In infectious pancreatic necrosis virus carrier Atlantic salmon, Salmo salar L., post-smolts, almost all kidney macrophages ex vivo contain a low level of non-replicating virus.

The level of infection by infectious pancreatic necrosis virus (IPNV) of kidney macrophages from 12 asymptomatic carrier Atlantic salmon post-smolts was studied. Kidney leucocytes were fractionated on 34/51% Percoll gradients, allowed to adhere to plastic wells overnight, washed to remove non-adherent cells and cultured for up to 7 days with or without renewal of medium on day 3. On day 1, supernatants were harvested, macrophages were counted, lysed and IPNV in the supernatants and lysates was titred in chinook salmon embryo (CHSE-214) cells. The multiplicity of infection ranged between 1:2.2 and 1:7.4 (virus:macrophages). On day 3, the titres of IPNV in macrophage lysates decreased and in wells where the medium was renewed on day 3, IPNV was no longer detectable on day 7. In the supernatants, one fish was positive for IPNV on day 1, four fish on day 3 but none were detectably positive on day 7. In parallel wells in which the medium was not renewed, on day 7 IPNV was detected in macrophage lysates of three fish and the supernatants were also IPNV positive in two of these fish. This suggests that virus might be shed from infected macrophages and then reinfect other macrophages. When macrophages were serially diluted in wells and cultured for 24 h, IPNV could be cultured from macrophage lysates of wells containing between two and 70 macrophages. These results indicate that a very high proportion of the adherent kidney macrophages must be infected with very few non-replicating virions.

A non-destructive test for detection of IPNV-carriers in Atlantic halibut, Hippoglossus hippoglossus (L.).

Over 18 months after infectious pancreatic necrosis virus (IPNV) was first detected in fish (80 g-4 kg) on a halibut farm, the stocks were found to be still carrying the virus. This suggests that long-term persistence of IPNV occurs in farmed Atlantic halibut. A non-destructive test was applied to blood adherent leucocytes by placing 100 microL of whole blood collected in a heparinized tube into 96-well plates. After overnight incubation, the non-adherent cells were washed off, the remaining adherent cells lysed in a lysis buffer and inoculated onto CHSE-214 cells. The resulting cytopathic effect was confirmed as IPNV positive by enzyme-linked immunosorbent assay. In a sample of 10 fish tested by this method, all were positive for IPNV while only two were positive by the standard method for virus culture from sonicated kidney homogenates and only one fish, which was positive by the standard method, was positive by reverse transcription polymerase chain reaction on kidney tissue. The test on blood leucocytes is shown to be simple to perform on samples taken under field conditions.

A sensitive non-destructive method for detecting IPNV carrier Atlantic salmon, Salmo salar L., by culture of virus from plastic adherent blood leucocytes.

In populations of Atlantic salmon in sea water, infectious pancreatic necrosis virus (IPNV) could be detected by standard virological culture methods in sonicated kidney homogenates and in mucus samples (gill, skin and rectum) from 14 and nine of 25 fish, respectively, but all fish were positive by virus culture from lysates of kidney macrophages and adherent blood leucocytes. In fish which tested negative for IPNV by the standard method of detection, the virus could be detected using adherent blood leucocytes isolated on a Percoll gradient from as little as 10 microL of blood. The blood sample could be stored for at least 3 days in a heparinized tube on ice before preparing the plastic adherent leucocytes. Furthermore, the latter could be prepared without prior fractionation on Percoll simply by incubating whole blood (33 microL) in cell culture medium (66 microL) in 96-well plates overnight and washing away the non-adherent cells before lysing the adherent cells and inoculation of the lysate onto CHSE-214 cells. This highly sensitive method for detecting IPNV-carriers is therefore very suitable for non-destructive sampling of fish in the field.

Septic peritonitis secondary to unilateral pyometra and ovarian bursal abscessation in a dog.

A seven-year-old, female entire Labrador retriever was presented for acute-onset vomiting and lethargy, associated with weakness and generalised tremors. The clinical, radiographic, ultrasonographic and histopathological findings revealed septic peritonitis which occurred secondarily to unilateral pyometra and ovarian bursal abscessation. However, in this case, the initial clinical findings, blood parameters, radiographic and ultrasonographic findings did not allow a specific diagnosis. Repeat monitoring was required, and abdominocentesis proved to be the most useful diagnostic test, allowing a definitive diagnosis and the decision to be made as to whether or not to carry out exploratory surgery.

Myasthenia gravis associated with cutaneous lymphoma in a dog.

A middle-aged golden retriever presenting with exercise intolerance and multifocal cutaneous nodules was diagnosed as having non-epitheliotropic cutaneous lymphoma with concurrent myasthenia gravis. Treatment with corticosteroids induced short-term remission of the lymphoma and alleviation of myasthenic signs.

Aetiology and diagnosis of persistent nasal disease in the dog: a retrospective study of 42 cases.

Forty-two dogs with a history of persistent nasal disease were evaluated by a combination of clinical examination, thoracic and nasal radiography, retroflexed endoscopy and biopsy, and anterograde rhinoscopy and blind nasal biopsy. A definitive diagnosis was made in 91 per cent of cases. Neoplasia was the most common diagnosis (33 per cent of cases), followed by inflammatory rhinitis (24 per cent). Other diagnoses included periodontal disease (10 per cent), aspergillosis (7 per cent) and foreign bodies (7 per cent). Adenocarcinoma was the most common tumour diagnosed. The clinical findings were found to be too variable to be used as specific diagnostic criteria. Anterograde rhinoscopy and retroflexed endoscopy had higher specificity and sensitivity than radiology for the diagnosis of neoplasia, inflammatory rhinitis, aspergillosis and foreign bodies. With a systematic approach to the investigation of persistent nasal disease, a definitive diagnosis can be successfully obtained in the vast majority of cases.

Osteomyelitis of the atlanto-occipital region as a sequela to a pharyngeal stick injury.

An eight-year-old female Belgian shepherd dog was referred for investigation of chronic neck pain. The dog had sustained a pharyngeal injury 12 weeks previously while catching a stick. Radiographs of the cervical spine revealed signs consistent with a septic arthritis of the atlanto-occipital joint and osteomyelitis of both occipital condyles and the atlas. A foreign body was identified ultrasonographically in the retropharyngeal soft tissues, and a stick was surgically removed from a site ventral to the right side of the atlanto-occipital joint. The signs of neck pain started to resolve within a week of surgery.

Depolarization and cAMP elevation rapidly recruit TrkB to the plasma membrane of CNS neurons.

Here, we describe a novel mechanism for the rapid regulation of surface levels of the neurotrophin receptor TrkB. Unlike nodose ganglion neurons, both retinal ganglion cells (RGCs) and spinal motor neurons (SMNs) in culture display only low levels of surface TrkB, though high levels are present intracellularly. Within minutes of depolarization or cAMP elevation, surface TrkB levels increase by nearly 4-fold, and this increase is not blocked by cycloheximide. These findings suggest that activity and cAMP elevation rapidly recruit TrkB to the plasma membrane by translocation from intracellular stores. We propose that a fundamental difference between peripheral nervous system (PNS) and central nervous system (CNS) neurons is the activity dependence of CNS neurons for responsiveness to their peptide trophic factors and that differences in membrane compartmentalization of the receptors underlie this difference.

Successful non operative management of late infrainguinal polytetrafluoroethylene vascular graft infection with antibiotics alone: a report of two cases.

Polytetrafluoroethylene (PTFE) prosthetic vascular graft infection is a serious complication of arterial reconstructive surgery. Traditional management is primarily surgical and may result in long hospital stay with significant morbidity and mortality. The authors describe two cases of infected femoropoliteal PTFE grafts which were managed non operatively with antibiotic therapy alone, resulting in early return to normal activities and excellent resolution of infection on Indium labelled leucocyte scanning. The authors suggest that in selected cases antibiotic therapy alone is a valuable tool in the management of infrainguinal PTFE prosthetic sepsis.

Inhibition of human vascular smooth muscle cell proliferation by lovastatin: the role of isoprenoid intermediates of cholesterol synthesis.

Restenosis remains the largest single obstacle to the long-term success of invasive vascular interventions. Lovastatin, an HMG-CoA reductase inhibitor, has been shown to reduce myointimal hyperplasia in animal models of restenosis and in one clinical coronary restenosis trial. We have assessed the effect of lovastatin on the growth of cultured human vascular smooth muscle cells derived from saphenous vein and vascular graft stenoses. Lovastatin (2 microM) inhibited proliferation over 14 days in saphenous vein (and graft stenoses) derived vascular smooth muscle cells by 42% and 32% respectively: this was not significantly different. Lovastatin (10 microM) reduced [methyl 3H]-thymidine uptake by 51% in saphenous vein-derived cells. These concentrations were significantly higher than those achieved in plasma during therapeutic dosage. Lovastatin-induced inhibition of vascular smooth muscle cell proliferation and [methyl 3H]-thymidine uptake was completely reversed by adding mevalonate (100 microM) but cholesterol (10-40 micrograms ml-1) had no effect. Isopentenyl adenine (25-50 microM) did not affect the inhibition of [methyl 3H]-thymidine uptake by lovastatin (10 microM), but farnesol (20 microM), another isoprenoid precursor of cholesterol synthesis, reversed the antiproliferative effect.

Consistent responses of the human vascular smooth muscle cell in culture: implications for restenosis.

PURPOSE: The mechanisms whereby restenoses occur at discrete sites within the vasculature remain uncertain. We have recently reported that vascular smooth muscle cells (VSMC) derived from patients with graft stenoses are resistant to growth inhibition by heparin. In this study, we have examined whether VSMC proliferation rates and responses to inhibition by heparin vary according to the individual or the anatomic site of origin. METHODS: Long saphenous veins from seven patients were divided into proximal, middle, and distal portions, and VSMC were cultured separately from each. VSMC proliferation in response to 15% fetal calf serum +/- 100 micrograms/ml heparin was measured by counting triplicate samples at 0, 3, 7, 10, and 14 days. This experiment was repeated from the second to the sixth passage (n = 6) and for artery and vein pairs derived from four additional patients. RESULTS: Differences between vein segment cultures of individual veins were found not to differ significantly from experimental error for either proliferation or heparin inhibition and were not altered by repeated passage (ANOVA). There were, however, significant differences in sensitivity to heparin inhibition between patients (p = 0.02) (ANOVA). There were no significant differences between paired samples of artery and vein for either proliferation or heparin inhibition (Mann-Whitney test). CONCLUSIONS: VSMC growth characteristics reflect the individual patient and are maintained in cell culture.

[Gynandroblastoma of the ovary. Review of the literature]. / Gynandroblastome de l'ovaire. Revue de la littérature.

Gynadroblastoma is an extremely rare tumour of the ovary containing both a granulosa cell and a Sertoli-Leydig cell tumour. We report a case of gynandroblastoma of the right ovary Stage IA (FIGO) in a 15-year-old girl. Microscopically, the tumour composed predominantly of granulosa cells with a minoritary component, although more than 10%, of Sertoli cells. The present knowledge about its nature, function and behaviour are reviewed.

Cellular biology of human intimal hyperplastic stenosis.

Restenosis after angioplasty and vascular surgery remains a major unsolved clinical problem. Vascular smooth muscle cell (VSMC) hyperplasia is an invariable response, but in 20-50% of cases proceeds to compromise the vessel lumen. We sought to identify cellular characteristics of human VSMC which are associated with restenosis. Human VSMC were grown from 135 samples of vascular tissue derived from patients undergoing primary cardiovascular surgery and revision surgery for restenosis. Cells derived from normal vein and artery, atherosclerotic plaques and from stenotic lesions were studied for successful proliferation in cell culture. Furthermore, growth rates were measured in response to 15% foetal calf serum +/- inhibition with heparin (100 micrograms/ml). Significantly fewer cells from atherosclerotic plaques progress to the third passage in cell culture than those derived from stenoses and controls (p < 0.001, Chi square) and growth rates after the third passage could not be studied in these cells. Of cells that progress to this stage, growth rates do not differ between stenosis-derived and normal cells under standard conditions. VSMC from mature atherosclerotic plaques may have undergone senescent changes. Stenosis-derived cells do not grow more rapidly than normal cells, but are significantly less sensitive to heparin (p < 0.001, Mann-Witney test), which is a major physiological inhibitor of VSMC growth. Differences in biological characteristics of human VSMC, observed in cell culture, may provide important insights into human vascular disease.

Abnormal growth regulation of vascular smooth muscle cells by heparin in patients with restenosis.

Proliferation of vascular smooth muscle cells (VSMC) underlies myointimal hyperplasia, which can lead to restenosis after angioplasty and vascular surgery. We propose that some individuals have an intrinsic capacity for this exaggerated response to vascular injury, partly through decreased sensitivity to the physiological growth inhibitor heparin. We investigated the effect of heparin on VSMC from restenotic lesions and from apparently normal vessels of the same patients, and VSMC from control patients undergoing primary bypass procedures. Cells from patients with restenosis (both restenotic lesion and undiseased vein) showed much lower sensitivity to growth inhibition by heparin than the controls (median inhibition 8 [95% Cl -2 to 25] vs 22 [15-44]%, p < 0.001); this finding suggests aberrant growth regulation in these cells.

Regional localization of human ecto-5' nucleotidase to chromosome 6q14-q21.

Human and mouse hybrids that contain fragments of human chromosome 6 as translocations were analysed for expression of ecto-5' nucleotidase enzymic activity measured by the conversion of AMP to adenosine and for antigenicity recognized by a monoclonal antibody specific for the human isozyme. Both methods allow a regional assignment of ecto-5'nucleotidase to 6q14-q21.

Duchenne muscular dystrophy in one of monozygotic twin girls.

Monozygotic twin girls are reported, one of whom has the typical clinical features of Duchenne muscular dystrophy despite a normal female karyotype. Although certain features of the biopsy were atypical, the clinical diagnosis was supported by persistent markedly raised blood creatine kinase levels and findings typical of DMD on electromyography and magnetic resonance spectroscopy. Analysis of an X linked DNA polymorphism in 16 independent somatic cell hybrids made between cells derived from each girl and a mouse line suggest that in one twin only the maternal X chromosome is active, whereas in the other the active X was paternally derived. More data are needed to exclude sampling error. These preliminary experimental results support the hypothesis that both girls are heterozygous for Duchenne muscular dystrophy. X inactivation, by chance, resulted in two contrasting cell masses with different active X chromosomes. This segregation was followed by, and may even have resulted in, twinning into a female pair, one normal and one with the full clinical features of the disease.