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Biomedical subjects

J R Fraser

Publications and source records attributed to J R Fraser.

At least 91 records · Page 5Linked to original sources

Epidemic polyarthritis and Ross River virus disease.

Ross River virus is a mosquito-transmitted alphavirus indigenous to Australia, Papua New Guinea and nearby islands, which recently appeared in other western and central South Pacific islands. Human infection can be manifest by varied constitutional disturbances, rash and rheumatic symptoms, known in Australia as epidemic polyarthritis and broadly similar to certain alphavirus diseases in other regions. Although usually short-lived, the rash can persist for 5 months. Rheumatic effects involve synovial joints, tendon and ligaments, and can continue or recur in peripheral joints and tissues as long as 6 years, though gradually improving without destructive changes. At different times, the disease can closely simulate rubella and other virus diseases, Henoch-Schönlein syndrome, rheumatoid and other chronic rheumatic diseases. Diagnosis rests upon geography, specific serology and judicious interpretation of clinical and supportive laboratory data. Skin and synovial lesions are characterized by infiltration of mononuclear cells. Their pathogenesis most likely depends on the reaction of these cells with persistent foci of virus disseminated during the early viraemic phase of infection.

Arthritis, Infectious↗

Removal of hyaluronic acid from the circulation in rheumatoid disease and primary biliary cirrhosis.

Recent evidence from metabolic studies in animals and from measurement of plasma and serum hyaluronic acid (HA) levels in humans indicates that the liver is the organ mainly responsible for the clearance of circulating HA. The clearance of HA from the bloodstream was therefore studied with tritium-labeled material of high specific activity in patients with rheumatoid arthritis or primary biliary cirrhosis (PBC). In patients with rheumatoid arthritis, HA clearance was similar to that reported in normal subjects and approached the range of hepatic blood flow. HA metabolism was also rapid. In patients with PBC, clearance was inversely related to the preexisting plasma HA levels and severity of disease. Renal excretion of HA remained low despite the slower decay of plasma concentrations. Metabolic degradation was not significantly impaired. Most of the injected material was accounted for as tritiated water within 24 hours. Taken with data from other species and with the raised plasma HA levels observed in a variety of liver diseases, our findings indicate that the liver may also be the major site for the uptake of circulating HA in humans. High levels of plasma HA in the absence of hepatic dysfunction can be taken as evidence of an increased input from the tissues.

Adult↗

Uptake of circulating hyaluronic acid by the rat liver. Cellular localization in situ.

The uptake of [3H]acetyl-labelled hyaluronic acid (HA) was examined in the liver, spleen and kidney of the rat after i.v. injection. 3H-activity was located by light- and electron-microscopic autoradiography after measurement by scintillation counting of tissue digests. In the liver, approximately 90% of the radioactivity was located in the sinusoidal endothelial cells, with autoradiographic grains distributed throughout the cytoplasm; 50% of the grains overlay vacuoles 0.3 to 1.2 micron in diameter. A few grains (4%) were located in Disse's space or nearby in the cytoplasm of hepatocytes. No grains were found in Kupffer cells. The remainder were randomly scattered across the sections in a pattern indicating nonspecific background activity. These observations are in accordance with the selective uptake of HA exhibited by dissociated liver cells in vitro. HA concentrations in the spleen and kidney were too low for detection by autoradiography. Splenic concentrations were much lower than in rabbits or mice; in this respect the uptake of circulating HA in the rat resembles that reported for chondroitin 4-sulphate.

Animals↗

Peripheral blood mononuclear cells stimulate prostacyclin levels of human synovial fibroblast-like cells.

Fibroblast-like synovial cells, isolated from intact joints of non-arthritic human donors and from explants of rheumatoid and non-rheumatoid synovial tissue, released prostacyclin (PGI2) when incubated in conditioned medium from human peripheral blood mononuclear cells (MCCM). The effect of MCCM on the rate of PGI2 synthesis (measured by radioimmunoassay as the stable product, 6-keto-PGF1 alpha) was clearly established within 2 h and appeared to require RNA and protein synthesis as judged by inhibition with actinomycin D and cycloheximide, respectively. Low concentrations of dexamethasone suppressed the increase in PGI2 levels. Prostaglandin E2 (PGE2) levels were also raised by the MCCM and reduced by dexamethasone. All-trans retinoic acid did not stimulate the levels of either prostanoid. These findings offer an explanation for some of the inflammatory events occurring in rheumatoid lesions.

6-Ketoprostaglandin F1 alpha↗

Persistent rubella virus infection of human synovial cells cultured in vitro.

Primary and passaged human synovial cell cultures inoculated with wild-type and vaccine strains of rubella virus were incubated at 32 C and 37 C. At 32 C, the temperature close to that in human peripheral joints, infection persisted with extracellular virus titers of approximately 10(4) fluorescent focus-forming units/ml. Extracellular titers at 37 C first stabilized at one-tenth the level of the titers at 32 C and then declined after 20 days to undetectable levels. Cellular expression of viral antigen rose at 32 C and fell at 37 C. Infection was noncytocidal at all stages. Virus yields reflected the temperature in the subsequent incubation rather than during virus adsorption. Interferon was found only in cultures held at 37 C and was mainly alpha with a minor beta fraction, a result suggesting retention of functional characteristics of the type A macrophage-like synovial cell. We conclude that persistent infection of synovial tissue in vivo is a feasible explanation for the presence of rubella virus in peripheral joints of patients with chronic arthritis.

Animals↗

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.

Alphavirus↗

Ross River virus infection of human synovial cells in vitro.

Ross River virus (RRV) strains T48 and SC18006 produced a self-limited cytopathic infection of primary and passaged human synovial cell culture. Extracellular (EC) virus titres reached peak levels at 2 days in cell lines and at 4 days in primary cultures, ranging between 10(4.5) and 10(6.6) fluorescent focus-forming units (ffu)/ml. Thereafter titres declined rapidly to undetectable levels at 10-12 days. The proportion of adherent cells showing virus antigen exceeded 60% at 3 days and decreased in all cultures to less than 1/500 after 12 days. Cytopathic effects (CPE) were greatest at 4-8 days and destroyed between 25 and 75% of the cell layer, with subsequent partial regeneration by division of surviving cells. In contrast to rubella virus infection of synovial cells, cultures at 32 degrees and 37 degrees revealed only minor differences and persistent infection was not established. CPE were more extensive at 37 degrees in nearly all synovial cell cultures and in Vero cultures. At 37 degrees synovial cells infected with T48 strain produced higher maximum titres and were more extensively infected than at 32 degrees.

Cells, Cultured↗

Studies in vitro on the uptake and degradation of sodium hyaluronate in rat liver endothelial cells.

Rat liver endothelial cells in primary cultures at 7 degrees C bind radioactively labelled sodium hyaluronate (HA; Mr 400 000) specifically and with high affinity (Kd = 6 X 10(-11) M). Maximal binding capacity is approx. 10(4) molecules per cell. Inhibition experiments with unlabelled HA and oligosaccharides from HA indicate that each molecule is bound by several receptors acting co-operatively and that the single receptor recognizes a tetra- or hexa-saccharide sequence of the polysaccharide. At 37 degrees C the liver endothelial cells endocytose the HA. The process combines the features of a receptor-mediated and a fluid-phase endocytosis. The rate of internalization does not show any saturation with increasing HA concentration, but is approximately proportional to the polysaccharide concentration at and above the physiological concentration. At 50 micrograms of free HA/l each liver endothelial cell accumulates 0.1 fg of the polysaccharide/min. Fluorescent HA accumulates in perinuclear granules, presumably lysosomes. Degradation products from HA appear in the medium about 30 min after addition of the polysaccharide to the cultures. The radioactivity from HA containing N-[3H]acetyl groups or 14C in the sugar rings is recovered mainly as [3H]acetate and [14C]acetate respectively. Estimations of the capacity of liver endothelial cells to internalize and degrade HA in vitro indicate that these cells may be primarily responsible for the clearance of HA from human blood in vivo.

Animals↗

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.

Adult↗

Stimulation of glycosaminoglycan production and lysosomal activity of human synovial cells in culture by low environmental pH.

Glycosaminoglycan production, acid hydrolase activity, proliferation, and morphology were examined in human synovial cells subjected to low environmental pH. The amount and the molecular size of newly synthesised glycosaminoglycan (GAG) were increased without significant change in the rate of cell proliferation. Lowered pH produced an increase in the size of cytoplasmic organelles. Some of these possessed ultrastructural features of lysosomes, but others were clearly nonlysosomal and were of uncertain identity. Intracellular activity of the lysosomal acid hydrolase N-acetyl-beta-glucosaminidase (NAG) was not altered by low pH, but a marked increase occurred in extracellular NAG activity, indicating enhanced release.

Acetylglucosaminidase↗

Leucine metabolism as a source of acetate in the synthesis of hyaluronic acid.

Synovial fibroblasts were incubated in nutrient medium with either L-[U-14C]leucine or L-[4,5-3H]leucine to reveal peptides associated with hyaluronic acid. Isotope was found in hyaluronic acid extracted from culture medium by repeated density gradient ultracentrifugation, but 76-98% of the labelled hyaluronic acid fraction was recovered as oligosaccharide by Sephadex G-25 gel chromatography after digestion with protease-free staphylococcal hyaluronidase (EC 4.2.99.1). Radioactivity in acid digests of the oligosaccharide fractions (i) was volatile, (ii) separated similarly to acetate in Biogel P2 chromatography, and (iii) was identified as acetate by melting point after conjugation as a p-phenylphenacyl ester crystallized to constant specific activity. The synovial fibroblast thus possesses metabolic pathways for complete transformation of leucine to acetylCoA, which operate freely in adequate nutrient medium and are a significant source of acetyl groups in hyaluronic acid synthesis.

Acetates↗

Tissue uptake of circulating hyaluronic acid. A whole body autoradiographic study.

Previous work in the rabbit has shown that there is a significant flux of plasma hyaluronic acid (HA) which is taken up and degraded mainly in the liver but also concentrated in the spleen. Purified 14C-labelled HA of high average molecular wt prepared by biosynthesis from D-[U-14C] glucose was injected i.v. in mice and its tissue distribution was determined by whole-body autoradiography during the next 24 h. As blood levels declined, radioactivity was concentrated in the liver and spleen as found in the rabbit, and also in bone marrow and lymph nodes. Distribution was uniform in liver tissue, concentrated and relatively persistent in the periphery of lymph nodes, and distinctly nodular within the spleen. Analysis of an aqueous liver extract taken 4 h after injection identified 14C in HA, in a macromolecular fraction resistant to fungal hyaluronidase, and in metabolites of low molecular wt. These findings confirm and extend observations based on tissue extraction in rabbits. The pattern of distribution through the body and the restricted localization within spleen and lymph nodes further suggest that HA is absorbed from plasma and tissue fluids by elements of the reticuloendothelial system.

Animals↗

Turnover of hyaluronate in the aqueous humour and vitreous body of the rabbit.

Sodium hyaluronates with molecular weights of 18000, 500000 and 4 x 10(6), labelled with either 3H or 14C, were injected into the anterior chamber or into the centre of the vitreous body of rabbits and their rates of disappearance were followed. The out-flow from the anterior chamber in anaesthetized animals was virtually independent of the molecular weight indicating that the disappearance of the polysaccharide is controlled by bulk flow. The rate constant for the disappearance of sodium hyaluronate was 0.0094/min and after treatment with indomethacin 0.0061/min. These figures are in general agreement with published flow-rates of aqueous humour in the rabbit. The disappearance from the vitreous body was strongly molecular weight dependent indicating a diffusion controlled transport. The rate constant for hyaluronate with mean molecular weight of 18000 was 0.16/day and 500000, 0.024/day. The rate constant for endogenous hyaluronate was estimated to be about 0.01/day. A calculation using these rate constants shows that the turnover of sodium hyaluronate in the rabbit anterior chamber is about 3 micrograms per 24 hr while the turnover in the vitreous body is only 15% of that. This confirms an earlier conclusion (Laurent and Granath, 1983) that the preponderant part of the hyaluronate in aqueous humour is not a general degradation product from the vitreous body.

Animals↗