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.

Blood glucose concentrations in critically ill neonatal foals.

REASONS FOR PERFORMING STUDY: Critical illness is associated with hyperglycemia in humans, and a greater degree and duration of hyperglycemia is associated with nonsurvival. Hypoglycemia is also seen in critically ill humans, and is associated with nonsurvival. This might also be true in the critically ill foal. OBJECTIVES: To investigate the association of blood glucose concentrations with survival, sepsis, and the systemic inflammatory response syndrome (SIRS). METHODS: Blood glucose concentrations at admission (515 foals) and 24 hours (159 foals), 36 hours (95), 48 hours (82), and 60 hours (45) after admission were analyzed. Logistic regression analyses were performed to investigate the association of glucose concentrations with survival, sepsis, a positive blood culture, or SIRS. RESULTS: 29.1% of foals had blood glucose concentrations within the reference range (76-131 mg/dL) at admission, 36.5% were hyperglycemic, and 34.4% were hypoglycaemic. Foals that did not survive to hospital discharge had lower mean blood glucose concentrations at admission, as well as higher maximum and lower minimum blood glucose concentrations in the 1st 24 hours of hospitalization, and higher blood glucose at 24 and 36 hours. Foals with blood glucose concentrations <2.8 mmol/L (50 mg/dL) or >10 mmol/L (180 mg/dL) at admission were less likely to survive. Hypoglycemia at admission was associated with sepsis, a positive blood culture, and SIRS. CONCLUSIONS AND POTENTIAL RELEVANCE: Derangements of blood glucose concentration are common in critically ill foals. Controlling blood glucose concentrations may therefore be beneficial in the critically ill neonatal foal, and this warrants further investigation.

Prophylactic administration of ponazuril reduces clinical signs and delays seroconversion in horses challenged with Sarcocystis neurona.

The ability of ponazuril to prevent or limit clinical signs of equine protozoal myeloencephalitis (EPM) after infection with Sarcocystis neurona was evaluated. Eighteen horses were assigned to 1 of 3 groups: no treatment, 2.5 mg/kg ponazuril, or 5.0 mg/kg ponazuril. Horses were administered ponazuril, once per day, beginning 7 days before infection (study day 0) and continuing for 28 days postinfection. On day 0, horses were stressed by transport and challenged with 1 million S. neurona sporocysts per horse. Sequential neurologic examinations were performed, and serum and cerebrospinal fluid were collected and assayed for antibodies to S. neurona. All horses in the control group developed neurologic signs, whereas only 71 and 40% of horses in the 2.5 and 5.0 mg/kg ponazuril groups, respectively, developed neurologic abnormalities. This was significant at P = 0.034 by using Fisher exact test. In addition, seroconversion was decreased in the 5.0 mg/kg group compared with the control horses (100 vs. 40%; P = 0.028). Horses with neurologic signs were killed, and a post-mortem examination was performed. Mild-to-moderate, multifocal signs of neuroinflammation were observed. These results confirm that treatment with ponazuril at 5.0 mg/kg minimizes, but does not eliminate, infection and clinical signs of EPM in horses.

Equine Protozoal Myeloencephalitis: An Updated Consensus Statement with a Focus on Parasite Biology, Diagnosis, Treatment, and Prevention.

Equine protozoal myeloencephalitis (EPM) remains an important neurologic disease of horses. There are no pathognomonic clinical signs for the disease. Affected horses can have focal or multifocal central nervous system (CNS) disease. EPM can be difficult to diagnose antemortem. It is caused by either of 2 parasites, Sarcocystis neurona and Neospora hughesi, with much less known about N. hughesi. Although risk factors such as transport stress and breed and age correlations have been identified, biologic factors such as genetic predispositions of individual animals, and parasite-specific factors such as strain differences in virulence, remain largely undetermined. This consensus statement update presents current published knowledge of the parasite biology, host immune response, disease pathogenesis, epidemiology, and risk factors. Importantly, the statement provides recommendations for EPM diagnosis, treatment, and prevention.

Arterial lactate concentration, hospital survival, sepsis and SIRS in critically ill neonatal foals.

REASONS FOR PERFORMING STUDY: Blood lactate concentration has been shown to be a useful clinical indicator in human patients, but has not been formally investigated in critically ill foals. OBJECTIVE: To investigate the association of blood lactate with hospital survival, markers of cardiovascular status, metabolic acid base status, sepsis and systemic inflammatory response syndrome (SIRS). METHODS: A database containing clinical, haematological, plasma biochemical and hospital outcome data on neonatal foals referred to an intensive care unit in 2000-2001 was analysed. Seventy-two foals for which arterial lactate was measured at admission were included in the study. RESULTS: Sixty-one foals had an admission lactate concentration > 2.5 mmol/l. Admission lactate was statistically associated with hospital survival, mean arterial pressure, blood creatinine concentration, bacteraemia, anion gap, lactate concentration at 18-36 h after admission and evidence of SIRS, but not with packed cell volume or heart rate. Lactate at 18-36 h was also associated with survival and evidence of SIRS. Anion gap, base excess, base excess due to unidentified anions (BEua), simplified strong ion gap or bicarbonate correctly classified foals for presence of hyperlactaemia (> 5 mmol/l) in < or = 80% of animals. CONCLUSIONS: Admission blood lactate gives important prognostic information. Lactate should be measured rather than assumed from the anion gap, base excess, BEua, simplified strong ion gap or bicarbonate. POTENTIAL RELEVANCE: Blood lactate concentrations at admission are clinically relevant in neonatal foals and warrant further investigation. This should include the clinical value of measuring changes in lactate in response to treatment.

Equine endotoxaemia--a state-of-the-art review of therapy.

The pathophysiology of endotoxaemia, a leading cause of death in the horse, is beginning to be understood in greater detail. Endotoxin may be absorbed into the systemic circulation in a number of different ways: most commonly the body's normal defense mechanisms are disrupted or bypassed, or the normal clearance mechanisms overwhelmed. Following this wide-spread effects are observed, although the most significant are seen in the cardiovascular system. Fever, arterial hypoxaemia and signs of abdominal pain are also common. With increased understanding of the disease new therapeutic agents have become available, however, while the newer agents offer some advantages it is important to recognise that supportive care is the mainstay of treatment for endotoxaemia. Supportive care consists of aggressive fluid therapy (crystalloid, colloid and hypertonic), the administration of non-steroidal antiinflammatory drugs and, where appropriate, antimicrobials. The principles of supportive care are discussed in detail. Other therapies such as hyperimmune plasma, polymyxin B, pentoxifylline, dimethyl sulfoxide and heparin are commonly used in the treatment of equine endotoxaemia and their use is reviewed here. Furthermore, newer agents such as anti-tumour necrosis factor antibodies, detergent, activated protein C and insulin, which have yet to gain widespread acceptance but may have an important role in the treatment of endotoxaemia in the future, are examined.

Glucocorticoid receptor density and binding affinity in healthy horses and horses with systemic inflammatory response syndrome.

BACKGROUND: Dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis occurs in horses with systemic inflammatory response syndrome (SIRS). Peripheral resistance to glucocorticoids has not been investigated in horses. OBJECTIVE: To determine if glucocorticoid receptor (GR) function in horses can be measured using flow cytometry, and to use this information to evaluate HPA axis dynamics. ANIMALS: Eleven healthy adult horses in parts 1 and 2. Ten horses with SIRS and 10 age and sex matched controls in part 3. METHODS: Flow cytometry was used to evaluate GR density and binding affinity (BA) in 3 healthy horses in part 1. In part 2, exogenous ACTH was administered to eight healthy horses. Their cortisol response and GR properties were measured. In part 3, CBC, serum biochemistry, cortisol and ACTH, and GR properties were compared between controls without SIRS (n = 10) and horses with SIRS (n = 10), and between survivors and nonsurvivors (n = 4 and n = 6 respectively). RESULTS: Flow cytometry can be used to measure GR properties in equine PBMCs. No correlation was observed between plasma cortisol concentration and GR density or BA in healthy horses (r = -0.145, P = .428 and r = 0.046, P = .802 respectively). Nonsurvivors with SIRS had significantly decreased GR BA (P = .008). Horses with triglyceride concentration > 28.5 mg/dL had increased odds of nonsurvival (OR=117; 95% CI, 1.94-7,060). GR BA <35.79% was associated with nonsurvival (OR = 30.33; 95% CI, 0.96-960.5). CONCLUSIONS AND CLINICAL IMPORTANCE: Tissue resistance to glucocorticoids contributes to HPA axis dysfunction in adult horses with SIRS. These horses might benefit from treatment with exogenous glucocorticoids.

An update on Sarcocystis neurona infections in animals and equine protozoal myeloencephalitis (EPM).

Equine protozoal myeloencephalitis (EPM) is a serious disease of horses, and its management continues to be a challenge for veterinarians. The protozoan Sarcocystis neurona is most commonly associated with EPM. S. neurona has emerged as a common cause of mortality in marine mammals, especially sea otters (Enhydra lutris). EPM-like illness has also been recorded in several other mammals, including domestic dogs and cats. This paper updates S. neurona and EPM information from the last 15 years on the advances regarding life cycle, molecular biology, epidemiology, clinical signs, diagnosis, treatment and control.

Immunologic studies of a horse with lymphosarcoma.

Immunological, clinical, and pathological investigations were conducted on a horse with lymphosarcoma. The immunological status was investigated by measuring the level of antibodies by single radial immunodiffusion test and the ability of lymphocytes to proliferate in response to mitogens. Multiple immunological abnormalities were noted in this horse. They were; (1) decreased IgM, IgG, and IgA levels in the serum despite hyperproteinemia; (2) increased in-vitro spontaneous lymphoproliferation which reflects augmented mitosis; (3) decreased lymphoproliferative response to T cell stimulants (e.g. Concanavalin-A (Con-A)) suggesting impaired T cell activation; (4) presence of immunosuppressive factors in serum as demonstrated by in-vitro lymphocyte culture systems. Clinical pathology findings revealed an unusual monoclonal alpha peak in the serum and morphologically abnormal lymphocytes distributed throughout the body. Serum fractionated by fast protein liquid chromatography (FPLC) revealed that the immunosuppressive factors were found in this abnormal alpha peak. The immunopathological findings in this horse are discussed.

Transforming growth factor beta concentrations and interferon gamma responses in cerebrospinal fluid of horses with equine protozoal myeloencephalitis.

The following experiment was performed to test the hypothesis that transforming growth factor beta (TGF-beta) concentration varies in the cerebrospinal fluid and serum of horses with EPM and to determine if cerebrospinal fluid (CSF) alters the interferon-gamma (IFN-gamma) rersponse of equine peripheral blood mononuclear cells (PBMCs). The concentration of transforming growth factor-beta (TGF-beta2) was investigated in the serum and cerebrospinal fluid (CSF) of 18 horses (9 normal, 9 affected with equine protozoal myeloencephalitis [EPM]). The TGF-beta2 assay was validated in a group of 6 normal horses. Intra-assay variability was 4.7%, and interassay variability was 10.7%. The slope of the curve of the unknown samples of various volumes demonstrated parallelism with a curve developed using equal volumes of assay kit standard. Assay of normal and EPM-affected horses found that TGF-beta2 was present in both the serum and CSF of all animals. However, the concentration of TGF-beta2 in the CSF was less (P = 0.03) in EPM-affected horses (144 pg/ml) than in normal horses (256 pg/ml). In addition, the effect of CSF from normal and EPM-affected horses on the production of interferon-gamma (IFN-gamma) by PHA-P stimulated PBMCs from normal horses was investigated using a bioassay. It was found that CSF from normal and EPM-affected horses enhanced IFN-gamma activity from PHA-P stimulated peripheral blood mononuclear cells (P < or = 0.05); however, the response to CSF from EPM-affected horses was no different than the response to CSF from normal horses. Treatment of cells with anti-TGF-beta2 monoclonal antibodies slightly increased the response when co-incubated with CSF from normal horses, and slightly decreased it when co-incubated with CSF from EPM-affected horses. These differences, however, did not achieve statistical significance (P > 0.05). Results of this study indicated that production of TGF-beta2 is altered in horses with EPM, and that CSF appears to contain substances which alter the inflammatory reaction to plant lectins. These findings confirm the immunomodulatory properties of CSF and suggest new techniques for future research regarding the pathophysiology of EPM.

The effects of stress on gastric ulceration, T3, T4, reverse T3 and cortisol in neonatal foals.

Sixteen neonatal foals stressed by disease underwent endoscopic examination of their stomachs and blood was assayed for triiodothyronine (T3), reverse T3 (rT3), thyroxine (T4) and cortisol, to determine the effects of severe physiological stress and the occurrence of gastric ulcers. compared with eight age-matched controls, six foals had abnormal cortisol, seven had abnormal T3 and 12 had abnormal T4. Eleven of 13 foals had rT3 outside the 95 per cent confidence interval for clinically normal foals of comparable ages. Gastric lesions were seen more frequently in stressed foals, and gastric glandular mucosal lesions were noted in 40 per cent of the stressed foals. Previous studies report low (3 per cent) occurrence of gastric mucosal lesions. The frequency of squamous mucosal lesions was not different from that reported previously, indicating that stress has little effect on the development of lesions at this site.

Comparison of lithium dilution and thermodilution cardiac output measurements in anaesthetised neonatal foals.

Knowledge of cardiac output is expected to help guide the treatment of hypotension associated with critical illness and/or anaesthesia in neonatal foals. However, a practical and safe method of measuring cardiac output has not been described for the foal. Lithum dilution, a new method of cardiac output determination not requiring cardiac catheterisation, has recently been reported in mature horses. We compared this method to thermodilution in isoflurane-anaesthetised foals age 30-42 h and found good agreement between the 2 methods in a range of cardiac outputs 5.4-20.4 l/min. The lithium dilution technique is a practical and reliable method of measuring cardiac output in anaesthetised neonatal foals, and warrants investigation in critically ill conscious foals.

Prognosis for neonatal foals in an intensive care unit.

This study was conducted to develop an equation for the prediction of outcome in neonatal foals undergoing treatment in an intensive care unit (ICU). Fifty-three physical examination, historical, and clinicopathologic variables were analyzed from the records of 99 neonatal foals (< 14 days of age) treated in the neonatal ICU of the Equine Medical Center. The outcome was recorded and the results were categorized into either surviving or nonsurviving groups. The mean values for the 2 groups were compared, and variables that differed significantly between the two groups were retained and used to construct a logistic regression equation. Retained variables were heart rate, temperature, and neutrophil count. The predictive equation then was tested prospectively in 2 additional groups of foals from 2 separate ICUs. The predicted outcome was compared to the actual outcome, and performance variables were calculated. Sensitivity (.83), specificity (.87), negative predictive value (.72), and positive predictive value (.93) were determined for foals from one neonatal ICU; the sensitivity (.83), specificity (.44), negative predictive value (.44), and positive predictive value (.83) were lower for foals at a second, separate ICU.

Neonatal equine herpesvirus type 1 infection on a thoroughbred breeding farm.

Of 17 foals born on a Thoroughbred breeding farm between March and April 1995, infection with equine herpesvirus type 1 (EHV-1) was associated with neonatal morbidity in 5 foals, 3 of which died or were euthanized. Morbidity and mortality were associated with pulmonary inflammation, and EHV-1 was identified in the lungs of the 3 foals that died. All neonatal EHV-1 infections occurred in foals of mares housed in the same pasture and barn. No other clinical manifestations of EHV-1 infection (e.g., abortion, neurologic disease, or respiratory disease) occurred during this outbreak. Three foals were treated with acyclovir (1 died, 2 survived), which may have influenced the clinical outcome in the surviving foals.

Diagnostic validity of electroencephalography in equine intracranial disorders.

Electroencephalography (EEG) is a valuable diagnostic test to identify functional disturbances in brain activity. The purpose of this study was to assess the validity of EEG as a diagnostic indicator of intracranial diseases in horses. The validity of EEG was estimated by comparing clinical, clinicopathologic, and histopathologic findings to EEG findings in 20 horses examined for seizures. collapse, or abnormal behavior between 1984 and 1997. A bipolar left-to-right, back-to-front montage and a bipolar circular montage were recorded from sedated (4) and anesthetized (16) horses. Visual and semiquantitative masked analysis of EEG recording Ist was validated on 10 horses presented for problems other than intracranial diseases. EEG pattern was normal in 7 of the 20 clinically affected horses. Abnormal EEG patterns included high-voltage slow waves and discrete paroxysmal activity with or without generalized activity in 13 horses. Histopathologic diagnoses in 10 horses included meningoencephalitis, neuronal necrosis, congenital anomalies. cerebral edema. and abscess. All of these horses had abnormal EEG patterns (sensitivity, 100%) with a positive neuroanatomic correlation in 7 animals. Localization of histopathologic and EEG abnormalities did not correlate in 15% of the horses (3/20). The cause of neurologic signs could not be explained at postmortem examination in 10 animals and the EEG pattern was normal in 7 of these horses (specificity, 70%). In conclusion, equine EEG was a sensitive tool in the diagnosis of intracranial disorders.

Cerebrospinal fluid variables in clinically normal foals from birth to 42 days of age.

To determine the effects of age on each analyte, CSF variables were evaluated in healthy foals from birth through 42 days of age. Cerebrospinal fluid was collected from 14 clinically normal, naturally delivered cross-bred foals and was analyzed for glucose, sodium, potassium, magnesium, and total protein concentrations, total and differential WBC counts, RBC count, and lactate dehydrogenase, aspartate transaminase, and creatine kinase activities. Samples were collected in 3 foals < 48 hours old, and at 11 to 14 days of age in 4 foals, 21 to 22 days of age in 3 foals, and 31 to 42 days of age in 4 foals. Each foal was tested only once, to avoid any effects of CSF sample collection on subsequent analysis. Regression analysis confirmed age-related effects on CSF glucose, protein, and magnesium concentrations, but did not indicate an effect of age on CSF sodium and potassium concentrations or cell counts. Results indicate that CSF glucose concentration decreases with age; foals < 2 days old had the highest CSF glucose values, 98.8 +/- 12.0 mg/dl (mean +/- 1 SD). In foals 10 to 14 days old, CSF glucose concentration was 67.3 +/- 12.0 mg/dl, was 65.3 +/- 4.5 mg/dl in foals 21 to 22 days old, was 70.0 +/- 5.4 mg/dl in foals 31 to 42 days old, and was 51.1 +/- 2.5 mg/dl in adults.(ABSTRACT TRUNCATED AT 250 WORDS)

High resolution protein electrophoresis of equine cerebrospinal fluid.

OBJECTIVE: To determine normal CSF electrophoresis patterns in horses, and to determine whether the electrophoretic scans from horses with cervical compression differ from those of neurologically normal horses. ANIMALS: 32 horses assigned to 1 of 2 groups: neurologically normal (n = 18) or cervical compression (n = 14). PROCEDURE: CSF was collected from 18 neurologically normal horses referred to the Marion duPont Scott Equine Medical Center, and protein electrophoresis was performed to describe the normal equine CSF electrophoretogram. Results of CSF electrophoresis from 14 horses with cervical compression were then compared with results for the neurologically normal horses. RESULTS: Horses with cervical compression had decreased beta-globulin fraction, and 1 or 2 prominent post-beta 2 peak(s). When the presence of post-beta peaks was used as a diagnostic criterion for cervical compression, the test had sensitivity of 71.4% and specificity of 81.8%. The positive and negative predictive values were 83.3 and 69.2%, respectively. CONCLUSION AND CLINICAL IMPLICATIONS: Electrophoresis of CSF may be a useful diagnostic aid in evaluation of horses with neurologic disease.

Cerebrospinal fluid creatine kinase activity in horses with central nervous system disease: 69 cases (1984-1989).

The CSF creatine kinase (CK) activity was determined in 70 CSF samples from 69 horses with CNS disease. Abnormal values (greater than or equal to 1 IU/L) were determined from 32 CSF samples, and normal values (less than 1 IU/L) were found in 38 samples. Increased CK activity was most frequently associated with a diagnosis of equine protozoal myelitis; CK activity was not increased in 11 horses with cervical compressive myelopathy. Other diagnoses, in which CSF CK activity was increased included trauma (n = 1), idiopathic epilepsy (n = 2), botulism (n = 2), articular facet fracture (n = 1), intervertebral disk protrusion (n = 1), and toxemia (n = 1).

Intervertebral disk prolapse and diskospondylitis in a horse.

Intervertebral disk prolapse was diagnosed in a mature horse with clinical signs of caudal ataxia. Radiography and myelography demonstrated a collapsed intervertebral space and loss of the dorsal and ventral dye columns. Results of CSF analysis were normal, as were a CBC and serum biochemical profile. High CSF WBC count and high CSF creatine kinase activity were noticed following acute neurologic deterioration. While common in certain breeds of dogs, intervertebral disk prolapse is rarely reported in horses. It should be considered in the differential diagnosis of horses with caudal ataxia.

Hepatoencephalopathy and hypocalcemia in a miniature horse mare.

A pregnant, miniature horse mare had clinical signs of hepatoencephalopathy and concurrent hypocalcemia. The signs included dullness, inappetence, blindness, head pressing, weakness, muscle fasiculations, uveitis, and urinary incontinence. Hepatic dysfunction and hypocalcemia were confirmed by serum biochemical analysis. The mare was treated successfully with a continuous infusion of Ringer solution, calcium gluconate, dextrose, B-complex vitamins, sodium ampicillin, and flunixin meglumine; topical administration of ophthalmic ointments; and decompression of the urinary bladder. Histologic examination of a liver biopsy specimen revealed acute hepatic necrosis, which likely was associated with administration of a vaccine of equine origin 2 months earlier.