Intraoperative tissue sampling for histology in chronic osteomyelitis shows high neutrophil infiltration centrally and low remains in debrided presumed infection-free regions.

INTRODUCTION: Histology of debrided bone tissue is a confirmatory diagnostic criterion for fracture related infection (FRI) and prosthetic joint infection (PJI). The aim of the present study was to describe the histopathology of the first and last debrided bone tissue in chronic osteomyelitis (CO) according to the international diagnostic guidelines for FRI and PJI. METHODS: 15 patients with CO were allocated to surgical treatment using a one-stage protocol including extensive debridement. Suspected infected bone tissue eradicated early in the debridement procedure was collected as a clearly infected sample (S1). Likewise, the last eradicated bone tissue was collected as a suspected non-infected sample (S2). The samples were processed for histology. HE-stained sections were patho-morphologically examinated. Immunohistochemistry with MAC-387 antibodies towards calprotectin was used for estimation of neutrophil granulocyte (NP) score (0, 1, 2 or 3). RESULTS: S1 samples showed a mean NP score of 2.6 (3 is confirmatory for infection). Following debridement, the NP score was significantly (p = 0.005) reduced to a mean NP score of 1.6. The S1 samples showed a mix of fibrovascular tissue, dense fibrosis, viable bone, bone necrosis and bone debris. S2 samples contained mostly viable bone tissue, however, often small fragments of necrotic bone or bone debris were present. CONCLUSION: The inflammatory response of CO still exists after debridement, although the response fades from the center. Therefore, sampling of debrided bone tissue for histology must be performed initially during surgery, otherwise there is a risk for underestimation of NP infiltration. The present results might also be highly relevant for FRI and PJI.

An observational field study of porcine post-weaning diarrhea: clinical and microbiological findings, and fecal pH-measurements as a potential diagnostic tool.

BACKGROUND: Recently, in-feed medicinal zinc has been phased out in pig production in the European Union. This makes updated knowledge about porcine post-weaning diarrhea (PWD) crucial. The objectives of the present study were to investigate (i) the clinical presentation of PWD in pigs housed in Danish herds that did not use medicinal zinc, specifically the prevalence of diarrhea and whether PWD was associated to clinical signs of dehydration or altered body temperature; (ii) which microorganism are associated to PWD; and iii) whether measurements of the fecal pH have a potential to be used diagnostically to differentiate between infectious etiologies in cases of PWD. RESULTS: The prevalence of diarrhea varied considerably between the outbreaks in the nine studied herds (median = 0.58, range = 0.10; 0.94). In a cross-sectional design (n = 923), diarrhea was associated with reduced rectal temperature and alkaline feces. Diarrhea was also associated with observably reduced skin elasticity, possibly indicating dehydration. In both diarrheic case pigs (n = 87) and control pigs (n = 86), the presence of Brachyspira pilosicoli, Clostridium perfringens, Cryptosporidium spp., Cystoisopora suis, enterotoxigenic Escherichia coli, Lawsonia intracellularis, porcine circovirus types 2 and 3, rotavirus A, B, C, and H, Samonella enterica spp. enterica, and Trichuris suis was described. PWD was associated with high levels of enterotoxigenic E. coli shedding (odds ratio versus no E. coli detection = 4.79 [CI 1.14; 12.62]). Diarrhea was associated with high levels of rotavirus A shedding (odds ratio versus no/low rotavirus A = 3.80 [CI 1.33; 7.97]). The association between microbiological findings in diarrheic pigs and fecal pH was negligible. CONCLUSIONS: Enterotoxigenic E. coli was confirmed to be a cause of PWD; however, cases of PWD where enterotoxigenic E. coli was not detected in high levels occurred commonly, and this adds to the increasing evidence suggesting that PWD is not necessarily a result of enteric colibacillosis. Rotaviral enteritis might be a differential diagnosis of PWD. pH-measurements cannot be used to differentiate between differential diagnoses for PWD.

Development of an aerogenous Escherichia coli infection model in adult broiler breeders.

Escherichia coli constitutes an immense challenge to the poultry industry due to its devastating effect on productivity, mortality, and carcass condemnations. To aid future studies on disease mechanisms and interventions, an aerogenous infection model was established in adult broiler breeders. Hens (n = 120) were randomly allocated into six groups receiving either aerosolised E. coli or vehicle, or intratracheal E. coli or vehicle. Replication of aerosol inoculation was performed on distinct days. Alternating euthanasia time points were predetermined in order to evaluate the progression of the disease. All animals were thoroughly necropsied, and bacteriological samples were collected as well as tissues for histopathology. Birds inoculated with E. coli exhibited clinical signs and developed characteristic gross and histopathological lesions of colibacillosis, including splenic fibrinoid necrosis, folliculitis, polyserositis and impaction of parabronchi with fibrinoheterophilic exudate and necrotic debris, as well as positive in situ localisation of intralesional E. coli by immunohistochemistry. This study presents a successful development of a discriminative colibacillosis model through aerosol inoculation of adult broiler breeders. Gross and histopathological lesions characteristic of colibacillosis were established in two independent experiments.

Evaluation of public and animal health risks in case of a delayed post-mortem inspection in ungulates.

The potential effects of a 24 or 72-h delay in post-mortem inspection (PMI) of ungulates on public health and monitoring of animal health and welfare was evaluated. The assessment used a survey of meat inspectors, expert opinion, literature search and a stochastic model for Salmonella detection sensitivity. Disease detection sensitivity at a delayed PMI is expected to reduce detection sensitivity to a variable extent, depending on the hazard and on the signs/lesions and organs involved. No reduction is expected for Trichinella detection in meat from susceptible animal species and any decrease in detection of transmissible spongiform encephalopathies (TSEs) will not exceed the current tolerance for fallen stock. A 24-h delay in PMI could result in a small reduction in sensitivity of detection for tuberculosis, echinococcosis and cysticercosis. A greater reduction is expected for the detection of pyaemia and Rift valley fever. For the detection of Salmonella, the median model estimates are a reduction of sensitivity of 66.5% (90% probability interval (PI) 0.08-99.75%) after 24-h delay and 94% (90% PI 0.83-100%) after 72-h delay of PMI. Laboratory testing for tuberculosis following a sampling delay of 24-72 h could result in no, or a moderate, decrease in detection depending on the method of confirmation used (PCR, culture, histopathology). For chemical contaminants, a delay in meat inspection of 24 or 72 h is expected to have no impact on the effectiveness of detection of persistent organic pollutants and metals. However, for certain pharmacologically active substances, there will be a reduced effectiveness to detect some of these substances due to potential degradation in the available matrices (tissues and organs) and the non-availability of specific preferred matrices of choice.

Inflammatory proteins in infected bone tissue - An explorative porcine study.

OBJECTIVE: To explore the in situ inflammatory proteins in the local extracellular fluid of infected bone tissue. MATERIAL AND METHODS: Seven pigs went through a two-step surgery performing a traumatically implant-associated Staphylococcus aureus osteomyelitis in the proximal tibia. Five days later, microdialysis catheters (membrane cut off: 20 kDa) were placed in the implant cavity, infected and healthy cancellous bone, and infected and healthy subcutaneous tissue. Plasma samples were collected simultaneously. We employed an antibody-based proximity extension assay (Olink Inflammatory panel) for the measurement of inflammatory molecules within plasma and extracellular fluid of the investigated tissue compartments. RESULTS: A higher normalized protein expression in the infected bone tissue in comparison to healthy bone tissue was identified for proteins associated with angiogenesis and bone remodeling: OPG, TGFα, MCP-1, VEGFA, and uPA. Moreover, a parallel detectability of the systemic range of cytokines and chemokines as from the investigated local tissue compartments was demonstrated, indicating the same occurrence of proteins in the local environment as within plasma. CONCLUSION: An angiogenic and osteogenic inflammatory protein composition within the extracellular fluid of infected bone tissue was described. The findings support the current histopathological knowledge and, therefore, microdialysis may represent a valid method for sampling of material for protein investigation of the in vivo inflammatory composition within the extracellular environment in infected bone tissue.

TL1A regulates adipose-resident innate lymphoid immune responses and enables diet-induced obesity in mice.

BACKGROUND/OBJECTIVES: TL1A is a pro-inflammatory cytokine that is homologous to TNFα and connected with the development of several chronic inflammatory disorders. The preliminary results of this study indicated reduced fat accumulation in 9-month-old TL1A-deficient mice at steady state. Thus, the objective was to investigate whether TL1A-deficient mice are resistant to the development of high-fat (HF) diet-induced obesity and to investigate the impact on lymphocyte infiltration in adipose tissue. METHODS: TL1A-deficient and TL1A-sufficient male BALB/cJ littermate mice were fed a 60% HF diet or a 10% low-fat control diet for 22 weeks. Mouse body composition and weight were monitored, and tissues were processed and evaluated by flow cytometry, qPCR, and histology. RESULTS: In this study, the TL1A-deficient HF-diet-fed mice had reduced whole-body weight gain, which was directly explained by a corresponding fat mass reduction (average 37.2%), compared with that of their TL1A-sufficient littermates. Despite previous data showing marked changes in the gut microbial community, TL1A-deficient GF mice also displayed reduced adiposity. Furthermore, the TL1A-deficient mice were resistant to hepatic steatosis and were shown to have improved glucose tolerance, as determined by oral glucose tolerance test (OGTT), and greater insulin sensitivity. In the epididymal white adipose tissue (eWAT), TL1A deficiency in HF-diet-fed mice resulted in a reduced abundance of IL-18Ra+ type-1 ILCs and γδT cells as well as markedly reduced expression of the mitochondria-regulating genes Ucp1, Ucp2, Ucp3, and Prdm16. Finally, to investigate the link of TL1A to obesity in humans, we identified a noncoding polymorphism (rs4979453) close to the TL1A locus that is associated with waist circumference in men (p = 0.00096, n = 60586). CONCLUSIONS: These findings indicate that TL1A plays an important role in regulating adipose tissue mass and that this role is independent of the gut microbiota. Furthermore, we show that TL1A regulates adipose-resident innate lymphocytes and mitochondria-mediated oxidative stress in eWAT.

Staphylococcus aureus infected embolic stroke upregulates Orm1 and Cxcl2 in a rat model of septic stroke pathology.

OBJECTIVE: Ischaemic brain lesions and brain abscesses are frequent in both human and animal cases of septic embolic stroke. However, existing models of brain infection do not reflect central aspects of septic embolic stroke. Our aim was to compare septic and non-septic embolic stroke in order to identify gene expressions, inflammatory mediators and brain damage in a rat model. METHODS: We created precisely located focal brain infarcts in a rat model of Staphylococcus aureus infected embolic stroke. To cause septic embolic stroke we used a fibrin-rich embolus with bacteria, while every rat in the control group received a non-infected embolus. 64 rats were randomized to receive sham-surgery, sterile embolic stroke or septic embolic stroke. All groups were compared for brain pathology, mortality, gene expressions and inflammatory mediators using histology and reverse transcription quantitative real-time PCR. RESULTS: Although infarct volumes did not differ, septic embolic stroke caused higher mortality than sterile embolic stroke (p=  0.002). Brain abscesses were observed only in the septic group. Approximately 400-500 fold increases were observed for Orm1 and Cxcl2 respectively (1.00E-08 < p < 1.92E-07) in the septic group compared to the sterile group, and these were the most dramatically regulated genes in septic embolic stroke compared to sterile embolic stroke. CONCLUSIONS: Septic embolic stroke caused brain abscesses, increased mortality and upregulated Orm1 and Cxcl2 gene expressions compared to non-infected embolic stroke. The dramatic Orm1 increase observed in the septic group is unprecedented and suggests a significant biological role of Orm1 during septic neuroinflammation.

Isavuconazole in a Successful Combination Treatment of Disseminated Mucormycosis in a Child with Acute Lymphoblastic Leukaemia and Generalized Haemochromatosis: A Case Report and Review of the Literature.

Invasive mucormycosis in immunocompromised children is a life-threatening fungal infection. We report a case of a 7-year-old girl treated for acute lymphoblastic leukaemia complicated by disseminated mucormycosis during induction therapy. Microscopic examination of surgically removed lung tissue revealed wide, pauci-septate hyphae suggesting a Mucorales infection. This diagnosis was confirmed immunohistochemically and by PCR analysis followed by a final identification of Cunninghamella sp. The patient was treated successfully with surgical debridement and antifungal combination therapy with amphotericin B, caspofungin and isavuconazole. The use of isavuconazole in a child was not previously reported. Additionally, case reports concerning pulmonary mucormycoses in paediatric population published after 2010 were reviewed. Nineteen out of 26 identified patients suffered from haematological diseases. Reported mortality reached 38.5%. By the fact of rising morbidity, unsatisfactory results of treatment and remaining high mortality of mucormycoses in immunocompromised patients, new therapeutic options are warrant. Isavuconazole, with its broad-spectrum activity, good safety profile and favourable pharmacokinetics, is a promising drug. However, further studies are necessary to confirm positive impact of isavuconazole on mucormycosis treatment in children.

The intensity of the inflammatory response in experimental porcine bruises depends on time, anatomical location and sampling site.

The assessment of the age of bruises inflicted on livestock is an important component of veterinary forensic pathology investigations. However, the sampling site within a bruise, the anatomical location and the mass and speed of the object inflicting the blunt trauma might influence the intensity of the inflammatory reaction. In the present study, the variation of the inflammatory reaction within and along experimental porcine bruises was evaluated in order to determine the optimal sampling site. Moreover, we evaluated if a combination of histological characteristics and gene expression signatures was able to differentiate bruises according to anatomical location, age of bruises and the speed and mass of the object used to cause the impact. Twelve experimental slaughter pigs were anesthetized, and on each animal four blunt traumas were inflicted on the back using either a plastic tube or an iron bar, respectively. The pigs were euthanized at 2, 5 or 8 h after infliction. Following gross examination, skin and underlying muscle tissue were sampled from the center and both ends of bruises and evaluated histologically. Subcutaneous fat tissue from the center of the bruises was sampled for quantitative real-time polymerase chain reaction to evaluate mRNA expression of 13 selected genes. Uninjured tissue was sampled from the right thigh of all pigs and served as control tissue. The amount of tissue damage and the intensity of the inflammatory reaction in bruises depended on the sampling site within and along a bruise, the anatomical location and the age of the bruise. The optimal site for sampling, i.e. the most pronounced inflammatory reaction, was at the center of the bruises where the plastic tube or iron bar first struck the skin. Moreover, bruises inflicted in areas with a thin layer of subcutaneous fat tissue showed more damage and inflammation in the underlying muscle tissue compared to bruises inflicted in areas with a thicker layer of subcutaneous fat tissue. In addition, hemorrhage in the muscle tissue was more likely present when bruises were inflicted with an iron bar compared to a plastic tube. Combining histology and mRNA expression of the 13 genes showed that the age of bruises could be determined with a precision of ±2.04 h. Moreover, the age of bruises could be determined with a precision of ±1.84 h based solely on mRNA expression of a selection of four genes.

Pigs are useful for the molecular study of bone inflammation and regeneration in humans.

Pigs are used with increased frequency to model different kinds of orthopedic surgical conditions. In order to show the full potential of porcine models in orthopedic research, it is therefore required to examine the expression of bone regulatory genes in pigs affected by orthopedic surgery and compare it to the expression in humans and mice as mice, are one of the most applied animal species in orthopedics today. In the present study, the local molecular response to drilling of a tibial implant cavity, and the subsequent insertion of a steel implant was examined in a porcine model. Pigs were euthanized five days after drilling of the bone. The molecular response of 73 different genes was analyzed using a high-throughput quantitative polymerase chain reaction platform and compared to histopathology. Histologically, it was found that bone remodeling was initiated on day 5 after surgery and was associated with upregulation of several genes involved in bone degradation and formation ( CTSK, ACP5, IBSP, RANK, RANKL and COL1A1). Interleukin-6 and several acute-phase proteins (C3, SAA and ITIH4) were significantly upregulated, indicating their importance in the initial process of healing and osseointegration. All tested bone morphogenic proteins (BMP2, -4 and -7) including their inhibitor noggin were also significantly upregulated. Surprisingly, vascular endothelial growth factor A was not found to be regulated five days after surgery while several other vascular growth factors (ANGPT1, ANGPT2 and PTN) were upregulated. The pig was found to be a useful model for elucidation of bone regulatory genes in humans.

Scrotal granulomatous aspergillosis in a dromedary camel (Camelus dromedarius).

BACKGROUND: This report describes a case of primary subcutaneous aspergillosis in a 7-year-old neutered male dromedary camel (Camelus dromedarius). CASE PRESENTATION: The animal developed a large nodular lesion in the right scrotum two years after surgical intervention for neutering. The mass had a firm consistency and was painful at palpation. Histopathology revealed dermal granulomatous inflammation with a necrotic centre, surrounded by plasma cells, macrophages, neutrophils, and sparse fungal hyphae characterised by parallel cell walls, distinct septa, and dichotomous branching. Fungal culture was not performed, but a panel of mono- and polyclonal antibodies specific for different fungal genera identified the hyphae as Aspergillus sp. CONCLUSIONS: The occurrence of subcutaneous lesions is a rare manifestation of aspergillosis in animals, and this appears to be the first case reported in the dromedary camel.

Early implant-associated osteomyelitis results in a peri-implanted bacterial reservoir.

Implant-associated osteomyelitis (IAO) is a common complication in orthopedic surgery. The aim of this study was to elucidate how deep IAO can go into the peri-implanted bone tissue within a week. The study was performed in a porcine model of IAO. A small steel implant and either 104 CFU/kg body weight of Staphylococcus aureus or saline was inserted into the right tibial bone of 12 pigs. The animals were consecutively killed on day 2, 4 and 6 following implantation. Bone tissue around the implant was histologically evaluated. Identification of S. aureus was performed immunohistochemically on tissue section and with scanning electron microscopy and peptide nucleic acid in situ hybridization on implants. The distance of the peri-implanted pathological bone area (PIBA), measured perpendicular to the implant, was significantly larger in infected animals compared to controls (p = 0.0014). The largest differences were seen after 4 and 6 days of inoculation, where PIBA measurements of up to 6 mm were observed. Positive S. aureus bacteria were identified on implants and from 25 µm to 6 mm into PIBA. This is important knowledge for optimizing outcomes of surgical debridement in osteomyelitis.

The impact of force on the timing of bruises evaluated in a porcine model.

In animal models developed in order to estimate the age of bruises, focus has been on the changes over time and not considering the force used to inflict the trauma. In the present study, gross and histological changes in 2, 4, 6 and 8 h old bruises which were inflicted with a low, moderate and high force were compared. Twelve experimental pigs were randomly assigned to three groups of force (low, moderate and high force). All pigs were anesthetized, and on each animal four blunt traumas were inflicted on the back with the low, moderate or high force according to the groups. The pigs were kept in anesthesia for 2, 4, 6 or 8 h, after which they were euthanized, and skin and muscle tissues were sampled for histology. As control, two pigs were included. The gross appearance of bruises developed similarly until 0.5 h after infliction at which time the visibility of the bruises depended on the force. The infiltration of subcutaneous neutrophils depended on the time and force used which was confirmed by both manual evaluation and image analysis of immunostained skin sections. In the muscle tissue, the number of macrophages was found useful for age determination in bruises inflicted with the highest force. Therefore, when evaluating forensic cases of bruises in both human and veterinary pathology the impact of force and not only the timing should be taken into consideration.

Effects of Implant-Associated Osteomyelitis on Cefuroxime Bone Pharmacokinetics: Assessment in a Porcine Model.

BACKGROUND: The prolonged antibiotic therapy that is often needed for successful management of osteomyelitis may be related to incomplete penetration of antibiotics into the target site. The objective of this study was to assess the effects of implant-associated osteomyelitis on cefuroxime penetration into bone. METHODS: Implant-associated osteomyelitis using a Staphylococcus aureus strain was induced in the right tibia in ten pigs. After five days and following administration of 1500 mg of cefuroxime, measurements of cefuroxime were obtained using microdialysis for eight hours in the implant-related bone cavity, in the adjacent infected cancellous bone and infected subcutaneous tissue, and in healthy cancellous bone and subcutaneous tissue in the contralateral leg. Measurements of the corresponding free plasma concentrations were also obtained. The extent of the infection was assessed by postmortem computed tomography (CT) scans and cultures of blood, swabs, and bone specimens. RESULTS: Bone destruction was found in the implant cavities. No structural bone changes in the adjacent infected cancellous bone were visible on CT scans. S. aureus was grown on culture of specimens from all implant cavities and from eight of ten swabs and seven of ten bone samples from the infected bone. The areas under the concentration-time curves for the different tissues differed significantly, with the lowest area under the curve found in the implant cavity (analysis of variance; p < 0.001). Although not as notable as for the implant cavity, cefuroxime penetration into infected cancellous bone was incomplete but comparable with that in healthy bone. Despite poorer tissue penetration, slightly increased time with concentrations above the minimal inhibitory concentration (MIC) was achieved in the implant cavity up to MICs of 2 mg/L compared with the other tissues, but the time was shorter for higher MICs. CONCLUSIONS: Cefuroxime penetration into infected cancellous bone was incomplete but comparable with that in healthy bone. The destructive bone processes associated with acute osteomyelitis reduced cefuroxime penetration further. CLINICAL RELEVANCE: These findings support the general clinical perception that fast diagnosis and early initiation of antibiotics before the development of implant-associated cavities is important in nonsurgical management of acute osteomyelitis.

A novel, comprehensive, and reproducible porcine model for determining the timing of bruises in forensic pathology.

PURPOSE: Calculating the timing of bruises is crucial in forensic pathology but is a challenging discipline in both human and veterinary medicine. A mechanical device for inflicting bruises in pigs was developed and validated, and the pathological reactions in the bruises were studied over time in order to identify gross and histological parameters that may be useful in determining the age of a bruise. METHODS: The mechanical device was able to apply a single reproducible stroke with a plastic tube that was equivalent to being struck by a man. In each of 10 anesthetized pigs, four strokes that resulted in bruises were inflicted on the back. In addition, 2 control pigs were included in the study. The pigs were euthanized consecutively from 1 to 10 h after the infliction of bruises. Following gross evaluation, skin, and muscle tissues were sampled for histology. RESULTS: Grossly, the bruises appeared uniform and identical to the tramline bruises seen in humans and pigs subjected to blunt trauma. Histologically, the number of neutrophils in the subcutis, the number of macrophages in the muscle tissue, and the localization of neutrophils and macrophages in muscle tissue showed a time-dependent response. Combining these parameters, bruises could be grouped as being either less than 4 h old or between 4 and 10 h of age. Gross lesions and changes in the epidermis and dermis were inconclusive with respect to time determination. CONCLUSIONS: The model was reproducible and resembled forensic cases of bruises in pigs and humans. Therefore, the histological parameters are suitable for age determination of bruises in pigs and likely also in humans.

Specific Antibodies to Staphylococcus aureus Biofilm Are Present in Serum from Pigs with Osteomyelitis.

BACKGROUND: The Achilles heel in osteomyelitis is that bacteria, primarily Staphylococcus aureus, grow as a biofilm in the bone lesions. MATERIALS AND METHODS: In the present study, we explored the serum level of specific antibodies to S. aurues biofilm in porcine models of osteomyelitis. RESULTS: Significantly increased levels of antibodies towards the specific biofilm antigen SA0688 were measured in serum from pigs with S. aureus-associated acute and chronic osteomyelitis 5-7 and 10-14 days after inoculation, respectively. Simultaneously with raised antibody levels, an increase in serum interleukin 6 (IL 6) levels was also seen. CONCLUSION: The observed biofilm-specific antibody response represents a T-helper cell 17 (Th17) response and potentially a T-helper cell 1 (Th1) response. This is in agreement with previous studies in mice and rabbits speculating that S. aureus induces a Th1- and Th17-biased adaptive immune response, instead of a protective Th2 response, in order to evade the immune system, resulting in a chronic infection.

Intrabronchial Microdialysis: Effects of Probe Localization on Tissue Trauma and Drug Penetration into the Pulmonary Epithelial Lining Fluid.

Recent intrabronchial microdialysis data indicate that the respiratory epithelium is highly permeable to drugs. Of concern is whether intrabronchial microdialysis disrupts the integrity of the respiratory epithelium and thereby alters drug penetration into the pulmonary epithelial lining fluid (PELF). The objective of this study was to investigate the effect of intrabronchial microdialysis on the integrity of the bronchial epithelium. Microdialysis sampling in PELF in proximal (n = 4) and distal bronchi (n = 4) was performed after intravenous inulin and florfenicol administration in anaesthetized pigs. Inulin was used as a marker molecule of permeability of the epithelium, and florfenicol was used as test drug. Bronchial tissue was examined by histopathology (distal and proximal bronchi) and gene expression analysis (RT-qPCR, proximal bronchi) at the termination of the experiment (6.5 hr). The microdialysis probe caused overt tissue trauma in distal bronchi, whereas no histopathological lesions were observed in proximal bronchi. A moderate up-regulation of the pro-inflammatory cytokines IL1B, IL6 and acute-phase reactant serum amyloid A was seen in proximal bronchi surrounding the microdialysis probes suggesting initiation of an inflammatory response. The observed up-regulation is considered to have limited impact on drug penetration during short-term studies. Inulin penetrated the respiratory epithelium in both proximal and distal bronchi without any correlation to histopathological lesions. Likewise, florfenicol penetration into PELF was unaffected by bronchial histopathology. However, this independency of pathology on drug penetration may not be valid for other antibiotics. We conclude that short-term microdialysis drug quantification can be performed in proximal bronchi without disruption of tissue integrity.

Cancer resistance as an acquired and inheritable trait.

AIM: To induce cancer resistance in wild-type mice and detect if the resistance could be inherited to the progeny of the induced resistant mice. Furthermore to investigate the spectrum and immunology of this inherited cancer resistance. MATERIALS AND METHODS: Resistance to with live S180 cancer cells in BALB/c mice was induced by immunization with inactivated S180 cancer cells. The immunization was performed by either frozen/thawed or irradiated cancer cells or cell-free ascitic fluid (CFAF). RESULTS: In all instances the induced resistance was demonstrated to be inheritable. The phenotype was named HICR (heritable induced cancer resistance) and was defined as primary resistant progeny from mice immunized with frozen/thawed or irradiated S180 cells or CFAF obtained from mice with S180 induced ascites. Notably, this resistance was transferred from both male and female mice to the offspring of the immunized mice for at least two generations. Although inheritable, the frequency of cancer-resistant pups was lost over a few generations. Cells from the J774A.1 and RAW cancer cell lines did not induce inheritable cancer resistance, and C57BL/6 mice could not pass on cancer resistance fluorescence-activated cell sorting (FACS) analyses of the peritoneal cells revealed an increased fraction of macrophages. In necropsies of resistant mice no histological signs of cancer or other disease was found. CONCLUSION: Only materials derived from S180 cells could give rise to HICR mice. The molecular basis of the resistance is unknown but may involve epigenetic mechanisms. Other examples of inheritability of acquired phenotypic changes exist but, to our knowledge, this is the first demonstration of acquired, inherited cancer resistance.

Successful treatment of an intra-pelvic fungal pseudomycetoma causing constipation and hypercalcaemia in a Persian cat.

This case report describes the successful treatment of a Persian cat diagnosed with intra-abdominal fungal pseudomycetoma causing hypercalcaemia and constipation due to an extra-luminal mechanical obstruction of the colon. Treatment included surgical excision, supportive care and itraconazole for 6 months.

Functional characterization of a porcine emphysema model.

BACKGROUND: Lung emphysema is a central feature of chronic obstructive pulmonary disease (COPD), a frequent human disease worldwide. Cigarette smoking is the major cause of COPD, but genetic predisposition seems to be an important factor. Mutations in surfactant protein genes have been linked to COPD phenotypes in humans. Also, the catalytic activities of metalloproteinases (MMPs) are central in the pathogenesis of emphysema/COPD. Especially MMP9, but also MMP2, MMP7, and MMP12 seem to be involved in human emphysema. MMP12-/- mice are protected from smoke-induced emphysema. ITGB6-/- mice spontaneously develop age-related lung emphysema due to lack of ITGB6-TGF-ß1 regulation of the MMP12 expression. METHODS: A mutated pig phenotype characterized by age-related lung emphysema and resembling the ITGB6-/- mouse has been described previously. To investigate the emphysema pathogenesis in this pig model, we examined the expression of MMP2, MMP7, MMP9, MMP12, and TGF-ß1 by quantitative PCR (qPCR). In addition, immunohistochemical stainings of the lungs with SP-B, SP-C, MMP9, and MMP12 antibodies were performed. The haematologic/immunologic status of the pigs also was studied. RESULTS: The qPCR study showed no difference between pigs with and without emphysema, and no systemic differences were indicated by the haematologic and immunologic studies. However, the immunohistochemical stainings showed an increased expression of MMP9 and MMP12 in older, mutated pigs (with emphysema) compared with normal and young mutated pigs (without emphysema). CONCLUSIONS: The pig model is comparable to human emphysema patients and the ITGB6-/- mouse model with respect to both morphology and functionality.