Anna Mykkänen

Plasma levels of heat shock protein 72 (HSP72) and β-endorphin as indicators of stress, pain and prognosis in horses with colic.

A prospective observational study was performed to evaluate whether the plasma concentration of heat shock protein 72 (HSP72) or beta-endorphin is related to clinical signs, blood chemistry, or severity of pain of colic. Seventy-seven horses with colic and 15 clinically healthy controls were studied. The horses were divided into four groups which reflected increasing severity of colic, from normal control horses to horses with mild, moderate and severe colic. Blood samples were collected before any treatment. Packed cell volume (PCV) and plasma HSP72, beta-endorphin, cortisol, adrenocorticotropic hormone (ACTH) and lactate concentrations were measured. Plasma beta-endorphin was related with severity of colic and survival, as well as with plasma cortisol, ACTH and lactate concentrations, heart rate, PCV and pain score. High plasma HSP72 concentration may indicate circulatory deficits, but was not associated with clinical signs of colic. Plasma lactate still seemed to be the most useful single prognostic parameter in horses with colic.

Muscle fibre type distribution of the thoracolumbar and hindlimb regions of horses: relating fibre type and functional role

BackgroundAlthough the majority of equine muscles have a mixed fibre type distribution indicative of diverse functional roles, the predominance of a fibre type can indicate the primary function of a muscle. The deep epaxial musculature has an important role in core spinal stability in humans, reflected as a predominantly muscle fibre type (MFT) I or postural fibre type. The fibre type of the deep epaxial musculature has not been determined in horses. The objective of the study was to determine the MFT distribution in selected muscles of thoracolumbar and hindlimb region of horses. This included deep epaxial and hypaxial muscles that were hypothesised to have a postural stabilising role. A second objective was to examine differences in MFT distribution between horses bred for endurance (Arabian) and sprinting (Quarter horse). Muscle biopsy samples were obtained from selected thoracolumbar and hind limb muscles of 5 Quarter horses, 4 Arabians, and 2 Thoroughbreds. The myosin heavy chain distribution was determined by gel electrophoresis. Mann–Whitney rank test was used to compare the proportional MFT and differences between breeds.ResultsMm. sacrocaudalis dorsalis medialis and diaphragm had the highest proportion of MFT-I. The remaining deep epaxial muscles and the hypaxial muscle m. psoas minor had approximately equal MFT I and II proportions. Mm. psoas major, iliocostalis, longissimus dorsi and the hind limb muscles contained mostly MFT-IIX. The fibre type distribution was similar between Arabians and Quarter horses, although Quarter horses had more MFT-IIX fibres in psoas major (P = 0.02) while Arabians had more MFT-I fibres in m. longissimus dorsi (P = 0.03).ConclusionsThe fibre type distribution of the deep epaxial muscles, mm psoas minor and diaphragm varied from approximately equal MFT-I and II proportions to predominantly MFT-I suggesting a postural stabilising role possibly important in core spinal stability. In contrast the fibre type proportions of mm psoas major, iliocostalis, longissimus dorsi and the hind limb muscles were mainly MFT-II suggesting a locomotory role. Knowledge of fibre type distribution in such a clinically important area can direct diagnosis, prevention and treatment of muscular or neuromotor dysfunction.

Immunohistochemical analysis of MCT1 and CD147 in equine skeletal muscle fibres

Monocarboxylate transporter 1 (MCT1) and its ancillary protein CD147 facilitate efflux of lactate from the muscle. Expression of MCT1 and CD147 were studied with immunohistochemistry in type I, IIA, IIAB and IIB fibres of equine gluteal muscle. Staining intensity of MCT1 in the cytoplasm as well as in the membranes of fibre types decreased in the order I=IIA>IIAB>IIB and correlated with the oxidative capacity. Capillaries were pronounced in the MCT1 staining. CD147 antibody stained plasma membranes of all fibre types evenly, whereas the staining in the cytoplasm followed that of MCT1. In the middle gluteal muscle the expression of MCT1 follows the oxidative capacity of muscle fibres, but the expression of CD147 in sarcolemma does not vary among fibre types. The use of horse specific MCT1 and CD147 antibodies can in future studies help to evaluate lactate efflux from different muscle fibre types.

Generalised tetanus in a 2-week-old foal: use of physiotherapy to aid recovery.

A 2-week-old Estonian Draft foal presented with signs of severe generalised tetanus, recumbency and inability to drink. The suspected source of infection was the umbilicus. Medical treatment was administered, including tetanus antitoxin, antimicrobial therapy and phenobarbital to control tetanic spasms. In addition, an intensive physiotherapy program was carried out during the recovery period. Techniques designed for syndromes involving upper motor neuron spasticity in humans were applied. Exercises aimed at weight-bearing and mobility were executed with the help of a walking-frame. The foal made a complete recovery. To our knowledge, this is the first report of the use of physiotherapy in the treatment of tetanus in horses.

MCT1, MCT4 and CD147 gene polymorphisms in healthy horses and horses with myopathy.

Polymorphisms in human lactate transporter proteins (monocarboxylate transporters; MCTs), especially the MCT1 isoform, can affect lactate transport activity and cause signs of exercise-induced myopathy. Muscles express MCT1, MCT4 and CD147, an ancillary protein, indispensable for the activity of MCT1 and MCT4. We sequenced the coding sequence (cDNA) of horse MCT4 for the first time and examined polymorphisms in the cDNA of MCT1, MCT4 and CD147 of 16 healthy horses. To study whether signs of myopathy are linked to the polymorphisms, biopsy samples were taken from 26 horses with exercise-induced recurrent myopathy. Two polymorphisms that cause a change in amino acid sequence were found in MCT1 (Val(432)Ile and Lys(457)Gln) and one in CD147 (Met(125)Val). All polymorphisms in MCT4 were silent. Mutations in MCT1 or CD147 in equine muscle were not associated with myopathy. In the future, a functional study design is needed to evaluate the physiological role of the polymorphisms found.

Expression of monocarboxylate transporters I and IV and the ancillary protein CD147 in the intestinal tract of healthy horses and ponies

OBJECTIVE To characterize the expression of monocarboxylate transporters (MCTs) 1 and 4 and the ancillary protein CD147 in the intestinal tract of healthy equids and determine the cellular location of CD147 in the intestinal epithelium. ANIMALS 12 healthy horses and ponies slaughtered for meat production or euthanized for reasons unrelated to gastrointestinal tract disease. PROCEDURES The entire gastrointestinal tract was removed from each equid within 45 minutes after slaughter or euthanasia. Tissue samples were obtained from the antimesenteric side of the duodenum, jejunum, ileum, middle part of the cecum, sternal flexure of the ventral colon, pelvic flexure, sternal flexure of the dorsal colon, and descending colon (small colon). Expressions of MCT1, MCT4, and the ancillary protein CD147 were examined in tissue samples from each of the 8 intestinal locations by means of quantitative PCR assay, immunoblotting, and immunohistochemical analyses. RESULTS Expression of MCT1 was most abundant in the cecum and colonic sites, whereas expression of MCT4 was predominantly in the proximal section of the intestine (small intestinal sites and cecum). Immunohistochemical analysis revealed that MCT1 and CD147 were present in the membranes of enterocytes (in crypts and villi). CONCLUSIONS AND CLINICAL RELEVANCE The anatomic distribution of MCT1 and MCT4 in the equine intestinal tract determined in this study together with the previous knowledge of the sites of substrate absorption indicated that MCT1 might predominantly contribute to the uptake of short-chain fatty acids in the large intestine and MCT4 might predominantly contribute to the uptake of lactate in the small intestine.

Expression of extracellular matrix metalloproteinase inducer and matrix metalloproteinase-2 and −9 in horses with chronic airway inflammation

OBJECTIVE To examine whether expression of extracellular matrix metalloproteinase inducer (EMMPRIN) can be detected in equine lungs and whether it correlates with matrix metalloproteinase (MMP)-2 and -9 expression in bronchoalveolar lavage fluid (BALF) of horses with chronic inflammation of the lungs (ie, lower airway inflammation [LAI]). ANIMALS 29 horses with signs of chronic respiratory tract disease, which were classified as the LAI (n = 17) and LAI with respiratory distress (RDLAI [12]) groups, and 15 control horses. PROCEDURES BALF, tracheal aspirate, and blood samples were obtained, and EMMPRIN expression was determined from BALF cells and RBCs by use of western blotting. Activities of MMP-2 and -9 were determined with zymography. RESULTS Expression of EMMPRIN protein was identified in BALF cells of all horses. Expression of EMMPRIN protein was highest for the RDLAI group and was correlated with MMP-2 and -9 protein expression, MMP-9 gelatinolytic activity, and airway neutrophilia. CONCLUSIONS AND CLINICAL RELEVANCE Results suggested that EMMPRIN was involved in the pathophysiologic processes of asthma in horses. However, additional studies of horses and other species are warranted to elucidate the regulation of EMMPRIN expression in asthmatic lungs.

Cardiopulmonary effects of vatinoxan in sevoflurane-anaesthetised sheep receiving dexmedetomidine

The effects of pre-treatment with vatinoxan (MK-467) on dexmedetomidine-induced cardiopulmonary alterations were investigated in sheep. In a crossover study design with a 20-day washout, seven sheep were anaesthetised with sevoflurane in oxygen and air. The sheep were ventilated with the pressure-limited volume-controlled mode and a positive end-expiratory pressure of 5cmH2O. Peak inspiratory pressure (PIP) was set at 25cmH2O. The sheep received either 150μg/kg vatinoxan HCl (VAT+DEX) or saline intravenously (IV) 10min before IV dexmedetomidine HCl (3μg/kg, DEX). Cardiopulmonary variables were measured before treatments (baseline), 3min after vatinoxan or saline, and 5, 15 and 25min after dexmedetomidine. Computed tomography (CT) of lung parenchyma was performed at baseline, 2min before dexmedetomidine, and 10, 20 and 30min after DEX. Bronchoalveolar lavage (BAL) was performed after the last CT scan and shortly before sheep recovered from anaesthesia. After VAT, cardiac output significantly increased from baseline. DEX alone significantly decreased partial arterial oxygen tension, total dynamic compliance and tidal volume, whereas PIP was significantly increased. With VAT+DEX, these changes were minimal. No significant changes were detected in haemodynamics from baseline after DEX. With VAT+DEX, mean arterial pressure and systemic vascular resistance were significantly decreased from baseline, although hypotension was not detected. On CT, lung density was significantly increased with DEX as compared to baseline. No visual abnormalities were detected in bronchoscopy and no differences were detected in the BAL fluid after either treatment. The pre-administration of vatinoxan alleviates dexmedetomidine-induced bronchoconstriction, oedema and hypoxaemia in sevoflurane-anaesthetised sheep.

Comparison of Tracheal Wash and Bronchoalveolar Lavage Cytology in 154 Horses With and Without Respiratory Signs in a Referral Hospital Over 2009−2015

Most equine lower respiratory diseases present as increased airway neutrophilia, which can be detected in tracheal wash (TW) or bronchoalveolar lavage fluid (BALF) cytology samples. The aim was to compare the TW and BALF results in a population of client-owned horses with and without clinical respiratory disease signs. A secondary aim was to determine the sensitivity (Se) and specificity (Sp) of TW and BALF neutrophilia in detecting respiratory disease. The cutoff values for neutrophils were also evaluated. Retrospective data from 154 horses of various breeds that had been subject to both TW and bronchoalveolar lavage (BAL) sampling at rest during 2009−2015 were used. The horses were divided into three groups based on the presenting signs, physical examination, and endoscopy mucus score. Neutrophil counts of >20% in TW and >5% in BAL were considered abnormal. Cytology results between groups, correlations between TW and BALF cell types, and tracheal mucus score were analyzed. Two graph receiving operating characteristic (ROC) curves of the neutrophil percentage values of TW and BALF were created to determine the optimal cutoff values and to calculate the diagnostic Se and Sp for diagnosing airway inflammation in horses with and without clinical respiratory signs. The Se and Sp of TW and BALF neutrophil percentages were further estimated using a two-test one-population Bayesian latent class model. The two tests showed substantial agreement, and only 17.5% of the horses were classified differently (healthy vs. diseased). The neutrophil percentage was found to correlate between TW and BALF. The Se and Sp of TW were generally higher than for BAL when estimated with area under the curve or Bayesian model. Cutoff values of 17.7% for TW and 7% for BALF were indicated by the ROCs. We conclude that TW is a more sensitive and specific method in our patient population. We suggest that the current neutrophil cutoff values of 20% for TW and 5% for BALF would still be appropriate to use in clinical diagnosis of airway inflammation. However, further studies with other cell types and in other populations are warranted to determine the best sampling method for individual horses.

Effects of MK-467 hydrochloride and hyoscine butylbromide on cardiorespiratory and gastrointestinal changes induced by detomidine hydrochloride in horses

OBJECTIVE To compare the effects of MK-467 and hyoscine butylbromide on detomidine hydrochloride-induced cardiorespiratory and gastrointestinal changes in horses. ANIMALS 6 healthy adult horses. PROCEDURES Horses received detomidine hydrochloride (20 μg/kg, IV), followed 10 minutes later by MK-467 hydrochloride (150 μg/kg; DET-MK), hyoscine butylbromide (0.2 mg/kg; DET-HYO), or saline (0.9% NaCl) solution (DET-S), IV, in a Latin square design. Heart rate, respiratory rate, rectal temperature, arterial and venous blood pressures, and cardiac output were measured; blood gases and arterial plasma drug concentrations were analyzed; selected cardiopulmonary variables were calculated; and sedation and gastrointestinal borborygmi were scored at predetermined time points. Differences among treatments or within treatments over time were analyzed statistically. RESULTS With DET-MK, detomidine-induced hypertension and bradycardia were reversed shortly after MK-467 injection. Marked tachycardia and hypertension were observed with DET-HYO. Mean heart rate and mean arterial blood pressure differed significantly among all treatments from 15 to 35 and 15 to 40 minutes after detomidine injection, respectively. Cardiac output was greater with DET-MK and DET-HYO than with DET-S 15 minutes after detomidine injection, but left ventricular workload was significantly higher with DET-HYO. Borborygmus score, reduced with all treatments, was most rapidly restored with DET-MK. Sedation scores and pharmacokinetic parameters of detomidine did not differ between DET-S and DET-MK. CONCLUSIONS AND CLINICAL RELEVANCE MK-467 reversed or attenuated cardiovascular and gastrointestinal effects of detomidine without notable adverse effects or alterations in detomidine-induced sedation in horses. Further research is needed to determine whether these advantages are found in clinical patients and to assess whether the drug influences analgesic effects of detomidine.