Systemic Administration of Rejuvenated Adipose-Derived Mesenchymal Stem Cells Improves Liver Metabolism in Equine Metabolic Syndrome (EMS)- New Approach in Veterinary Regenerative Medicine.

Equine metabolic syndrome (EMS) is characterized by adiposity, insulin dysregulation and increased risk for laminitis. Increased levels of specific liver enzymes in the peripheral blood are typical findings in horses diagnosed with EMS. Current management of EMS is based on caloric restriction and increased physical activity. However, new potential treatment options are arising such as the transplantation of autologous adipose stem cells (ASC). However, cytophysiological properties of ASC derived from EMS horses are impaired which strongly limits their therapeutic potential. We hypothesized, that in vitro pharmacotherapy of those cells with 5-azacytidine (AZA) and resveratrol (RES) before their clinical application can reverse the aged phenotype of those cells and improve clinical outcome of autologous therapy. A 9 year old Dutch Warmblood Horse used for driving, was presented with severe obesity, insulin resistance. After EMS diagnosis, the animal received three intravenous injections of autologous, AZA/RES treated ASCs at weekly intervals. The therapeutic effect was assessed by the analysis of liver specific enzymes in the blood. ASC-transplantation reduced levels of glutamate dehydrogenase (GLDH), gamma-glutamyltransferase (GGT), lactate dehydrogenase (LDH) and aspartate transaminase (AST). This case report demonstrates the therapeutic potential of this intervention for EMS as well as apt utility of autologous, rejuvenated ASC injections.

Effects of positive end-expiratory pressure titration on intestinal oxygenation and perfusion in isoflurane anaesthetised horses.

REASONS FOR PERFORMING STUDY: High airway pressures, necessary to keep equine lungs open, can have a detrimental impact on central and peripheral perfusion. OBJECTIVE: The aim of this study was to assess the effects of stepwise increasing airway pressure recruitment on central and intestinal perfusion and oxygenation during isoflurane anaesthesia in horses. STUDY DESIGN: In vivo experimental study. METHODS: Ten anaesthetised horses were ventilated using intermittent positive pressure ventilation immediately after induction. After 90 min, end-expiratory pressure (PEEP) was increased by steps of 5 cmH2 O every 10 min up to a PEEP of 30 cmH2 O and decreased back to zero maintaining a constant airway pressure difference of 20 cmH2 O. Mean arterial blood pressure (MAP), heart rate, central venous pressure, pulmonary artery pressure, expiratory isoflurane concentration and cardiac output (thermodilution method) were measured. Cardiac index (CI) was calculated. Arterial blood gases were taken to measure arterial partial oxygen pressure (PaO2 ) and calculate arterial oxygen saturation (SaO2 ). Intestinal microperfusion and oxygenation were measured by laser Doppler flowmetry and white-light spectrophotometry. After ventral median laparotomy, a probe was placed on the stomach, jejunum and pelvic flexion of the colon. An ANOVA for repeated measurements and Tukey's post hoc test were used for statistical analysis (α = 5%). RESULTS: Recruitment of the lungs resulted in a significant increase in PaO2 from 201 ± 58 mmHg (baseline) to a maximum of 495 ± 75 mmHg. The CI and MAP decreased continuously with increasing airway pressures. When CI and MAP were 37 ± 9 ml/kg/min and 52 ± 8 mmHg (at PEEP of 25 cmH2 O), respectively, a sudden decrease in intestinal perfusion followed by a delayed decrease in oxygenation occurred. CONCLUSIONS: There was linear correlation between airway pressures and CI and MAP but not between central and gastrointestinal perfusion. Despite improvement of arterial oxygenation the decrease in CI and, therefore, in oxygen delivery PEEP resulted in a decrease in gastrointestinal oxygenation.

Temporal and concentration effects of isoflurane anaesthesia on intestinal tissue oxygenation and perfusion in horses.

The aim of this study was to assess the effect of duration of anaesthesia and concentration of isoflurane on global perfusion as well as intestinal microperfusion and oxygenation. Nine Warmblood horses were premedicated with xylazine; anaesthesia was induced with midazolam and ketamine, and maintained with isoflurane. Horses were ventilated to normocapnia. During 7 h of anaesthesia, mean arterial blood pressures (MAP), heart rate, central venous pressure, pulmonary artery pressure, expiratory isoflurane concentration (ETIso) and cardiac output using lithium dilution were measured; cardiac index (CI) was calculated. Intestinal microperfusion and oxygenation were measured using laser Doppler flowmetry and white-light spectrophotometry. Surface probes were placed via median laparotomy on the serosal and mucosal site of the jejunum and the pelvic flexion of the colon. After 3 h of constant ETIso (1.4%), ETIso was increased in 0.2% increments up to 2.4%, followed by a decrease to 1.2% and an increase to 1.4%. The CI and MAP decreased continuously with increasing ETIso to 40 ± 5 mL/kg/min and 52 ± 8 mmHg, respectively. Microperfusion and oxygenation remained unchanged until an ETIso of 2.0% resulted in CI and MAP of 48 ± 5 mL/kg/min and 62 ± 6 mmHg, respectively, and then decreased rapidly. When ETIso decreased back to baseline, CI, MAP, microperfusion and oxygenation recovered to baseline. Isoflurane concentration but not duration of isoflurane anaesthesia influenced central and intestinal oxygenation and perfusion in healthy horses. Under isoflurane, intestinal perfusion appeared to be preserved until a threshold MAP or blood flow was reached.

Three-dimensional anatomy of the equine sternum.

The sternum is a frequently used anatomical site to obtain bone marrow for diagnostic and therapeutic purposes in equine medicine and surgery. For a safe and reproducible aspiration of sternal bone marrow, a reliable anatomical description of the sternum is mandatory. However, the anatomical literature provides very heterogeneous information concerning the structure and number of sternebrae. Isolated sterna (horses of different ages) underwent clinical computed tomography, and single sternebrae were scanned by microcomputed tomography. Data sets were analysed in detail, the dimensions of each sternebra were determined, and correlations to the age and weight were generated. A uniform arrangement of seven sternebrae within the equine sternum was obtained, whereas the 6th and 7th sternebrae were fused in all sterna. The cranial sternebrae (sternebrae 1-3) had a lentiform shape with flattened lateral sides, while the caudal sternebrae (6 and 7) were flattened dorso-ventrally. In contrast, sternebrae 4 and 5 were spherical. The single sternebrae were well demarcated to the chondral sternum and showed two different zones. The periphery consisted of radiodense woven tissue, while in the centre the radiodense tissue was loosely arranged and contained large cavities with radiolucent tissue. A thin lamina (substantia corticalis) of <1 mm was arranged around the peripheral zone. There was no correlation between the body weight and the dimensions of the sternebrae, but there was a positive correlation to the age of the horses. The obtained data provide a sufficient basis to establish a standard nomenclature of the equine sternum.

Impact of tetrodotoxin application and lidocaine supplementation on equine jejunal smooth muscle contractility and activity of the enteric nervous system in vitro.

By blocking the enteric nervous system (ENS) using tetrodotoxin (TTX), previous studies have documented the contractility-enhancing (CE) effects of lidocaine in equine intestinal smooth muscle (SM) at the level of SM cells and/or interstitial cells of Cajal (ICC). The present study examined the impact of ENS deactivation on CE lidocaine effects, and investigated the effects of lidocaine on ENS activity. TTX application did not affect the CE effects of lidocaine, indicating that these were not mediated by TTX-sensitive sodium channels. Application of TTX or ≥100 mg/L lidocaine reduced ENS activity. Although such concentrations of lidocaine exceed therapeutic blood concentrations, tissue concentrations may be higher with the potential to reduce ENS activity and impair intestinal motility in vivo. Improved understanding of underlying mechanisms is relevant for therapeutic use of lidocaine in horses with postoperative ileus.

Perosomus elumbis, cerebral aplasia, and spina bifida in an aborted thoroughbred foal.

Perosomus elumbis represents a rare congenital anomaly characterized by aplasia of the lumbosacral spinal cord and vertebrae. This anomaly is often associated with arthrogryposis and malformations of the urogenital and intestinal tract. This report describes the first case of perosomus elumbis in an aborted Thoroughbred foal associated with cerebral aplasia with meningocele, cranioschisis, spina bifida, a fused urogenital and intestinal tracts lined by a cutaneous mucosa without uterine glands, atresia ani, and arthrogryposis of the hind legs. Immunohistochemistry detected no abnormalities in the GFAP, S-100, vimentin, NeuN, doublecortin, and neurofilament expression pattern in the remnants of the brainstem. Pathogenic bacteria or equine herpesviruses one and four were not found using a microbiological investigation or polymerase chain reaction, respectively. The observed malformations may be induced by a severe developmental disturbance during the formation of the neural tube with secondary distortion of the surrounding mesoderm. Although the aetiology of this dysorganogenesis remains undetermined, inherited chromosomal mutations, an intake of different xenobiotics, and/or a disturbed metabolism of the mare may have caused the defect.

[Regenerative therapy for tendon and ligament disorders in horses. Terminology, production, biologic potential and in vitro effects]. / Regenerative Therapie von Sehnen- und Banderkrankungen bei Pferden. Terminologie, Herstellung, biologisches Potenzial und In-vitro-Effekte.

Conventional treatments of equine tendon injuries lead to an unsatisfactory healing process that usually results in a relatively high recurrence rate. Therefore, in recent years so-called regenerative therapeutics were studied scientifically in vitro and in laboratory animals. These include substances that ideally lead to the formation of replacement tissue, which in contrast to the low quality scar, has similar functional properties as the original intact tendon. Currently, a plethora of different substrates is either commercially available or can be produced in practice with the help of kits. The current knowledge on the production and the regenerative potential of nucleated cells like stem cells from bone marrow and fat tissue, of the blood products PRP (platelet rich plasma), ACP (autologous conditioned plasma), ACS (autologous conditioned serum) and of the scaffold substance UBM (urinary bladder matrix) are presented. Finally, the potential of some growth factors and of gene therapy is considered. Currently, it is assumed that the regeneration of tendon tissue is promoted by a complex interaction of scaffolds, growth factors and cells. At present, only very few studies are available which allow a comparison between these substances. Studies on the effect of regenerative substrates on tendons in live horses are presented elsewhere.

In vitro effects of lidocaine on the contractility of equine jejunal smooth muscle challenged by ischaemia-reperfusion injury.

REASONS FOR PERFORMING STUDY: Post operative ileus (POI) in horses is a severe complication after colic surgery. A commonly used prokinetic drug is lidocaine, which has been shown to have stimulatory effects on intestinal motility. The cellular mechanisms through which lidocaine affects smooth muscle activity are not yet known. OBJECTIVES: To examine the effects of lidocaine on smooth muscle in vitro and identify mechanisms by which it may affect the contractility of intestinal smooth muscle. HYPOTHESIS: Ischaemia and reperfusion associated with intestinal strangulation can cause smooth muscle injury. Consequently, muscle cell functionality and contractile performance is decreased. Lidocaine can improve basic cell functions and thereby muscle cell contractility especially in ischaemia-reperfusion-challenged smooth muscle. METHODS: To examine the effects of lidocaine on smooth muscle function directly, isometric force performance was measured in vitro in noninjured and in vivo ischaemia-reperfusion injured smooth muscle tissues. Dose-dependent response of lidocaine was measured in both samples. To assess membrane permeability as a marker of basic cell function, release of creatine kinase (CK) was measured by in vitro incubations. RESULTS: Lidocaine-stimulated contractility of ischaemia-reperfusion injured smooth muscle was more pronounced than that of noninjured smooth muscle. A 3-phasic dose-dependency was observed with an initial recovery of contractility especially in ischaemia-reperfusion injured smooth muscle followed by a plateau phase where contractility was maintained over a broad concentration range. CK release was decreased by lidocaine. CONCLUSION: Lidocaine may improve smooth muscle contractility and basic cell function by cellular repair mechanisms which are still unknown. Improving contractility of smooth muscle after ischaemia-reperfusion injury is essential in recovery of propulsive intestinal motility. POTENTIAL RELEVANCE: Characterisation of the cellular mechanisms of effects of lidocaine, especially on ischaemia-reperfusion injured smooth muscle, may lead to improved treatment strategies for horses with POI.

Electrophysiological characterization of electrolyte and nutrient transport across the small intestine in horses.

The aim of this study was to characterize the transport mechanisms of electrolytes and nutrients across the jejunum of nine healthy horses electrophysiologically. The stripped mucosa was mounted in Ussing chambers and tissue conductances (G(t)) and short circuit currents (I(sc)) were continuously monitored. After blocking the sodium and potassium channels with amiloride, tetraethylammonium chloride (TEA) and barium, chloride secretion was stimulated by carbachol and forskolin. Subsequently, chloride channels were inhibited by 4,4'-diisothiocyanato-stilbene-2,2'-disulfonic acid, 5-nitro-2-(3-phenylpropylamino)benzoic acid, CFTR(inh)-172, N-(2-naphtalenyl)-(3.5-dibromo-2.4-dihydroxyphenyl)methylene glycine hydrazide (GlyH-101) and glibenclamide and their dose-response effect was investigated. The response to glucose, l-alanine and glycyl-l-glutamine was determined at two different mucosal pH values (pH 7.4 and 5.4 respectively). Mean basal I(sc) was -0.47 +/- 0.31 microEq/cm(2)h and mean G(t) was 22.17 +/- 1.78 mS/cm(2). Amiloride and TEA did not alter the baseline I(sc). Barium, carbachol and forskolin significantly increased I(sc). Irrespective of the dose, none of the chloride inhibitors changed I(sc). All nutrients induced a significant increase in I(sc) with the increase being significantly higher at pH 7.4 than at pH 5.4. In conclusion, there is evidence that chloride secretion in horses may be different from respective transport mechanisms in other species. The glucose absorption is suggestive of a sodium-dependent glucose cotransporter 1. However, a decrease in luminal pH did not stimulate current response to peptides as shown for other mammals.

[Verrucous pastern dermatitis syndrome in heavy draught horses. Part II: Clinical findings]. / Zur Entstehung von Warzenmauke bei Kaltblutpferden. Teil II: Klinische Untersuchungen.

In the present field study the skin of the feet of 37 heavy draught horses of different breeds showing verrucous pastern dermatitis was examined clinically. Included were the degree of severity of the disease and the prevalence of anatomically normal structures associated with the skin: fetlock tufts of hair ("feathering"), ergots, chestnuts, bulges in the pastern region, cannon circumference. Each horse was examined for Chorioptes sp. skin mites. Information was also collected on the development of the skin alterations and housing conditions and feeding. These individual data were correlated with the clinical degree of severity of verrucous pastern dermatitis, which was evaluated using a numerical code (scoring system). In addition, punch biopsies were taken from the diseased skin of the feet and from healthy skin of the neck for comparative patho-histological examination (see Part III). Verrucous pastern dermatitis is a chronic disease which can be divided into four groups: scaling (group I), hyperkeratotic and hyperplastic plaque-like lesions (group II), tuberous skin masses (group III), and verrucous skin lesions with rugged surfaces (group IV). No correlation was found between the clinical degree of severity of the skin lesions and sex, breed, amount of work, use of stallions for breeding, grooming condition of the hair, white markings in the foot region, or Chorioptes sp. infestation. In regard to feeding it was found that the amount of maize and oats fed had some influence on the clinical degree of severity. Statistical analysis revealed a significant correlation between the clinical degree of severity and the age, the grooming condition of the hooves, and the mean cannon circumference. The prevalence of fetlock tufts of hair, chestnuts, ergots, and anatomically normal bulges in the pastern region also increased significantly with the clinical degree of severity. Furthermore the study revealed that the clinical degree of severity depended on the hygienic conditions of the stables and of the ground where the horses were kept outdoors.

[The development of verrucous pastern dermatitis syndrome in heavy draught horses. Part I: Review of the literature]. / Zur Entstehung von Warzenmauke bei Kaltblutpferden. Teil I: Literaturübersicht.

Verrucous pastern dermatitis is a chronic hyperplastic dermatopathy of the feet of horses which is characterized by a greasy, odorous coating of the skin and, in advanced stages, by clearly demarcated calluses and wart-like proliferations. The disease occurs almost exclusively in cold-blooded and other heavy horses, with certain breeds affected most frequently. It is considered a distinct disease entity within the framework of pastern dermatitis syndrome. There is no consensus in the literature about relationship of the disease to the sex and age of the horse. Horses with a high cannon circumference and pronounced fetlock tufts of hair seem to be affected most severely. In chiefly anecdotal reports the cause of the disease or its aggravation has been ascribed to housing conditions and environmental influences, chemicals, mechanical insults, feeding, or infections with Chorioptes skin mites. It thus seems likely that the disease is affected by a variety of factors. Although verrucous pastern dermatitis has been known for hundreds of years, its etiology and pathogenesis remain unclear.

Polioencephalomalacia in captive harbour seals (Phoca vitulina).

In a colony of 11 harbour seals (Phoca vitulina Linné 1758) two episodes of central nervous disorders occurred within 2 years causing fatalities in seven adult animals. Clinical signs comprised dyspnoea, anorexia, apathy, incoordination and lateral recumbency. Vitamin B complex therapy was successful once. Pathomorphological examination of seven carcasses revealed acute and subacute malacia of the cerebellar grey matter. Additional acute malacic lesions located in the cerebral cortices and basal ganglia were observed. Mesencephalic nuclei were less severely affected and displayed acute changes. Despite intense search for environmental toxins and infectious agents, the cause of the fatalities remained undetermined. However, the type and pattern of the lesions are most suggestive of a thiamine deficiency.

[Nerve sheath tumors in cattle: literature review and case report]. / Nervenscheidentumoren beim Rind: Schrifttumsübersicht und Fallbericht.

Incidence, clinical signs, pathomorphological findings, differential diagnosis and prognosis of bovine nerve sheath tumours are reviewed. Own clinical and pathomorphological findings are described in an eight years old cow with a benign SCHWANNoma of the eight spinal nerve.