Interleukin-31 in serum and cerebrospinal fluid of dogs with syringomyelia.

BACKGROUND: Syringomyelia is a spinal cord cavity containing cerebrospinal fluid (CSF)-like fluid. If syringomyelia asymmetrically involves the dorsal horn grey matter of the spinal cord, affected dogs show increased signs of dysesthesia and neuropathic pain, like increased itching behaviour. In the dorsal horn, amongst others, receptors for Interleukin-31 (IL-31) can be found. IL-31 is one of the main cytokines involved in the pathogenesis of pruritus in atopic dermatitis in different species. This study investigates suspected elevated levels of IL-31 in serum and CSF of dogs showing signs of pain or increased itching behaviour related to syringomyelia. The IL-31 were measured in archived samples (52 serum and 35 CSF samples) of dogs with syringomyelia (n = 48), atopic dermatitis (n = 3) and of healthy control dogs (n = 11) using a competitive canine IL-31 ELISA. RESULTS: Mean serum IL-31 level in dogs with syringomyelia was 150.1 pg/ml (n = 39), in dogs with atopic dermatitis 228.3 pg/ml (n = 3) and in healthy dogs 80.7 pg/ml (n = 10). Mean CSF IL-31 value was 146.3 pg/ml (n = 27) in dogs with syringomyelia and 186.2 pg/ml (n = 8) in healthy dogs. Individual patients with syringomyelia (especially dogs with otitis media or otitis media and interna or intervertebral disc herniation) showed high IL-31 levels in serum and CSF samples, but the difference was not statistically significant. IL-31 serum and CSF levels did not differ significantly in dogs with syringomyelia with or without itching behaviour and with or without signs of pain. CONCLUSION: Based on this study, increased IL-31 levels seem not to be correlated with itching behaviour or signs of pain in dogs with syringomyelia, but might be caused by other underlying diseases.

Elevated Interleukin-31 Levels in Serum, but Not CSF of Dogs with Steroid-Responsive Meningitis-Arteritis Suggest an Involvement in Its Pathogenesis.

Steroid-responsive meningitis-arteritis (SRMA) is a predominantly Th-2 immune-mediated disease, but the exact pathomechanism remains unclear. Interleukin-31 (IL-31) is predominantly produced by T cells with a Th-2 phenotype during proinflammatory conditions. We hypothesize that IL-31 might be involved in the pathogenesis of SRMA. IL-31 was measured in archived samples (49 serum and 52 CSF samples) of dogs with SRMA, meningoencephalitis of unknown origin (MUO), infectious meningoencephalitis, and atopic dermatitis, and of healthy control dogs using a competitive canine IL-31 ELISA. The mean serum IL-31 level in dogs with SRMA (n = 18) was mildly higher compared to dogs with atopic dermatitis (n = 3, p = 0.8135) and MUO (n = 15, p = 0.7618) and markedly higher than in healthy controls (n = 10, p = 0.1327) and dogs with infectious meningoencephalitis (n = 3, no statistics). Dogs with SRMA in the acute stage of the disease and without any pre-treatment had the highest IL-31 levels. The mean CSF IL-31 value for dogs with SRMA (n = 23) was quite similar to that for healthy controls (n = 8, p = 0.4454) and did not differ markedly from dogs with MUO (n = 19, p = 0.8724) and infectious meningoencephalitis. Based on this study, an involvement of IL-31 in the pathogenesis of the systemic Th-2 immune-mediated immune response in SRMA can be assumed as a further component leading to an aberrant immune reaction.

Atypically located arachnoid cyst in a five-year-old cat.

Arachnoid cysts are cystic lesions that occur in spinal or intracranial locations in the leptomeningeal space. Four intracranial cases have been described in cats, three of which were diagnosed by imaging techniques alone. We now report the clinical, gross and histopathological findings in a 5-year-old, male-neutered European Shorthair cat that presented with chronic, asymmetrical encephalopathy. Using magnetic resonance imaging, a focal, fluid-filled cavity that did not show contrast enhancement was identified in the left temporal and piriform lobes. Necropsy confirmed the presence of a cystic, meningeal cavity filled with clear, serous fluid. Histologically, the cyst had an irregular, hypereosinophilic surface and single psammoma bodies with moderate perivascular oedema in the adjacent neuroparenchyma. Immunohistochemical evidence of meningeal tissue surrounding the cyst confirmed the diagnosis of an arachnoid cyst, which should be considered as a differential diagnosis of intracranial, fluid-filled cavities.

Investigation on solubilization protocols in the refolding of the thioredoxin TsnC from Xylella fastidiosa by high hydrostatic pressure approach.

The lack of efficient refolding methodologies must be overcome to take full advantage of the fact that bacteria express high levels of aggregated recombinant proteins. High hydrostatic pressure (HHP) impairs intermolecular hydrophobic and electrostatic interactions, dissociating aggregates, which makes HHP a useful tool to solubilize proteins for subsequent refolding. A process of refolding was set up by using as a model TsnC, a thioredoxin that catalyzes the disulfide reduction to a dithiol, a useful indication of biological activity. The inclusion bodies (IB) were dissociated at 2.4 kbar. The effect of incubation of IB suspensions at 1-800 bar, the guanidine hydrochloride concentration, the oxidized/reduced glutathione (GSH/GSSG) ratios, and the additives in the refolding buffer were analyzed. To assess the yields of fully biologically active protein obtained for each tested condition, it was crucial to analyze both the TsnC solubilization yield and its enzymatic activity. Application of 2.4 kbar to the IB suspension in the presence of 9 mM GSH, 1mM GSSG, 0.75 M guanidine hydrochloride, and 0.5M arginine with subsequent incubation at 1 bar furnished high refolding yield (81%). The experience gained in this study shall help to establish efficient HHP-based protein refolding processes for other proteins.

Refolding by high pressure of a toxin containing seven disulfide bonds: bothropstoxin-1 from Bothrops jararacussu.

Aggregation is a serious obstacle for recovery of biologically active heterologous proteins from inclusion bodies (IBs) produced by recombinant bacteria. E. coli transformed with a vector containing the cDNA for Bothropstoxin-1 (BthTx-1) expressed the recombinant product as IBs. In order to obtain the native toxin, insoluble and aggregated protein was refolded using high hydrostatic pressure (HHP). IBs were dissolved and refolded (2 kbar, 16 h), and the effects of protein concentration, as well as changes in ratio and concentration of oxido-shuffling reagents, guanidine hydrochloride (GdnHCl), and pH in the refolding buffer, were assayed. A 32% yield (7.6 mg per liter of bacterial culture) in refolding of the native BthTx-1 was obtained using optimal conditions of the refolding buffer (Tris-HCl buffer, pH 7.5, containing 3 mM of a 2:3 ratio of GSH/GSSG, and 1 M GdnHCl). Scanning electron microscopy (SEM) showed that that disaggregation of part of IBs particles occurred upon compression and that the morphology of the remaining IBs, spherical particles, was not substantially altered. Dose-dependent cytotoxic activity of high-pressure refolded BthTx-1 was shown in C2C12 muscle cells.

Concurrent Helicobacter bilis infection in C57BL/6 mice attenuates proinflammatory H. pylori-induced gastric pathology.

Because coinfections can alter helicobacter gastritis, we investigated whether enterohepatic Helicobacter bilis modulates Helicobacter pylori gastritis in C57BL/6 mice. Thirty mice per group were sham dosed, H. bilis or H. pylori infected, or H. bilis infected followed in 2 weeks by H. pylori and then evaluated at 6 and 11 months postinfection (mpi) for gastritis and premalignant lesions. Compared to H. pylori-infected mice, H. bilis/H. pylori-infected mice at 6 and 11 mpi had less severe gastritis, atrophy, mucous metaplasia and hyperplasia (P < 0.01) and, additionally, at 11 mpi, less severe intestinal metaplasia and dysplasia (P < 0.05). H. bilis/H. pylori-infected mice at 11 mpi exhibited less Ki67 labeling of proliferating epithelial cells, reduced numbers of FoxP3(+) T-regulatory (T(REG)) cells, and lower FoxP3(+) mRNA levels than did H. pylori-infected mice (P < 0.05). Proinflammatory interleukin-1beta (IL-1beta), gamma interferon, and tumor necrosis factor alpha mRNA levels were attenuated in H. bilis/H. pylori-infected mice at 6 and 11 mpi (P < 0.01), although anti-inflammatory IL-10, IL-13, and transforming growth factor beta1 mRNA levels were not consistently impacted by H. bilis coinfection. Decreased pathology in H. bilis/H. pylori-infected mice correlated with higher gastric H. pylori colonization at 6 mpi (P < 0.001) and lower Th1-associated immunoglobulin G2c responses to H. pylori at 6 and 10 mpi (P < 0.05). We hypothesized that reduced pathology in H. bilis/H. pylori-infected mice was due to H. bilis-primed T(REG) cells in the lower bowel that migrated to the gastric compartment and inhibited Th1 responses to subsequent H. pylori infection. Thus, H. pylori-induced gastric lesions may vary in mouse models of unknown enteric helicobacter infection status and, importantly, variable sequelae to human H. pylori infection, particularly in developing countries, may occur where coinfection with lower bowel helicobacters and H. pylori may be common.

Obesity and non-insulin-dependent diabetes mellitus in Swiss-Webster mice associated with late-onset hepatocellular carcinoma.

Genetic mutations resulting in obesity and type 2 diabetes mellitus (T2D) are described for both inbred and outbred mice. However, no known mouse model completely recapitulates human T2D and its comorbidities. We identified a cohort of obese, male, outbred Swiss-Webster (SW) mice as polyuric, polydipsic, glucosuric, and hyperglycemic. Prevalence of glucosuria in the SW colony reached 60% (n=70) in males 8 weeks to 6 months of age. Despite severe obesity in some females, no females were diabetic. Pathologic findings in affected males included cachexia, dilated gastrointestinal tracts with poor muscular tone, pancreatic islet degeneration and atrophy with compensatory metaplasia and/or neogenesis, bacterial pyelonephritis, membranous glomerulopathy, and late-onset hepatic tumors with macrosteatosis, microsteatosis, and hydropic change in aged males. Serum insulin correlated with blood glucose in a nonlinear pattern, suggestive of islet exhaustion. Circulating leptin levels showed a weak inverse correlation with glucose. Diabetic males were bred with obese colony females to produce 20 male and 20 female offspring. Prevalence of diabetes in male offspring was 80% (16/20) with a median age of onset of 18 weeks. By contrast, no diabetic females were identified, despite being significantly more obese than males. Male predominance is likewise a feature of T2D in humans. To our knowledge, this is the first documentation of hepatocellular carcinoma and islet metaplasia and/or neogenesis in a spontaneous outbred mouse model of T2D. The SW availability and histopathologic features represent a promising new model for the study of T2D.

Infestation of wild-caught American bullfrogs (Rana catesbeiana) by multiple species of metazoan parasites.

The American bullfrog (Rana catesbeiana) is an aquatic, carnivorous member of the family Ranidae that is used extensively in physiology education programs and in various physiology, toxicology, sensorineural, and genetics research. Eleven bullfrogs purchased from a vendor distributing wild-caught frogs for use in a physiology research protocol were emaciated but otherwise showed no apparent clinical signs of illness. Necropsies performed on selected emaciated frogs indicated heavy infestation with multiple species of endoparasites. Identified helminths included Gorgodera amplicava, Haematolechus breviplexus, Clinostomum spp, Contracaecum spp, Cosmocercoides dukae, and Eustrongyloides spp. Grossly, parasitized bullfrogs showed encysted trematode larvae within skeletal muscle, nematode impaction of the intestinal tract, and lack of coelemic fat stores. Histopathologic lesions were restricted primarily to the gastrointestinal tract and consisted of parasitic granulomas associated with Contracaecum spp. The parasitic lesions may have been associated with the poor body condition of the bullfrogs. Food crickets maintained in-house were negative for parasite larvae or ova. Heavy parasitism of wild-caught bullfrogs may confound research protocols and markedly impair animal health. We encourage researchers to purchase laboratory-bred and -reared bullfrogs and to routinely monitor the parasite status of colony frogs.

Crouched gait in myelomeningocele: a comparison between the degree of knee flexion contracture in the clinical examination and during gait.

The purpose of this study was to quantitatively evaluate, in patients with low lumbar and sacral level myelomeningocele who have knee flexion contractures, whether there are significant differences between the degree of knee flexion contracture measured clinically and the degree of actual knee flexion during gait, measured by computerized gait analysis. Patients were divided into two groups, those who walked with ankle-foot orthoses (AFOs) alone and those who walked with AFOs and crutches. In both groups, the patient's knee flexion contractures were measured clinically, and the degree of knee flexion was measured dynamically at two representative points in the gait cycle. In both groups and at both points of the gait cycle, the degree of knee flexion during gait was significantly greater than the degree of clinical knee flexion contracture. This should be taken into account when evaluating the crouch gait of children with myelomeningocele and planning the proper treatment.

The surgical management of pediatric fractures of the upper extremity.

The vast majority of pediatric fractures of the upper extremity can and should be treated with closed reduction, immobilization, and close follow-up. However, there is an ongoing debate in the orthopaedic community regarding the exact role of surgical management in the treatment of pediatric fractures. In the past 2 decades, surgical management of certain fractures (e.g., percutaneous pinning of displaced supracondylar fractures) has provided better results than closed management. Surgical management is clearly indicated for certain injuries, such as those requiring anatomic realignment of the physis or articular surface. Increasingly, however, surgical management is being used to maintain optimal alignment or to allow early motion. In many such cases, both nonsurgical and surgical methods have yielded good results and have vocal advocates. Certain technical advances, such as flexible intramedullary fixation and bioreabsorbable implants, have further increased enthusiasm for surgical management of pediatric fractures.