Hypoxia aggravates non-alcoholic steatohepatitis in mice lacking hepatocellular PTEN

The metabolic disorders that predispose patients to NASH (non-alcoholic steatohepatitis) include insulin resistance and obesity. Repeated hypoxic events, such as occur in obstructive sleep apnoea syndrome, have been designated as a risk factor in the progression of liver disease in such patients, but the mechanism is unclear, in particular the role of hypoxia. Therefore we studied the influence of hypoxia on the development and progression of steatohepatitis in an experimental mouse model. Mice with a hepatocellular-specific deficiency in the Pten (phosphatase and tensin homologue deleted on chromosome 10) gene, a tumour suppressor, were exposed to a 10% O2 (hypoxic) or 21% O2 (control) atmosphere for 7 days. Haematocrit, AST (aspartate aminotransferase), glucose, triacylglycerols (triglycerides) and insulin tolerance were measured in blood. Histological lesions were quantified. Expression of genes involved in lipogenesis and mitochondrial beta-oxidation, as well as FOXO1 (forkhead box O1), hepcidin and CYP2E1 (cytochrome P450 2E1), were analysed by quantitative PCR. In the animals exposed to hypoxia, the haematocrit increased (60+/-3% compared with 50+/-2% in controls; P<0.01) and the ratio of liver weight/body weight increased (5.4+/-0.2% compared with 4.7+/-0.3% in the controls; P<0.01). Furthermore, in animals exposed to hypoxia, steatosis was more pronounced (P<0.01), and the NAS [NAFLD (non-alcoholic fatty liver disease) activity score] (8.3+/-2.4 compared with 2.3+/-10.7 in controls; P<0.01), serum AST, triacylglycerols and glucose were higher. Insulin sensitivity decreased in mice exposed to hypoxia relative to controls. The expression of the lipogenic genes SREBP-1c (sterol-regulatory-element-binding protein-1c), PPAR-gamma (peroxisome-proliferator-activated receptor-gamma), ACC1 (acetyl-CoA carboxylase 1) and ACC2 (acetyl-CoA carboxylase 2) increased significantly in mice exposed to hypoxia, whereas mitochondria beta-oxidation genes [PPAR-alpha (peroxisome-proliferator-activated receptor-alpha) and CPT-1 (carnitine palmitoyltransferase-1)] decreased significantly. In conclusion, the findings of the present study demonstrate that hypoxia alone aggravates and accelerates the progression of NASH by up-regulating the expression of lipogenic genes, by down-regulating genes involved in lipid metabolism and by decreasing insulin sensitivity.

Ablation of the tumor suppressor histidine triad nucleotide binding protein 1 is protective against hepatic ischemia/reperfusion injury

The identification of cellular pathways capable of limiting ischemia/reperfusion (I/R) injury remains a frontier in medicine, and its clinical relevance is urgent. Histidine triad nucleotide binding protein 1 (HINT1) is a tumor suppressor that influences apoptosis. Because apoptotic pathways are a feature of I/R injury, we asked whether Hint1 influences hepatic I/R injury. Hint1(-/-) and C57BL/6 mice were subjected to 70% liver ischemia followed by reperfusion for 3 or 24 hours or to a sham operation. The serum aminotransferase levels, histological lesions, apoptosis, reactive oxygen species, and expression of B cell lymphoma 2-associated X protein (Bax), heme oxygenase 1 (HO-1), interleukin-6 (IL-6), IL-10, tumor necrosis factor-a, Src, nuclear factor kappa B (p65/RelA), and c-Jun were quantified. The responses to toll-like receptor ligands and nicotinamide adenine dinucleotide phosphate oxidase activity in Kupffer cells were compared in Hint1(-/-) mice and C57BL/6 mice. After I/R, the levels of serum aminotransferases, parenchymal necrosis, and hepatocellular apoptosis were significantly lower in Hint1(-/-) mice versus control mice. Furthermore, Bax expression decreased more than 2-fold in Hint1(-/-) mice, and the increases in reactive oxygen species and HO-1 expression that were evident in wild-type mice after I/R were absent in Hint1(-/-) mice. The phosphorylation of Src and the nuclear translocation of p65 were increased in Hint1(-/-) mice, whereas the nuclear expression of phosphorylated c-Jun was decreased. The levels of the protective cytokines IL-6 and IL-10 were increased in Hint1(-/-) mice. These effects increased survival after I/R in mice lacking Hint1. Hint1(-/-) Kupffer cells were less activated than control cells after stimulation with lipopolysaccharides. CONCLUSION: The Hint1 protein influences the course of I/R injury, and its ablation in Kupffer cells may limit the extent of the injury.

Multiple small "imaging" branch-duct type intraductal papillary mucinous neoplasms (IPMNs) in familial pancreatic cancer: indicator for concomitant high grade pancreatic intraepithelial neoplasia?

Most screening programs for familial pancreatic cancer are currently based on endoscopic ultrasonography and/or magnetic resonance imaging (MRI). Cystic lesions, especially those suspicious for small intraductal pancreatic mucinous neoplasms (IPMNs) of the branch ducts, can be visualized in up to 40 % of individuals at risk, but their pathological importance in the setting of FPC is yet not well established. Individuals at risk from a prospective screening program for familial pancreatic cancer with small "imaging" IPMNs of the branch-duct type (BD-IPMN) who underwent pancreatic resection were analysed regarding clinico-pathological data and the locations of pancreatic lesions. Five of 125 individuals at risk who underwent screening had multiple small (size 2-10 mm) unicystic lesions and/or multicystic single lesions in the pancreatic body and tail suspicious for BD-IPMNs upon MRI imaging and decided to undergo surgical resection after interdisciplinary counselling, although none fulfilled the consensus criteria for IPMN resection. Histological examination revealed BD-IPMNs with low or moderate dysplasia of the gastric type in combination with multifocal PanIN2 and PanIN3 lesions in 4 individuals. The remaining patient had only tiny ductectasias in the pancreatic tail with multifocal PanIN 2 lesions in the entire gland and one PanIN3 lesion in the pancreatic head. Intriguingly, the location of the most dysplastic histological lesions (PanIN3) did not correspond to the preoperatively detected lesions and were not visible in preoperative imaging. In the setting of FPC, the presence of multiple small "imaging" BD-IPMNs may indicate the presence of high-grade PanIN lesions elsewhere in the pancreas.

[Treatment of nonalcoholic steatohepatitis]

NASH associates steatosis with parenchymal inflammation and signs of hepatocellular injuy and even apoptosis. This leads in a minority of patients to fibrosis and in the long term to cirrhosis. NASH regularly occurs in a metabolic context characterized by insulin resistance. Several drugs have been tested in randomized controlled studies. Glitazones improve insulin resistance and also NASH, but are associated with side effects particularly unwelcome in NASH patients. Ursodesoxycholic acid, an hydrophilic biliary acid with hepatoprotective properties, does not improve the histological lesions of NASH. Vitamin E is the only compound which showed so far a positive effect without relevant side effects. However, it is too early to recommend its long-term use in this indication. Finally, the best treatment is not with drugs, but relies on behavioural changes: NASH patients should regularly exercise!

Histopathological alterations, EROD activity, CYP1A protein and biliary metabolites in gilthead seabream Sparus aurata exposed to benzo(a)pyrene

This study compared for seabream, Sparus aurata exposed to benzo(a)pyrene-B(a)P-, the response of molecular cytochrome P450 1A (CYP1A) and cellular histopathology biomarkers. Male gilthead seabream, Sparus aurata specimens were exposed for 20 days via water to a series of high B(a)P concentrations. CYP1A was assessed by measuring enzymatic activity (EROD) and CYP1A protein content, and cellular responses were evaluated by routine histopathological methods. In addition, biliary metabolites were measured in order to verify that B(a)P was absorbed and metabolised. Histological lesions, both in liver and gills, increased in parallel to B(a)P concentrations, with the majority of changes representing rather non-specific alterations. Hepatic EROD and CYP1A proteins data showed a concentration-dependent induction, while in the gills, EROD activity but not CYP1A proteins showed a non-monotonous dose response, with a maximum induction level at 200 microg B(a)P.L-1 and decreasing levels thereafter. The findings provide evidence that short-term, high dose exposure of fish can result in significant uptake and metabolism of the lipophilic B(a)P, and in pronounced pathological damage of absorptive epithelia and internal organs.

Irradiation of intracerebral 9L gliosarcoma by a single array of microplanar x-ray beams from a synchrotron: balance between curing and sparing

The purpose of this work was the understanding of microbeam radiation therapy at the ESRF in order to find the best compromise between curing of tumors and sparing of normal tissues, to obtain a better understanding of survival curves and to report its efficiency. This method uses synchrotron-generated x-ray microbeams. Rats were implanted with 9L gliosarcomas and the tumors were diagnosed by MRI. They were irradiated 14 days after implantation by arrays of 25 microm wide microbeams in unidirectional mode, with a skin entrance dose of 625 Gy. The effect of using 200 or 100 microm center-to-center spacing between the microbeams was compared. The median survival time (post-implantation) was 40 and 67 days at 200 and 100 microm spacing, respectively. However, 72% of rats irradiated at 100 microm spacing showed abnormal clinical signs and weight patterns, whereas only 12% of rats were affected at 200 microm spacing. In parallel, histological lesions of the normal brain were found in the 100 microm series only. Although the increase in lifespan was equal to 273% and 102% for the 100 and 200 microm series, respectively, the 200 microm spacing protocol provides a better sparing of healthy tissue and may prove useful in combination with other radiation modalities or additional drugs.

Small ruminant lentivirus genotype B and E interaction: evidences on the role of Roccaverano strain on reducing proviral load of the challenging CAEV strain

Live attenuated vaccines provide the most consistent protective immunity in experimental models of lentivirus infections. In this study we tested the hypothesis that animals infected with a naturally attenuated small ruminant lentivirus field strain of genotype E may control a challenge infection with a virulent strain of the caprine arthritis encephalitis virus (CAEV-CO). Within genotype E, Roccaverano strain has been described as attenuated since decreased arthritic pathological indexes were recorded in Roccaverano-infected animals compared to animals of the same breed infected with genotype B strains. Moreover, under natural conditions, animals double-infected with genotypes B and E appear less prone to develop SRLV-related disease, leading to a putative protective role of Roccaverano strain. Here we present evidence that goats experimentally infected with the avirulent genotype E SRLV-Roccaverano strain control the proviral load of a pathogenic challenge virus (CAEV-CO strain) more efficiently than naïve animals and appear to limit the spread of histological lesions to the contralateral joints.

Proliferative kidney disease in brown trout – infection level, pathology and mortality under field conditions

Proliferative kidney disease (PKD) is an emerging disease threatening wild salmonid populations. In temperature-controlled aquaria, PKD can cause mortality rates of up to 85% in rainbow trout. So far, no data about PKD-related mortality in wild brown trout Salmo trutta fario are available. The aim of this study was to investigate mortality rates and pathology in brown trout kept in a cage within a natural river habitat known to harbor Tetracapsuloides bryosalmonae. Young-of-the-year (YOY) brown trout, free of T. bryosalmonae, were exposed in the River Wutach, in the northeast of Switzerland, during 3 summer months. Samples of wild brown trout caught by electrofishing near the cage location were examined in parallel. The incidence of PKD in cage-exposed animals (69%) was not significantly different to the disease prevalence of wild fish (82 and 80% in the upstream and downstream locations, respectively). The mortality in cageexposed animals, however, was as low as 15%. At the termination of the exposure experiment, surviving fish showed histological lesions typical for PKD regression, suggesting that many YOY brown trout survive the initial infection. Our results at the River Wutach suggest that PKD in brown trout does not always result in high mortality under natural conditions.

Apoptosis in Bovine viral diarrhea virus (BVDV)-induced mucosal disease lesions: a histological, immunohistological, and virological investigation

Cattle persistently infected with a noncytopathic Bovine viral diarrhea virus (BVDV) are at risk of developing fatal "mucosal disease" (MD). The authors investigated the role of various apoptosis pathways in the pathogenesis of lesions in animals suffering from MD. Therefore, they compared the expression of caspase-3, caspase-8, caspase-9, and Bcl-2L1 (Bcl-x) in tissues of 6 BVDV-free control animals, 7 persistently infected (PI) animals that showed no signs of MD (non-MD PI animals), and 11 animals with MD and correlated the staining with the localization of mucosal lesions. Caspase-3 and -9 staining were markedly stronger in MD cases and were associated with mucosal lesions, even though non-MD PI animals and negative controls also expressed caspase-9. Conversely, caspase-8 was not elevated in any of the animals analyzed. Interestingly, Bcl-x also colocalized with mucosal lesions in the MD cases. However, Bcl-x was similarly expressed in tissues from all 3 groups, and thus, its role in apoptosis needs to be clarified. This study clearly illustrates ex vivo that the activation of the intrinsic, but not the extrinsic, apoptosis pathway is a key element in the pathogenesis of MD lesions observed in cattle persistently infected with BVDV. However, whether direct induction of apoptosis in infected cells or indirect effects induced by the virus are responsible for the lesions observed remains to be established.

In situ and in vitro comparison of laser fluorescence with visual inspection in detecting occlusal caries lesions

The aim of this study was to compare the in situ and in vitro performances of a laser fluorescence (LF) device (DIAGNOdent 2095) with visual inspection for the detection of occlusal caries in permanent teeth. Sixty-four sites were selected, and visual inspection and LF assessments were carried out, in vitro, three times by two independent examiners, with a 1-week interval between evaluations. Afterwards, the occlusal surfaces were mounted on the palatal portion of removable acrylic orthodontic appliances and placed in six volunteers. Assessments were repeated and validated by histological analysis of the tooth sections under a stereomicroscope. For both examiners, the highest intra-examiner values were observed for the visual inspection when in vitro and in situ evaluations were compared. The inter-examiner reproducibility varied from 0.61 to 0.64, except for the in vitro assessment using LF, which presented a lower value (0.43). The methods showed high specificity at the D(1) threshold (considering enamel and dentin caries as disease). In vitro evaluations showed the highest values of sensitivity for both methods when compared to the in situ evaluations at D(1) and D(2) (considering only dentinal caries as the disease) thresholds. For both methods, the results of sensitivity (at D(1) and D(2)) and accuracy (at D(1)) showed significant differences between in vitro and in situ conditions. However, the sensitivity (at D(1) and D(2)), specificity and accuracy (both at D(1)) of the methods were not significantly different when the same condition was considered. It can be concluded that visual inspection and LF showed better performance in vitro than in situ.

Endoscopic resection of laryngeal and tracheal lesions using the microdebrider

CONCLUSION: Endoscopic resection of laryngeal and tracheal lesions using the microdebrider is a safe, accurate and reliable method. OBJECTIVE: The microdebrider is an important tool for endoscopic nasal and sinus surgery and over the last few years a powered blade with a long shaft has been developed for endoscopic laryngeal and tracheal surgery. The aim of this non-randomized prospective study was to determine the advantages and disadvantages of the microdebrider for treating patients with different laryngeal and tracheal pathologies. MATERIAL AND METHODS: The laryngeal microdebrider was used under endoscopic control in 37 patients. In 29 cases a benign laryngeal lesion was removed endoscopically. In four patients debulking of a malignant obstructive endolaryngeal tumor was performed in order to avoid a tracheotomy. In four cases a bulky obstructing endotracheal lesion was removed. RESULTS: All laryngotracheal lesions could be removed, and this was facilitated by the use of angled rigid telescopes and the laryngeal blade. No traumatic lesions to normal laryngeal tissue occurred as a result of use of the microdebrider and no postoperative endolaryngeal bleeding was observed. The histological diagnosis of the biopsies taken with the microdebrider was accurate in every case. In three of the four cases with obstructive laryngeal malignancies, a tracheotomy was avoided until definitive therapy was undertaken. Normal breathing was restored in all patients with endotracheal lesions.

MRI characteristics and histology of bone marrow lesions in dogs with experimentally induced osteoarthritis

Signal changes within the bone marrow adjacent to osteoarthritic joints are commonly seen on magnetic resonance (MR) images in humans and in dogs. The histological nature of these lesions is poorly known. In this study, we describe the MR imaging of bone marrow lesions adjacent to the stifle joints of dogs with experimental osteoarthritis over 13 months. Histology of the proximal tibia at the end of the study was compared with the last MR imaging findings. In five adult dogs, the left cranial cruciate ligament was transected. Post-operatively, MR imaging was performed at 1, 2, 3, 4, 6, 8, and 13 months. Dogs were euthanised after 13 months and histological specimen of the proximal tibia were evaluated. Bone marrow edema like MR imaging signal changes were seen in every MR examination of all dogs in one or more locations of the proximal tibia and the distal femur. Lesions varied in size and location throughout the whole study with the exception of constantly seen lesions in the epiphyseal and metaphyseal region at the level of the tibial eminence. On histology, hematopoiesis and myxomatous transformation of the bone marrow and/or intertrabecular fibrosis without signs of bone marrow edema were consistent findings in the areas corresponding to the MR imaging signal changes. We conclude that within the bone marrow, zones of increased signal intensity on fat suppressed MR images do not necessarily represent edema but can be due to cellular infiltration. Contrary to humans, hematopoiesis is seen in bone marrow edema-like lesions in this canine model of osteoarthritis.

Mucosal lesions in a sheep infected with the Border Disease Virus (BDV)

A 28-week-old sheep was presented at the animal hospital because of chronic emaciation, anemia and slight diarrhea. Due to poor general condition and bad prognosis the animal was euthanized and submitted for postmortem investigation. Multiple erosions and ulcerations were found in the dorsal region of the tongue, the pharynx, the hard palate, in the esophagus and the ruminal pillars. Histologically, these lesions consisted of necrosuppurative inflammation. The animal was tested positive for pestivirus antigen both by immunohistochemical and by virological examination (cell culture, antigen capture ELISA and RT-PCR). A non-cytopathic Border Disease Virus was identified, and sequencing revealed a virus belonging to the BDV-3 cluster. Based on the macroscopical, histological, immunohistological and virological results this case was diagnosed as Border Disease with mucosal lesions. This is the first report of such a case in Switzerland.

Multiple osteosclerotic lesions in an Iron Age skull from Switzerland (320 - 250 BC) - An unusual case

The single Hochdorf burial was found in 1887 during construction work in the Canton of Lucerne, Switzerland. It dates from between 320 and 250 BC. The calvarium, the left half of the pelvis and the left femur were preserved. The finding shows an unusual bony alteration of the skull. The aim of this study was to obtain a differential diagnosis and to examine the skull using various methods. Sex and age were determined anthropologically. Radiological examinations were performed with plain X-ray imaging and a multislice computed tomography (CT) scanner. For histological analysis, samples of the lesion were taken. The pathological processing included staining after fixation, decalcification, and paraffin embedding. Hard-cut sections were also prepared. The individual was female. The age at death was between 30 and 50 years. There is an intensely calcified bone proliferation at the right side of the os frontalis. Plain X-ray and CT imaging showed a large sclerotic lesion in the area of the right temple with a partly bulging appearance. The inner boundary of the lesion shows multi-edged irregularities. There is a diffuse thickening of the right side. In the left skull vault, there is a mix of sclerotic areas and areas which appear to be normal with a clear differentiation between tabula interna, diploë and tabula externa. Histology showed mature organised bone tissue. Radiological and histological findings favour a benign condition. Differential diagnoses comprise osteomas which may occur, for example, in the setting of hereditary adenomatous polyposis coli related to Gardner syndrome.

Cutaneous reactive angiomatosis with combined histological pattern mimicking a cellulitis

Cutaneous reactive angiomatoses (CRA) encompass a distinct group of rare benign reactive vascular proliferations that include reactive angioendotheliomatosis, diffuse dermal angiomatosis and reactive intralymphatic histiocytosis. The etiology of these conditions, often associated with either localized or systemic diseases, is poorly understood. We report a 72-year-old woman who presented giant diffuse cellulitis-like plaques on the right lower limb and the pelvis and a reduction of her general condition with fever. Light microscopy studies revealed combined features of reactive angioendotheliomatosis, diffuse dermal angiomatosis and reactive intralymphatic histiocytosis. A small arteriovenous fistula of the right lower leg was thought to act as trigger. Systemic corticosteroids resulted in the clinical remission of the skin lesions. Our observation provides strong evidence that reactive angioendotheliomatosis, diffuse dermal angiomatosis and reactive intralymphatic histiocytosis, previously regarded as distinct forms of CRA, may show overlapping histopathological features and most likely represent facets of the same disease.