Uncertainties in carbon dioxide radiative forcing in atmospheric general circulation models.

Global warming caused by an increase in the concentrations of greenhouse gases, is the direct result of greenhouse gas-induced radiative forcing. When a doubling of atmospheric carbon dioxide is considered, this forcing differed substantially among 15 atmospheric general circulation models. Although there are several potential causes, the largest contributor was the carbon dioxide radiation parameterizations of the models.

Effect of prostaglandin E2 and hyaluronan on mesangial cell proliferation. A potential contribution to glomerular hypercellularity in diabetes.

Proliferation of mesangial cells is a feature of several forms of human and experimental glomerulopathy, including that seen in diabetes. The nonsulfated glycosaminoglycan hyaluronan participates in the regulation of pericellular matrix assembly and is a mitogen in some cell types. We have shown previously that hyaluronan production is increased in the glomerulus in a glucose- and prostaglandin-dependent manner. We have investigated the effect of diabetes and of addition of hyaluronan and prostaglandin E2 (PGE2) on the uptake of [3H]thymidine by glomerular core preparations enriched in mesangial cells. When compared with nondiabetic controls, it was shown that [3H]thymidine uptake was significantly increased in glomerular core preparations from streptozotocin-induced diabetic rats (to 169 +/- 5%, P < 0.001). In glomerular cores from both experimental groups, hyaluronan (50-250 ng/ml) or PGE2 (10(-12) to 10(-8) mol/l) increased the uptake of [3H]thymidine. Further, mesangial cells from nondiabetic control glomerular cores, when maintained in culture in early passage, responded with increased [3H]thymidine uptake to raised glucose (5.6-25 mmol/l) and to added hyaluronan and PGE2. We propose that prostaglandin and hyaluronan production in response to a raised glucose environment in diabetes can contribute to mesangial hypercellularity.

Absorption of hyaluronan applied to the surface of intact skin.

Hyaluronan has recently been introduced as a vehicle for topical application of drugs to the skin. We sought to determine whether hyaluronan acts solely as a hydrophilic reservoir on the surface of intact skin or might partly penetrate it. Drug-free hyaluronan gels were applied to the intact skin of hairless mice and human forearm in situ, with and without [3H] hyaluronan. [3H]hyaluronan was shown by autoradiography to disseminate through all layers of intact skin in mouse and human, reaching the dermis within 30 min of application in mice. Cellular uptake of [3H]hyaluronan was observed in the deeper layers of epidermis, dermis, and in lymphatic endothelium. Absorption through skin was confirmed in mice by chromatographic analysis of blood, urine, and extracts from skin and liver, which identified 3H as intact hyaluronan and its metabolites, free acetate and water. Hyaluronan absorption was similarly demonstrated without polyethylene glycol, which is usually included in the topical formulation. [3H]hyaluronan absorption was not restricted to its smaller polymers as demonstrated by the recovery of polymers of (360-400 kDa) from both blood and skin. This finding suggests that its passage through epidermis does not rely on passive diffusion but may be facilitated by active transport. This study establishes that hyaluronan is absorbed from the surface of the skin and passes rapidly through epidermis, which may allow associated drugs to be carried in relatively high concentration at least as far as the deeper layers of the dermis.

Immunolocalisation of 14-3-3 isoforms in normal and scrapie-infected murine brain.

The appearance of 14-3-3 proteins in the cerebrospinal fluid is characteristic of some neurodegenerative conditions which include sporadic Creutzfeldt-Jakob disease. Although 14-3-3 proteins are physiochemically well characterised and are known to be present in neuronal cells little is known of the neuroanatomical localisation of the individual isoforms. Using 14-3-3 isoform specific antibodies we have examined the distribution of the isoforms in normal murine brain and the changes observed during neurodegeneration as a result of ME7 scrapie infection. In normal brain there are two major patterns of immunolabelling. The beta, gamma, eta and zeta isoforms which exhibit a similar distribution pattern showing labelling of neuronal cell bodies often in particular anatomical nuclei. However the individual isoforms exhibit variation revealing subtle differences in location. The tau isoform was found only in the hippocampus and medulla, and the epsilon isoform was found throughout grey matter of the CNS. In the scrapie-infected murine brain, where severe pathological changes occur during the course of the disease, significant differences in the 14-3-3 isoform distribution were observed in the hippocampus and in the thalamus. Importantly, both the 14-3-3 eta isoform and prion protein were seen in the same neurones in both the cerebellar roof nuclei and in the lateral hypothalamic nuclei. Our study of 14-3-3 isoform distribution in adult murine brain clearly demonstrates a heterogeneous pattern of neurolocation for specific 14-3-3 isoforms. The fact that isoform labelling in terminal scrapie CNS is lost in some brain areas, but increases in others, suggests that the processing of these proteins during neurodegeneration may be much more complex than previously recognised.

The kinetics of hyaluronan in normal and acutely inflamed synovial joints: observations with experimental arthritis in sheep.

The metabolic half-life of hyaluronan (HA) in synovial fluid was estimated in sheep from the rate of appearance of 3H2O in plasma after injection of highly polymerized labeled HA. This material is substituted with 3H in its acetyl group and is rapidly and almost completely degraded in sheep and other species to yield 3H2O. Previously sensitized sheep were studied before and after induction of acute monoarticular arthritis by intraarticular challenge with type II collagen. In both circumstances 3H was released from the joint in a monophasic exponential pattern and appeared in plasma only as 3H2O. Before challenge, the mean metabolic half-life of [3H]HA was 20.8 hours (range, 15.8 to 27.9 hours, n = 5); an estimate in a single unsensitized sheep (27.0 hours) fell within this range. After challenge, swelling occurred around the joint without frankly increased synovial fluid. The mean half-life fell to 11.5 hours (range, 9.0 to 16.8 hours), with a corresponding increase in mean fractional turnover from 3.5%/h to 6.3%/h; an increased amount of the label was also retained within the peripheral tissues. It is concluded that a relatively mild acute inflammation can induce major changes in the metabolic turnover of synovial HA without the development of gross effusions. In the course of this study, mean synovial fluid volume in the normal sheep hock joint was estimated to be 1.54 mL; the concentration and content of HA were 0.54 mg/mL and 0.84 mg, respectively. These data add to other evidence that the volume and HA content of normal synovial fluid vary widely in different joints and species.(ABSTRACT TRUNCATED AT 250 WORDS)

The exanthem of Ross River virus infection: histology, location of virus antigen and nature of inflammatory infiltrate.

The exanthem of epidemic polyarthritis, a disease caused by Ross River (RR) virus, was examined three days after onset of the common erythematous and the rare purpuric forms of the eruption. The dermis showed a light perivascular infiltrate of mononuclear cells in both, with extravasation of erythrocytes in the latter. No immunoglobulins (IgM, IgG, IgA) or complement components (Clq, C3) were detected. Most of the infiltrating cells were T lymphocytes of the T suppressor-cytotoxic class. Their perivascular location, the scarcity of other lymphocytes or phagocytes, and rapid resolution of the rash indicated that the T lymphocytes were responsible for cytotoxic destruction of virus-infected cells. A few monocyte-macrophage cells were identified in the perivascular infiltrate. RR virus antigen was found in the basal epidermal and eccrine duct epithelial cells of both types of lesion and in the perivascular zone of the erythematous lesion, but appeared to have been eliminated from this region in the purpuric lesion. It is suggested that secondary effects of the T-cytotoxic reaction on nearby capillaries are responsible for erythema, oedema and purpura in the exanthem.

Activation of Fas and caspase 3 precedes PrP accumulation in 87V scrapie.

The sequence of events involved in the neurodegeneration caused by transmissible spongiform encephalopathies is not yet known. Using a murine scrapie model in which neurodegeneration in the hippocampus is restricted to the CA2, we show an up-regulation of the proapoptotic markers Fas and caspase 3 early in the incubation period prior to disease-specific prion protein (PrP) deposition and clinical signs. These results suggest that activation of Fas and caspase 3 are involved in the early pathological sequence of events during murine scrapie, and that these proapoptotic markers may be a specific method for early detection of neurodegeneration.

Early loss of dendritic spines in murine scrapie revealed by confocal analysis.

Confocal analysis of dye-filled neurons has revealed a significant early loss of dendritic spines in a murine scrapie model in which neuron loss occurs in the hippocampus. An 18% loss of spines was found at 109 days, > 50 days before neuron loss occurs, and by 126 days a 51% spine loss was found. Spine loss is concurrent with synapse loss, axon terminal degeneration and a decrease in long term potentiation in this model. Preceding these changes is the deposition of disease specific PrP at 70 days, which may initiate the damage to dendritic spines and the subsequent degeneration of synapses. We suggest that these changes underlie the development of clinical disease in the transmissible spongiform encephalopathies.

Scrapie-induced neuron loss is reduced by treatment with basic fibroblast growth factor.

Neuron loss can be a prominent feature of the pathology of transmissible spongiform encephalopathies (TSEs); recent evidence indicates that this loss occurs through apoptosis. Growth factor treatment of other neurodegenerative diseases has been shown to protect neurons destined for apoptosis, and several types of experimental retinopathy have been successfully treated with basic fibroblast growth factor (bFGF). In a murine scrapie model which develops a severe loss of photoreceptors, we administered a single intravitreal injection of bFGF four-fifths of the way through the disease process; this doubled the number of photoreceptors surviving for up to 5 weeks, i.e. to the terminal stages of the disease. This is the first time that a potential late-stage therapy for the TSEs has been demonstrated.

Apoptosis and dendritic dysfunction precede prion protein accumulation in 87V scrapie.

The sequence of events involved in the neurodegeneration caused by transmissible spongiform encephalopathies (TSEs) is not yet known. Using a murine scrapie model in which neurodegeneration in the hippocampus is restricted to CA2, we show that pyramidal neuron damage and death by an apoptotic mechanism occur early in the incubation period, prior to the appearance of CA2 disease-specific accumulation of PrP and the onset of clinical disease. We suggest that the initial hippocampal pathological event in this model is dendritic dysfunction and activation of an apoptotic pathway rather than PrP accumulation.

Fetal cell grafts provide long-term protection against scrapie induced neuronal loss.

We have transplanted fetal neurons to prolong hippocampal pyramidal cell survival in a mouse scrapie model in which 50% of CA1 pyramidal cells have died by day 180 of the 250-day incubation period. Cells prepared from embryonic PrP deficient mice were intracerebrally injected into infected mice on day 150 and groups killed on day 171 and with terminal disease. Neuron counts and CA1 depth measurements were made on semi-serial sections using an image analysis system. Both grafted groups retained more CA1 neurons than controls injected with medium alone, and showed greater depth of CA1 than controls. This new approach may have potential as a late-stage therapy for TSEs for which there are currently no available treatments.

Rubella arthritis in adults. Isolation of virus, cytology and other aspects of the synovial reaction.

Rubella virus was isolated from three consecutive samples of synovial fluid after 1 to 4 days of arthritis in adults with typical naturally acquired rubella. In the two samples suitably prepared, cells in the synovial fluid were mononuclear, with numerous monocytes and macrophages. Detailed studies in one case revealed a close similarity in all respects to the illness caused by Ross River virus, another member of the Togaviridae. Cells in the synovial exudate of this patient showed intense phagocytosis. A fine nodular cytoplasmic distribution of virus antigen was detected by immunofluorescence in 45% of the cells. There was no depletion of the complement components C'3 and C'4 in serum or synovial fluid, and no evidence of excessive immune complexes in the circulation or of immune-complex uptake by cells in the synovial exudate or intima. Synovial intima from the suprapatellar region merely showed slight hyperplasia without cellular infiltration, indicating that the lodgement of virus and the inflammatory cellular reaction to it can be confined to limited regions of the synovial lining, at least in the early stages.

Detection of Ross River virus immunoglobulin M antibodies by enzyme-linked immunosorbent assay using antibody class capture and comparison with other methods.

An enzyme-linked immunosorbent assay based on antibody class capture was developed for the detection of Ross River virus-specific immunoglobulin M antibodies (RRV IgM). The assay was specific, reproducible and precise. When compared with conventional tests for the detection of RRV IgM, such as hemagglutination inhibition following sucrose density gradient centrifugation and indirect enzyme-linked immunosorbent assay, the class capture assay was more sensitive. In 186 sera which were collected from 39 patients with RRV infection over a period of 1-4 yr from onset of initial symptoms, RRV IgM persisted for at least 1-2 yr. Sera were tested both at a single dilution from which the results were expressed as a binding index and in a dilution series in which they were expressed as an antibody titre. Binding index values gave better discrimination between sera collected during acute and later phases of the disease and may be of greater value than antibody titres in the diagnosis of RRV infection.

The potential of phase and amplitude images in determining the boundary of the left ventricle.

The phase and amplitude images derived from multiple gated blood pool studies of the heart have been used to aid the delineation of the boundary of the left ventricle. The results of left ventricular ejection fraction (LVEF) using this method have been compared with those obtained using edge detection algorithms, dual region of interest techniques, and x-ray contrast ventriculography, in a series of patients with well documented left ventricular wall motion. The results indicate that the sole use of phase and amplitude images does not improve the correlation or accuracy of LVEF estimation in relation to those obtained by x-ray contrast ventriculography, and that the dual region of interest technique is likely to remain the preferred method.

Glomerulonephritis in the acute phase of Ross River virus disease (epidemic polyarthritis).

Hematuria and proteinuria were detected at the peak of symptoms in a case of Ross River virus (RRV) disease. No other infective cause was identified. A renal biopsy 28 days after the onset of nephritis showed mild mesangial proliferative changes and one segmental sclerotic lesion. Immunofluorescence showed widespread linear deposition of IgG in glomerular capillary walls with similar but weak staining for IgM, complement (C3) and fibrinogen; granular deposits of IgM and C3 in several arterioles; and IgM in a few mesangial cells. No electron-dense deposits were detected, nor was RRV antigen found in the renal tissue. Anti-glomerular basement membrane antibodies were not detected in the serum. Recovery from the renal disturbance was complete within three months although rheumatic symptoms persisted for 30 months.

Concentration and turnover of intraperitoneal hyaluronan during inflammation.

Aseptic peritonitis was induced in rabbits by intraperitoneal injection of irritating agents, mainly starch suspensions. The inflammatory response was followed in the peritoneal lavage fluid by cell counts (average increase about 800-fold the first day) and hyaluronan concentration (average increase about 200-fold on the second and third days). The turnover rate of hyaluronan was studied by injecting tritium-labeled hyaluronan intraperitoneally and by following the appearance of tritiated water in serum. In control animals given trace amounts of hyaluronan, half-lives of 1-14 h were recorded. When the labeled polysaccharide had been mixed with 10 mg/ml of unlabeled hyaluronan, the half-life was approximately one day. Rabbits with ongoing peritonitis exhibited half-lives between 1 and 16 h. It was concluded that there was a large individual variation in uptake kinetics, that the removal process could be receptor mediated, and that the increase in intraperitoneal hyaluronan in peritonitis mainly was due to an increased production of the polysaccharide rather than a decreased rate of removal.

Onset of accumulation of PrPres in murine ME7 scrapie in relation to pathological and PrP immunohistochemical changes.

In a murine scrapie model, three different methods (immunohistochemistry, Western blotting and histoblotting) for determining disease-specific PrP accumulation were compared. The incubation period of ME7 scrapie in the F1 cross of C57 BL and VM/Dk mice is about 230 days. Mice show hippocampal neuronal loss from 160-180 days post-inoculation (dpi), CA1 neuron dendritic spine atrophy at 126 dpi, and axon terminal degeneration and synaptic loss from 84-98 dpi. Infectivity titres of at least 100 are present from 40 dpi. PrP was detected immunohistochemically at 60 dpi in the hippocampus and in the thalamus. Thus, PrP accumulation in the hippocampus precedes even the earliest neurodegenerative changes. Low amounts of PrP immunolabelling were found between 60 dpi and 126 dpi, after which the intensity increased markedly. The histoblot method detected PrPres in one of four mice at 100 dpi. Western blotting of whole brains first identified the PrPres at 80 dpi. Thus, in our hands, the most sensitive method for detecting disease-specific accumulations of PrP was immunohistochemical examination. However, immunohistochemical methods are unable to distinguish the normal and abnormal isoforms of PrP. It is therefore possible that the initial accumulation of PrP takes place as PrPsen and that the translation of PrPsen to PrPres does not take place until the later stages of the disease process. The accumulation of disease-specific PrP lags behind the development of infectivity titres. The relative rates of increase of infectivity titre and PrP accumulation are different, suggesting that these parameters may be measures of different biological events.

Determination of the frequency and distribution of vascular and parenchymal amyloid with polyclonal and N-terminal-specific PrP antibodies in scrapie-affected sheep and mice.

Brains from 17 histopathologically confirmed cases of scrapie, five of which had congophilic vascular amyloid, were stained immunohistochemically for prion protein (PrP) using a polyclonal antibody. Two clinically suspect but pathologically unconfirmed cases of natural sheep scrapie and the brains of four mice infected with the 111A murine scrapie strain were also examined. Selected sections containing amyloid were stained with each of two peptide antibodies which recognise the N-terminal amino acid residues which are lost following protease digestion of the disease-specific isoform of PrP. The mice infected with the 111A murine scrapie strain had large numbers of hypermature plaques. All the amyloid plaques from both natural sheep scrapie brains and experimental murine brains were heavily immunostained by the polyclonal and both peptide antibodies. In addition, disease-specific accumulations of PrP were detected in endothelial cells or in the intima of blood vessels of the cerebral cortex of sheep scrapie brains. The affected blood vessels were located in areas which otherwise lacked typical scrapie pathology. Vascular accumulations of PrP were also found in leptomeningeal and choroid plexus blood vessels. Vascular amyloid was found mainly in the neocortex. Vascular amyloid and disease-specific parenchymal accumulations of PrP were found in two sheep which showed clinical signs of scrapie but lacked its typical vacuolar pathology. These results show that the mature amyloid of scrapie is composed of, or contains a substantial proportion of, whole length PrP protein. Thus truncation of PrP is not essential for the aggregation of PrP into amyloid. The vascular amyloid of natural sheep scrapie originates from the accumulation and release of PrP from endothelial cells presumably following systemic scrapie infection. The topography of vascular amyloid distribution in Great Britain differs from that reported in the Netherlands. As amyloid deposition in mice is largely controlled by the strain of the infecting agent it is possible that the strain of the agent may influence vascular amyloid deposition.

Tobacco use and smoking policy perceptions onboard an aircraft carrier.

Prior to implementing a shipwide no-smoking policy, the crew of U.S.S. Theodore Roosevelt (CVN 71) participated in a voluntary survey on tobacco-related matters. The survey queried participants on their tobacco-use history, subjective exposure to environmental tobacco smoke (ETS), and attitudes related to smoking policy prior to the cessation of all smoking activities aboard ship. Of the 2,221 crewmembers who participated (74% response rate), 36% classified themselves as current cigarette smokers. Nonsmokers estimated their general exposure to ETS between "low" to "moderate." Of all participants, 57% were in favor of the current restricted smoking policy, including 18% of currently smoking personnel. Follow-up research is being conducted to assess the long-term impact of the no-smoking policy on changes in attitudes regarding policy, tobacco-use rates, and ETS exposure.