et al. 2004). Resistin is another adipokine that affects insulin sensitivity. This hormone has been examined in mice and human and hyper-resistinaemia is associated with IR and type 2 diabetes mellitus (Radin et al. 2009). It should be noted that some insulin resistant horses exhibit a leaner overall body condition. Abnormal fat deposition is present regionally in some lean animals and may be responsible for the production of inflammatory mediators and adipokines that lower insulin sensitivity. Other horses appear normal and the mechanisms underlying IR in these animals require further study. Finally, laminitis may occur more readily in obese horses because they carry more weight on their hooves, which increases forces exerted upon dermo-epidermal attachments. Insulin resistance and vascular dynamics Insulin possesses vasoregulatory properties and this might explain why IR predisposes horses to laminitis. Slow vasodilation occurs in response to insulin through increased synthesis of nitric oxide (NO) by endothelial cells (Muniyappa et al. 2007). However, insulin also promotes vasoconstriction by stimulating endothelin-1 (ET-1) synthesis and activating the sympathetic nervous system. Under normal conditions, the opposing actions of NO (vasodilation) and ET-1 (vasoconstriction) are in balance because both arms of the insulin signalling cascade are active; activation of the insulin receptor therefore stimulates 2 different signalling pathways within the vascular endothelial cell (Fig 1). Nitric oxide is secreted when the phosphatidylinositol 3-kinase (PI3K) pathway is activated, whereas activation of the mitogen-activated protein kinase (MAPK) pathway leads to release of ET-1 (Muniyappa et al. 2008). Both the vasodilatory effects of insulin and insulin-dependent stimulation of glucose uptake are mediated by PI3K and this pathway becomes disrupted when IR develops. Consequently, vasoconstriction is promoted in insulin resistant animals because only the MAPK pathway

4 E. M. Tadros and N. Frank 155 remains fully functional. Development of compensatory hyperinsulinaemia in response to IR can further stimulate MAPK signalling and increase ET-1 synthesis (Kim et al. 2006). Eades et al. (2007) detected an increase in plasma ET-1 concentration within blood collected from digital veins 12 h after carbohydrate was administered to induce laminitis in healthy horses. This finding suggests that digital vessels undergo vasoconstriction as a result of carbohydrate overload in horses, which may contribute to the development of laminitis. Horses with chronic IR may be more likely to develop laminitis since vasoconstriction is already promoted. Hyperinsulinaemia and vascular dynamics Laminitis has been experimentally-induced in healthy ponies and Standardbred horses by inducing hyperinsulinaemia (Asplin et al. 2007; de Laat et al. 2010). In both studies, glucose and insulin were infused i.v. according to the euglycaemic-hyperinsulinaemic clamp procedure, with mean serum insulin concentrations exceeding 1000 mu/ml. Mean time to onset of Obel grade 2 laminitis was 46 h in horses and 55 h in ponies and hoof wall surface temperature increased in response to insulin infusion, indicating that vasodilation occurred within the foot. These results suggest that hyperinsulinaemia itself induces laminitis through a mechanism involving vasodilation. If this is the case, then hyperinsulinaemia-induced vasodilation would overcome vasoconstriction promoted by IR. It has also been proposed that hyperinsulinaemia-induced vasodilation increases glucose delivery to hoof tissues, leading to local glucotoxicity (de Laat et al. 2010). This may lead to the formation of advanced glycation end products that damage tissues. Advanced glycation end products develop as glucose reacts with amino acids within tissues and these products play an important role in the development of diabetic angiopathy in human (Yamagishi 2009). Clinical diagnosis and management of pituitary pars intermedia dysfunction Presenting complaints Classical clinical signs of PPID include hypertrichosis (commonly referred to as hirsutism) and affected horses are relatively easy to identify once the condition has become advanced. Early disease, however, is more challenging to recognise. Early signs of PPID include delayed shedding of the winter haircoat, a perceived shift in metabolism, regional adiposity, infertility and laminitis. Delayed shedding of the winter haircoat Affected horses exhibit delayed shedding of the winter haircoat, increased hair length (hypertrichosis) and dullness of the haircoat. If this problem is suspected, owners should record the time that their horse sheds its winter haircoat and compare it with other horses in the same barn. Retention of winter hairs in certain regions of the body Some horses with early PPID shed most of their winter haircoat, but retain hairs along the palmar or plantar aspects of the legs, behind the elbow or beneath the Fig 2: Photograph of the right side of a horse with retained winter hairs. Longer hairs that were lighter in colour were present in this region. mandibles. These hairs can become lighter in colour over time as a result of exposure to sunlight (Fig 2). Shift in metabolism Owners sometimes report that their horse has undergone a shift in metabolism and is now losing weight and body condition. This problem is difficult to confirm unless a complete dietary history has been recorded but clinicians should suspect PPID when a middle-aged horse begins to lose muscle mass and requires more calories than it did in the past. Regional adiposity This clinical sign is associated with both EMS and PPID. In the case of EMS, regional adiposity first develops at an earlier age and then persists. In contrast, detection of regional adiposity for the first time in an older (>15 years) horse indicates the development of PPID. As discussed below, horses with EMS can develop PPID as they get older. These patients have fat deposits that were previously associated with EMS and then become a component of PPID. Infertility Pituitary pars intermedia dysfunction should be considered if an aged mare develops fertility problems. More research is required to determine the effects of PPID on the reproductive cycle and uterine environment. At present, only anecdotal evidence is available to suggest that reproductive performance improves in mares with PPID that receive treatment. Diagnostic testing Resting ACTH concentrations and the overnight dexamethasone suppression test (DST) are easily performed (Table 2), but the sensitivity of these tests must be questioned for early PPID. In a study performed by Miller et al. (2008),

5 156 Endocrine disorders and laminitis TABLE 2: Recommended diagnostic tests for pituitary pars intermedia dysfunction PPID: Resting ACTH concentration Considerations Higher concentrations are detected in healthy horses in the late summer and autumn Pain and stress raise ACTH concentrations Procedure Collect blood in a tube containing EDTA Keep the blood sample cool at all times Centrifuge the same morning or afternoon Mail to the laboratory with a cool pack Interpretation Positive if ACTH >35 pg/ml (>50 pg/ml in August to October) Results may fall within reference range for horses with early/ mild PPID. Use the fall as a natural stimulation test PPID: Dexamethasone suppression test Considerations False positives are detected in healthy horses in the late summer and autumn Owner concerns about inducing laminitis Procedure Collect a preinjection blood sample (optional) Inject 0.04 mg/kg bwt (20 mg for a 500-kg horse) dexamethasone i.v. or i.m. Collect a blood sample 19 or 24 h later Interpretation Positive if cortisol concentration >10 ng/ml (27 nmol/l) at 19 or 24 h Results may be normal in horses with early/mild PPID Only the negative result is significant when testing in the late summer or autumn resting ACTH concentrations were within reference range for 5 of 5 horses with grade 3/5 pituitary pars intermedia lesions, 6 of 12 horses with grade 4/5 lesions and 1 of 4 horses with grade 5/5 lesions. In addition, measurement of plasma ACTH during the late summer and early fall has been avoided to minimise the likelihood of obtaining false-positive results due to seasonal increases in ACTH concentration (McFarlane et al. 2004; Donaldson et al. 2005). However, recent studies have suggested that differences in plasma ACTH concentrations between normal horses and those affected by PPID may be most pronounced during this time (Frank et al. 2010a), therefore establishing season-specific reference ranges to facilitate testing in the late summer and early fall could improve the sensitivity of this diagnostic procedure. The DST had high specificity when examined by Beech et al. (2007), but sensitivity ranged from 23 66% depending upon which groups were compared. Other tests for PPID are being developed, including the thyrotropin-releasing hormone stimulation test (Beech et al. 2007) and oral domperidone challenge (Miller et al. 2008) and these tests warrant consideration. A complicating factor in the development of antemortem diagnostic tests is the lack of a reliable gold standard for identifying PPID. Results have been compared to post mortem histological evaluation of the pituitary; however, there is little consensus among pathologists on the histological criteria that define PPID and therefore only moderate agreement between antemortem and post mortem diagnosis (McFarlane et al. 2005). Pituitary lesions have also been identified in clinically normal animals (van der Kolk et al. 2004) and overlap between normal ageing changes and early disease further complicates diagnosis. It should also be noted that all available antemortem diagnostic tests are limited in their ability to detect early or mild disease so the clinician must sometimes decide to institute treatment on the basis of clinical judgement alone. In these cases, pergolide can be administered as a therapeutic trial. Management Pergolide mesylate This ergot alkaloid dopamine receptor agonist is administered to horses with PPID to restore dopaminergic inhibition of melanotrophs. Interaction of the drug with D2 receptors inhibits hormone secretion and may also slow the progression of disease (Donaldson et al. 2002). Pergolide is prescribed on a total dose basis, using a starting dose of 1.0 mg/day for horses and larger ponies with PPID. A lower starting dose of mg/kg bwt (range of mg/kg bwt daily) can be calculated for small ponies or miniature horses. Some owners report loss of appetite when pergolide is first started. If anorexia develops, treatment should be halted for 2 days or until appetite improves and then restarted at 0.25 mg/day for 2 days, 0.5 mg/day for 2 days and 0.75 mg/day for 2 days. Another side effect of pergolide treatment is temporary dullness after initiating therapy and this can be addressed using the same approach. Horses with advanced PPID receive higher dosages, with a maximum daily dosage of 5 mg/day for horses with severe disease. Pergolide and cyproheptadine can also be administered in combination. It must be recognised that the goals of medical treatment vary with disease severity; the dosage selected for horses with early PPID should be adjusted until normal diagnostic test results are obtained, whereas treatment is palliative in horses with advanced disease. It is only reasonable to expect medical treatment to ameliorate disease in these advanced cases of PPID. Cyproheptadine Both pergolide and cyproheptadine lower plasma ACTH concentrations in horses with PPID (Perkins et al. 2002), but pergolide is more effective (Donaldson et al. 2002). Donaldson et al. (2002) reported that 17 of 20 horses (85%) with PPID improved with pergolide treatment, compared with only 2/7 horses treated with cyproheptadine. Cyproheptadine antagonises serotonin, which is thought to be a stimulatory neurotransmitter for pars intermedia melanotrophs. Treated horses occasionally exhibit sedation when treatment is initiated. A dosage of 0.25 mg/kg bwt per os q. 12 h is recommended.

6 E. M. Tadros and N. Frank 157 Trilostane This drug is a 3-beta hydroxysteroid dehydrogenase inhibitor and acts by inhibiting corticosteroid synthesis within the adrenal cortex. Trilostane is commonly used in dogs for the management of pituitary-dependent hyperadrenocorticism when increased ACTH secretion from the pars distalis induces adrenal hyperplasia. However, the reported incidence of adrenal hyperplasia is low (20%) in horses with PPID (Schott 2002) so this may limit the usefulness of trilostane in this species. With this stated, McGowan and Neiger (2003) reported improved clinical signs in 20 horses with PPID treated with trilostane at a mean dosage of 0.5 mg/kg bwt. Lethargy resolved, polyuria/polydipsia decreased and recurrent or chronic laminitis improved in 13 of 16 affected horses. Combined dexamethasone/thyrotropin-releasing hormone test results improved after treatment but did not return to normal. The recommended dosage for trilostane has subsequently been increased to 1.0 mg/kg bwt (q. 24 h; given in the evening) for horses with PPID. Combination therapy Some horses with PPID respond to combination treatment with pergolide and cyproheptadine. One approach is to reach the maximum pergolide dosage of 5 mg/day and then add cyproheptadine treatment (0.25 mg/kg bwt per os q.12 h) to the regimen. Other clinicians add cyproheptadine when the pergolide dosage reaches 3 mg/day (Schott 2006). Dietary management of PPID Since some horses with PPID are insulin-resistant and others have normal insulin sensitivity, the diet of each affected horse must be adjusted accordingly. Insulin-resistant PPID horses are challenging to manage if the animal is losing weight or being worked. In these cases caloric intake must be increased without exacerbating IR. A diet of hay and low-sugar/low-starch pellets is recommended with one half to 1 cup vegetable oil added twice daily to increase digestible energy intake. Pasture access should be controlled to avoid sudden changes in carbohydrate intake. In contrast, insulin-sensitive PPID horses can be fed as normal, with senior feeds, sweet feed or oats fed with hay to provide additional calories if needed. Clinical diagnosis and management of equine metabolic syndrome Presenting complaints Age Equine metabolic syndrome first develops in the younger horse (<15 years of age) and it is likely that this condition has a genetic basis. Some horses become obese when they reach maturity whereas others develop obesity later in life when they are overfed and inadequately exercised. Fig 3: Photograph of the enlarged neck crest of a horse with Equine Metabolic Syndrome. In general, EMS should be suspected in any obese horse with regional adiposity that the owner describes as an easy keeper or good doer. Obesity and regional adiposity Most horses with EMS are obese and show enlargement of adipose tissues within the neck crest (Frank et al. 2006; Carter et al. 2009a), commonly referred to as a cresty neck (Fig 3). Fat pads can also develop close to the tailhead and within the prepuce or mammary gland region and horses sometimes present with the complaint of preputial swelling or mammary gland enlargement. Affected horses occasionally have randomly distributed subcutaneous adipose tissue deposits. If regional adiposity first develops in an older horse (>15 years), it is more likely to be associated with PPID. Laminitis Acute laminitis is the presenting complaint for many horses with EMS. Laminitis appears to occur spontaneously without any history of grain overload, bacterial disease or retained placenta. However, further questioning of the owner or examination of the surroundings may reveal that laminitis is associated with changes in the pasture grass, including rapid growth in the spring, drought in the summer, frost in the winter or transfer to a new pasture. Laminitis may also occur at a subclinical level and lead to structural changes within the hoof that manifest as divergent hoof rings or radiographic evidence of distal (third) phalanx rotation. Infertility Impaired reproductive performance may be a presenting complaint for EMS. Alterations in reproductive cycling have been detected in obese insulin-resistant mares (Vick et al. 2006).

7 158 Endocrine disorders and laminitis TABLE 3: Recommended diagnostic tests for insulin resistance EMS: Resting insulin concentration Considerations Use as a screening test Also measure the glucose concentration; persistent hyperglycaemia is a major concern Procedure Ask the owner to leave only one flake of hay in the stall after h the night before and do not feed the horse until after samples are collected Collect blood in the morning Submit samples for glucose and insulin concentrations Interpretation Hyperinsulinaemia if the insulin concentration exceeds 20 mu/ml (mu/l) Glucose concentrations are within reference range in most cases (euglycaemia) Detection of persistent hyperglycaemia indicates type 2 diabetes mellitus; insulin concentration may be normal or elevated EMS: Combined glucose insulin test Considerations Used to detect insulin resistance (IR) when the fasting insulin concentration is normal or when you want to quantify insulin sensitivity Risk of inducing clinical hypoglycaemia in insulin-sensitive horses Administer ml 50% dextrose and feed the horse if hypoglycaemia occurs Procedure Collect a preinjection blood sample Infuse 150 mg/kg bwt glucose (50% dextrose) and then 0.10 iu insulin i.v. Collect blood samples at 1, 5, 15, 25, 35, 45, 60, 75, 90, 105, 120, 135 and 150 min (or until normal result) and measure glucose on a hand-held glucometer. Measure insulin concentrations at 0 and 45 min Interpretation Insulin resistance is present if the blood glucose concentration is above the preinjection value for 45 min An insulin concentration >100 mu/ml at 45 min indicates an excessive pancreatic response consistent with compensated IR New tests in development Oral sugar test: Corn syrup (0.15 ml/kg bwt) is administered orally via dose syringe and a blood sample is drawn min later. Higher blood glucose (>115 mg/dl [6.4 mmol/l]) and insulin (>60 mu/ml) concentrations are detected in insulin-resistant horses. Infeed oral glucose challenge test: Dextrose powder (1 g/kg bwt) is mixed with a nonglycaemic feed (e.g. chaff) and fed to the horse. A blood sample is collected 2 h later and an insulin concentration >85 mu/ml is supportive of IR. Diagnostic testing Diagnostic testing for EMS currently focuses upon detection of IR and hyperinsulinaemia. Tests vary in their complexity and a compromise must be reached between accuracy and ease of testing. It must also be recognised that all of the available tests are affected by pain and stress, which accompanies laminitis and testing should be delayed until the animal has stabilised. Unfortunately this prevents the clinician from assessing the animal in its exacerbated state, which is when laminitis first developed. Results must be interpreted accordingly as horses with mild IR at the time of testing may have been more severely affected when laminitis developed several weeks beforehand. Resting insulin concentration This is a screening test for IR and easy to perform. Horses must undergo a short fast before samples are collected and this is accomplished by leaving only one flake of hay with the animal after h the night before and then collecting a blood sample the following morning. Reference ranges for insulin vary among laboratories according to the assay used and control population. With the radioimmunoassay used in our laboratory, a cut-off value of 20 mu/ml defines hyperinsulinaemia (Table 3). Because factors such as stress, duration of fasting and possibly diurnal variation can impact insulin sensitivity over short periods of time (Ralston 2002), single resting insulin concentrations outside of reference range or small variations in serial measurements should be interpreted with caution. Results that fall far outside the normal range are more likely to indicate a true positive. Glucose concentrations should be measured at the same time. Most insulin-resistant horses maintain euglycaemia and hyperglycaemia is a significant concern. Hyperglycaemia signals loss of glycaemic control, which is seen in some patients with pancreatic insufficiency. Resting insulin concentrations may be high or within reference range in these patients and hyperglycaemia is the key finding. This is referred to as type 2 diabetes mellitus if hyperglycaemia persists (Durham et al. 2009). Dynamic testing There are occasions when EMS is strongly suspected on the basis of the history and clinical findings, yet resting insulin concentrations fall within the reference range. In these situations, a dynamic test should be performed to challenge the system. This can be accomplished by performing an i.v. glucose tolerance test, administering dextrose orally or feeding a test meal. For all of these tests, the horse or pony should be fasted by leaving only one flake of hay in the stall after h the night before. A combined glucose-insulin test (CGIT) has been developed for use in horses and can be used to assess insulin sensitivity in approximately 1 h (Eiler et al. 2005; Frank 2009). This test involves insertion of an i.v. catheter and administration of dextrose followed immediately by insulin. An oral sugar test

8 E. M. Tadros and N. Frank 159 (OST) has also been introduced in the USA where corn syrup is readily available in supermarkets. Corn syrup is administered orally after a short fast at a dosage of 0.15 ml/kg bwt (150 mg sugar/kg bwt) and a blood sample is drawn min later. Higher blood glucose (>115 mg/dl [6.4 mmol/l]) and insulin (>60 mu/ml) concentrations are detected in insulin-resistant horses. An in-feed oral glucose challenge test (OGT) can be performed as an alternative by mixing dextrose powder at a dosage of 1 g/kg bwt with a nonglycaemic feed. A blood sample is collected 2 h later and insulin concentrations >85 mu/ml are consistent with IR. Although administration of sugar to insulin-resistant horses may be of concern to some owners, the authors are not aware of any cases in which testing procedures have induced laminitis. Management Equine metabolic syndrome is a disorder that should be managed with diet, housing and exercise changes, rather than drugs in most cases. The 2 principal strategies for addressing IR in horses are to 1) induce weight loss in obese horses and ponies and 2) improve insulin sensitivity through dietary management and exercise (Geor and Harris 2009). Obese horses should be placed on a weight reduction diet composed of hay plus a protein, vitamin and mineral supplement. Horses should initially receive hay in amounts equivalent to 1.5% of ideal bwt per day (i.e. 7.5 kg for a 500 kg horse). If the horse or pony does not begin to lose weight after one month on this diet, the amount of hay fed should be lowered to 1.0% of ideal bwt (5 kg for a 500 kg horse). The weight reduction diet should be continued until an ideal body condition score has been attained. Grain or pellets should be completely eliminated from the diet and there should be no access to pasture during the weight loss period. Analysis of hay is recommended to ensure that the nonstructural carbohydrate (NSC) content of the forage is low. Hay with NSC content less than 10% on a dry matter basis is ideal for insulin-resistant horses. Hay can be soaked for 1 h in cold water to reduce NSC content; however, this strategy does not reliably reduce NSC content below 10% and may not be adequate when managing an insulin-resistant animal (Longland et al. 2009). Obese horses should also be exercised because exercise induces weight loss (Carter et al. 2010) and is likely to lower appetite and improve insulin sensitivity. A horse with EMS should ideally be housed in a small paddock 0.25 to 0.5 acre so that there is sufficient room for exercise. With the exception of severely affected animals with recurrent laminitis, horses with EMS can be housed in grass paddocks, as long as a grazing muzzle is used to restrict grass consumption. One additional benefit of grazing muzzles is that the horse or pony exercises more as it works harder to graze on grass. Interacting with a companion horse in the same paddock also increases exercise. Refer to the section on PPID for dietary management of lean insulin-resistant horses. Veterinarians have a responsibility to recommend management changes and discourage horse owners from administering drugs as a substitute. However, there are 2 indications for pharmacological intervention: 1) short-term (3 6 months) treatment while management changes are taking effect and 2) refractory cases. Two drugs have been examined to date. Levothyroxine sodium When administered at high dosages, levothyroxine induces weight loss in horses and is accompanied by increased insulin sensitivity (Sommardahl et al. 2005; Frank et al. 2008a,b). Pretreatment with levothyroxine for 14 days has also been shown to prevent horses from developing IR following endotoxin infusion (Tóth et al. 2010). Levothyroxine has been administered at an approximate dosage of 0.1 mg/kg bwt, which is rounded to 48 mg/day for horses weighing kg. It is assumed that levothyroxine induces weight loss by raising circulating thyroxine concentrations and stimulating basal metabolic rate. Weight loss is enhanced by restricting caloric intake and increasing exercise at the same time that levothyroxine is administered. Horses should not be permitted to graze on pasture because levothyroxine is likely to induce hyperphagia, which offsets the effects of treatment. Levothyroxine is primarily administered for the purpose of accelerating weight loss in obese horses and is prescribed for 3 6 months while other management practices are instituted. This drug is reasonably priced in the United States but is considerably more expensive in Europe. Metformin hydrochloride Metformin has been used for decades in human medicine. This biguanide drug is administered to control hyperglycaemia and increase tissue insulin sensitivity in human with diabetes mellitus. It suppresses hepatic glucose production by activating AMP-activated protein kinase (AMPK), which inhibits gluconeogenesis and lipogenesis while increasing fatty acid oxidation and lipolysis (Kim et al. 2008). Two key gluconeogenesis enzymes, phosphoenolpyruvate carboxykinase and glucose-6-phosphatase are inhibited by metformin through this mechanism. The insulin-sensitising effects of metformin may also be mediated by skeletal muscle AMPK, causing increased GLUT4 abundance within cell membranes and enhanced glucose uptake (Musi et al. 2002). One study also describes AMPK-independent effects of metformin on cardiac muscle, with results indicating that p38 mitogen-activated protein kinase and protein kinase C pathways are activated (Saeedi et al. 2008). Only a small number of studies have been performed to explore the use of metformin in horses. Durham et al. (2008) reported that resting insulin concentrations and proxy measures of insulin sensitivity improved in horses and ponies with presumed insulin resistance when treated with metformin at a dosage of 15 mg/kg bwt per os q. 12 h. Administration of metformin at this dosage was associated with positive clinical outcomes, but a subsequent study evaluating the same dose did not demonstrate improved insulin sensitivity or changes in indices of glucose and insulin dynamics in nonobese insulin-resistant ponies (Tinworth et al. 2011). Hustace et al. (2009) reported that the oral bioavailability of 3 g metformin was % in fasted horses and % in fed animals and both trough and

9 160 Endocrine disorders and laminitis predicted peak concentrations were lower in ponies receiving metformin at 15 mg/kg bwt per os q. 12 h for 21 days than values associated with therapeutic efficacy in human (Tinworth et al. 2010). The current recommendation for metformin is, therefore, 30 mg/kg bwt per os q h. Response to treatment Weight loss can be monitored using a weight tape or walk-on scale and neck circumference measurements may be useful for assessing improvement in regional adiposity. Fasting blood glucose and insulin concentrations and dynamic test results should return to normal with treatment. However, there are some patients that remain insulin-resistant in the face of management changes and medical treatments. These animals remain at higher risk for laminitis and every effort should be made to avoid triggers for this disease. Severely affected horses and ponies should be held off pasture and housed in dry lots or small grass paddocks. Type 2 diabetes mellitus is a concern because glucosuria causes electrolyte depletion and anorexia. Diabetic horses are also more susceptible to hypertriglyceridaemia when lipids are mobilised in response to stress and negative energy balance. Horses with diabetes mellitus that are in good general heath should be managed with dietary changes and administration of metformin, whereas patients in crisis require exogenous insulin. Short-acting regular (soluble) insulin can be administered i.v. via constant rate infusion or long-acting insulin can be given by subcutaneous injection. The decision to administer insulin is based upon the degree of glucosuria and hypertriglyceridaemia. Diabetes mellitus is sometimes associated with PPID and pergolide should be administered to these patients. Future directions The endocrine disorders discussed increase the risk of laminitis and diagnostic testing should be performed to detect these conditions. Body condition scoring should also be conducted as part of biannual wellness evaluations and owners must recognise that obesity threatens the health of their horse. It is relatively easy to diagnose endocrine disorders in advanced cases but greater emphasis should be placed upon identifying and treating endocrinopathies in their earlier stages. Research should focus upon the development of more sensitive screening tests for EMS and PPID in horses, with the aim of preventing laminitis. Authors declaration of interests Dr Frank is a consultant for Boehringer Ingelheim, manufacturer of pergolide. The trade name and product name are not mentioned. References Asplin, K.E., Sillence, M.N., Pollitt, C.C. and McGowan, C.M. (2007) Induction of laminitis by prolonged hyperinsulinaemia in clinically normal ponies. Aust. vet. J. 174, Bailey, S.R., Habershon-Butcher, J.L., Ransom, K.J., Elliott, J. and Menzies-Gow, N.J. 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