Histological and molecular characterisation of feline humeral condylar osteoarthritis

<p>Background: Osteoarthritis (OA) is a clinically important and common disease of older cats. The pathological changes and molecular mechanisms which underpin the disease have yet to be described. In this study we evaluated selected histological and transcriptomic measures in the articular cartilage and subchondral bone (SCB) of the humeral condyle of cats with or without OA.</p> <p>Results: The histomorphometric changes in humeral condyle were concentrated in the medial aspect of the condyle. Cats with OA had a reduction in articular chondrocyte density, an increase in the histopathological score of the articular cartilage and a decrease in the SCB porosity of the medial part of the humeral condyle. An increase in LUM gene expression was observed in OA cartilage from the medial part of the humeral condyle.</p> <p>Conclusions: Histopathological changes identified in OA of the feline humeral condyle appear to primarily affect the medial aspect of the joint. Histological changes suggest that SCB is involved in the OA process in cats. Differentiating which changes represent OA rather than the aging process, or the effects of obesity and or bodyweight requires further investigation.</p&gt

Lung injury caused by aspiration of organophosphorus insecticide and gastric contents in pigs

INTRODUCTION: Patients who require mechanical ventilation after self-poisoning with ingested organophosphorus (OP) insecticides often die. Aspiration of stomach contents may contribute to lung injury and lethality. This study was designed to assess the severity of direct and indirect pulmonary injury created by pulmonary instillation of mixtures of OP insecticide, solvent (Solv) and porcine gastric juice (GJ) compared to controls. METHODS: Terminally anaesthetised minipigs (groups n = 5) were exposed to sham bronchoscopy or given mixtures (0.5 mL/kg) of: saline, GJ, OP insecticide and GJ (OP + GJ), or Solv and GJ (Solv + GJ), placed into the right lung, and monitored for 48 h. Lung injury was assessed through analysis of bronchoalveolar lavage fluid (BALF), computed tomography and histopathology. RESULTS: OP + GJ created a direct lung injury consisting of neutrophil infiltration, oedema and haemorrhage, as well as indirect injury to the other lung. OP + GJ directly-injured lung parenchyma had increased concentrations of BALF protein, albumin, IL-6, IL-8 and C-reactive protein (CRP) at 24 h (p < 0.05), and BALF protein, albumin and CRP at 48 h (p < 0.05), when compared with controls. Aspiration of GJ produced similar direct effects to OP + GJ but less indirect lung injury. Lung injury was less severe after Solv + GJ, for combined lung histopathology scores (vs. OP + GJ, p < 0.05) and for the proportion of directly-injured lung that was poorly/non-aerated at 48 h. CONCLUSION: Pulmonary instillation of OP + GJ created more lung damage than controls or Solv + GJ. In patients with severe OP insecticide poisoning and reduced consciousness, early airway protection is likely to reduce pulmonary damage

Development of a histopathology scoring system for the pulmonary complications of organophosphorus insecticide poisoning in a pig model

Organophosphorus (OP) insecticide self-poisoning causes over 100,000 global deaths annually. Around a third of patients are intubated and up to half of these can die. Post-mortem analysis of OP poisoned patients' lungs reveals consolidation, edema and hemorrhage, suggesting that direct or indirect lung damage may contribute to mortality. The lung injury caused by these formulated agricultural preparations is poorly characterised in humans, and a valid histopathology scoring system is needed in a relevant animal model to further investigate the disease and potential treatments. We conducted two pilot studies in anesthetized minipigs, which are commonly used for toxicological studies. In the first, pigs were given 2.5 mL/kg of either OP (n = 4) or saline (n = 2) by gavage and compared with positive controls (iv oleic acid n = 2). The second study simulated ingestion followed by gastric content aspiration: mixtures of OP (n = 3) or saline (n = 2) (0.63-0.71mL/kg) were placed in the stomach, and then small volumes of the gastric content were placed in the lung. At post-mortem examination, lungs were removed and inflation-fixed with 10% neutral buffered formalin. Samples (n = 62) were taken from cranial and caudal regions of both lungs. Two experienced lung histopathologists separately scored these samples using 8 proposed features of damage and their scores related (Kendall rank order). Two elements had small and inconsistent scores. When these were removed, the correlation increased from 0.74 to 0.78. Eight months later, a subset of samples (n = 35) was re-scored using the modified system by one of the previous histopathologists, with a correlation of 0.88. We have developed a reproducible pulmonary histopathology scoring system for OP poisoning in pigs which will assist future toxicological research and improve understanding and treatment of human OP poisoning

Down-regulation of mechanisms involved in cell transport and maintenance of mucosal integrity in pigs infected with Lawsonia intracellularis

Lawsonia intracellularis is an obligate intracellular bacterium, responsible for the disease complex known as proliferative enteropathy (PE). L. intracellularis is associated with intestinal crypt epithelial cell proliferation but the mechanisms responsible are yet to be defined. Microarray analysis was used to investigate the host-pathogen interaction in experimentally infected pigs to identify pathways that may be involved. Ileal samples originating from twenty-eight weaner pigs experimentally challenged with a pure culture of L. intracellularis (strain LR189/5/83) were subjected to microarray analysis. Microarray transcriptional signatures were validated using immunohistochemistry and quantitative real time PCR of selected genes at various time points post challenge. At peak of infection (14 days post challenge) 86% of altered transcripts were down regulated, particularly those involved in maintenance of mucosal integrity and regulation of cell transport. Among the up-regulated transcripts, CD163 and CDK1 were novel findings and considered to be important, due to their respective roles in innate immunity and cellular proliferation. Overall, targeted cellular mechanisms included those that are important in epithelial restitution, migration and protection; maintenance of stable inter-epithelial cell relationships; cell transport of nutrients and electrolytes; innate immunity; and cell cycle

Inflammatory infiltration into placentas of Neospora caninum challenged cattle correlates with clinical outcome of pregnancy

International audienceInfection with Neospora caninum stimulates host cell-mediated immune responses, which may be responsible for placental damage leading to bovine abortion. The aim of this study was to compare immune responses in the bovine placenta, following experimental infection in different stages of pregnancy. Placentomes were examined by immunohistochemistry and inflammation in early gestation was generally moderate to severe, particularly in the placentas carrying non-viable foetuses, whereas it was milder in later stages, mainly characterised by the presence of CD3+, CD4+ and γδ T-cells. This distinctive cellular immune response may explain the milder clinical outcome observed when animals are infected in later gestation

Lawsonia intracellularis infection of intestinal crypt cells is associated with specific depletion of secreted MUC2 in goblet cells

AbstractThe expression patterns of secreted (MUC2 and MUC5AC) and membrane-tethered (MUC1, MUC4, MUC12 and MUC13) mucins were monitored in healthy pigs and pigs challenged orally with Lawsonia intracellularis. These results showed that the regulation of mucin gene expression is distinctive along the GI tract of the healthy pig, and may reflect an association between the function of the mucin subtypes and different physiological demands at various sites. We identified a specific depletion of secreted MUC2 from goblet cells in infected pigs that correlated with the increased level of intracellular bacteria in crypt cells. We concluded that L. intracellularis may influence MUC2 production, thereby altering the mucus barrier and enabling cellular invasion

The safety profile of a cationic lipid-mediated cystic fibrosis gene transfer agent following repeated monthly aerosol administration to sheep

Clinically effective gene therapy for Cystic Fibrosis has been a goal for over 20 years. A plasmid vector (pGM169) that generates persistent expression and reduced host inflammatory responses in mice has raised prospects for translation to the clinic. The UK CF Gene Therapy Consortium is currently evaluating long-term repeated delivery of pGM169 complexed with the cationic lipid GL67A in a large Multidose Trial. This regulatory-compliant evaluation of aerosol administration of nine doses of pGM169/GL67A at monthly intervals, to the sheep lung, was performed in preparation for the Multidose Trial. All sheep tolerated treatment well with no adverse effects on haematology, serum chemistry, lung function or histopathology. Acute responses were observed in relation to bronchoalveolar cellularity comprising increased neutrophils and macrophage numbers 1 day post-delivery but these increases were transient and returned to baseline. Importantly there was no cumulative inflammatory effect or lung remodelling with successive doses. Molecular analysis confirmed delivery of pGM169 DNA to the airways and pGM169-specific mRNA was detected in bronchial brushing samples at day 1 following doses 1, 5 and 9. In conclusion, nine doses of pGM169/GL67A were well tolerated with no significant evidence of toxicity that would preclude adoption of a similar strategy in CF patients

Lawsonia intracellularis exploits β-catenin/Wnt and Notch signalling pathways during infection of intestinal crypt to alter cell homeostasis and promote cell proliferation

Lawsonia intracellularis is an obligate intracellular bacterial pathogen that causes proliferative enteropathy (PE) in pigs. L. intracellularis infection causes extensive intestinal crypt cell proliferation and inhibits secretory and absorptive cell differentiation. However, the affected host upstream cellular pathways leading to PE are still unknown. β-catenin/Wnt signalling is essential in maintaining intestinal stem cell (ISC) proliferation and self-renewal capacity, while Notch signalling governs differentiation of secretory and absorptive lineage specification. Therefore, in this report we used immunofluorescence (IF) and quantitative reverse transcriptase PCR (RTqPCR) to examine β-catenin/Wnt and Notch-1 signalling levels in uninfected and L. intracellularis infected pig ileums at 3, 7, 14, 21 and 28 days post challenge (dpc). We found that while the significant increase in Ki67+ nuclei in crypts at the peak of L. intracellularis infection suggested enhanced cell proliferation, the expression of c-MYC and ASCL2, promoters of cell growth and ISC proliferation respectively, was down-regulated. Peak infection also coincided with enhanced cytosolic and membrane-associated β-catenin staining and induction of AXIN2 and SOX9 transcripts, both encoding negative regulators of β-catenin/Wnt signalling and suggesting a potential alteration to β-catenin/Wnt signalling levels, with differential regulation of the expression of its target genes. We found that induction of HES1 and OLFM4 and the down-regulation of ATOH1 transcript levels was consistent with the increased Notch-1 signalling in crypts at the peak of infection. Interestingly, the significant down-regulation of ATOH1 transcript levels coincided with the depletion of MUC2 expression at 14 dpc, consistent with the role of ATOH1 in promoting goblet cell maturation. The lack of significant change to LGR5 transcript levels at the peak of infection suggested that the crypt hyperplasia was not due to the expansion of ISC population. Overall, simultaneous induction of Notch-1 signalling and the attenuation of β-catenin/Wnt pathway appear to be associated with the inhibition of goblet cell maturation and enhanced crypt cell proliferation at the peak of L. intracellularis infection. Moreover, the apparent differential regulation of apoptosis between crypt and lumen cells together with the strong induction of Notch-1 signalling and the enhanced SOX9 expression along crypts 14 dpc suggest an expansion of actively dividing transit amplifying and/or absorptive progenitor cells and provide a potential basis for understanding the development and maintenance of PE

Presence of Systemic Inflammatory Response Syndrome Predicts a Poor Clinical Outcome in Dogs with a Primary Hepatitis

<div><p>Primary hepatopathies are a common cause of morbidity and mortality in dogs. The underlying aetiology of most cases of canine hepatitis is unknown. Consequently, treatments are typically palliative and it is difficult to provide accurate prognostic information to owners. In human hepatology there is accumulating data which indicates that the presence of systemic inflammatory response syndrome (SIRS) is a common and debilitating event in patients with liver diseases. For example, the presence of SIRS has been linked to the development of complications such as hepatic encephalopathy (HE) and is associated with a poor clinical outcome in humans with liver diseases. In contrast, the relationship between SIRS and clinical outcome in dogs with a primary hepatitis is unknown. Seventy dogs with histologically confirmed primary hepatitis were enrolled into the study. Additional clinical and clinicopathological information including respiratory rate, heart rate, temperature, white blood cell count, sodium, potassium, sex, presence of ascites, HE score, alanine aminotransferase (ALT), alkaline phosphatase (ALP), bilirubin and red blood cell concentration were available in all cases. The median survival of dogs with a SIRS score of 0 or 1 (SIRS low) was 231 days compared to a median survival of 7 days for dogs with a SIRS score of 2, 3 or 4 (SIRS high) (p<0.001). A Cox proportional hazard model, which included all other co-variables, revealed that a SIRS high score was an independent predictor of a poor clinical outcome. The effect of modulating inflammation on treatment outcomes in dogs with a primary hepatitis is deserving of further study.</p></div