Biomedical subjects

K Watson

Publications and source records attributed to K Watson.

At least 55 records · Page 3Linked to original sources

PubMed1998

De novo protein synthesis is essential for thermotolerance acquisition in a Saccharomyces cerevisiae trehalose synthase mutant.

Heat shock (25 degrees C to 37 degrees C for 30 min) acquisition of thermotolerance (at 50 degrees C) was observed in a yeast trehalose synthase mutant and the corresponding control strain. The acquisition of thermotolerance in the control strain was maintained for a significantly longer time than in the trehalose synthase mutant. The heat shock was associated with the synthesis of specific heat shock proteins and, in the case of the control strain, also trehalose accumulation. Inhibition of protein synthesis during the heat shock totally abolished acquisition of thermotolerance in both strains but not trehalose accumulation in the control. It was concluded that trehalose may only be required for prolonged stress protection while heat shock proteins are required for heat shock acquisition of thermotolerance.

Fungal Proteins↗

PubMed1998

Heat shock response in the thermophilic enteric yeast Arxiozyma telluris.

Heat stress tolerance was examined in the thermophilic enteric yeast Arxiozyma telluris. Heat shock acquisition of thermotolerance and synthesis of heat shock proteins hsp 104, hsp 90, hsp 70, and hsp 60 were induced by a mild heat shock at temperatures from 35 to 40 degrees C for 30 min. The results demonstrate that a yeast which occupies a specialized ecological niche exhibits a typical heat shock response.

Ascomycota↗

PubMed1998

Transcriptional and translational regulation of major heat shock proteins and patterns of trehalose mobilization during hyperthermic recovery in repressed and derepressed Saccharomyces cerevisiae.

Patterns of heat shock gene transcription and translation, as well as trehalose content, were investigated in both glucose (repressed) and acetate (derepressed) grown cells of Saccharomyces cerevisiae during heat shock and subsequent return of cells to 25 degrees C. Heat-shocked cells (37 degrees C for 30 min), grown in either glucose- or acetate-supplemented media, initially acquired high thermotolerance to a 50 degrees C heat stress, which was progressively lost when cultures were allowed to recover at 25 degrees C and subsequently exposed to a second heat stress. In all cases, with the notable exception of repressed cells of a relatively thermosensitive strain, inhibition of protein synthesis and coincident decrease in trehalose accumulation during the heat shock had little effect on the kinetics of loss of thermotolerance. Heat shock at 37 degrees C elicited a marked increase in transcription and translation of genes encoding major heat shock proteins (hsps). During recovery at 25 degrees C, both metabolic activities were suppressed followed by a gradual increase in hsp mRNA transcription to levels observed prior to heat shock. De novo translation of hsp mRNAs, however, was no longer observed during the recovery phase, although immunodetection analyses demonstrated persistence of high levels of hsps 104, 90, 70, and 60 in cells throughout the 240-min recovery period. In addition, while heat shock induced trehalose was rapidly degraded during recovery in repressed cells, levels remained high in derepressed cells. Results therefore indicated that the progressive loss of induced thermotolerance exhibited by glucose- and acetate-grown cells was not closely correlated with levels of hsp or trehalose. It was concluded that both constitutive and de novo synthesized hsps require heat shock associated activation to confer thermotolerance and this modification is progressively reversed upon release from the heat-shocked state.

Acetates↗

PubMed1998

Laboratory and field evaluation of efficacy of VectoBac 12AS against Culex sitiens (Diptera: Culicidae) larvae.

Laboratory bioassay studies of the efficacy of VectoBac 12AS (active ingredient: 1,200 International Toxic Units [ITU]/mg Bacillus thuringiensis var. israelensis) against field-collected late 3rd/early 4th-instar larvae of Culex sitiens indicated excellent control potential. A 95% lethal concentration (LC95) value of 1.381 x 10(7) ITU was calculated, which equated to a dosage of 0.011 liters/ha. This dosage represented 1.8% of the recommended lowest dosage rate for the product. A field trial of VectoBac 12AS against late 3rd/early 4th-instar field specimens of Cx. sitiens in floating mesh cylinders was then conducted in salt-marsh pools near Coomera Marina, southeast Queensland, Australia. At a rate of 0.5 liters/ha, 100% mortality of Cx. sitiens larvae was recorded at 24 h posttreatment.

Animals↗

PubMed1997

Stress proteins and stress tolerance in an Antarctic, psychrophilic yeast, Candida psychrophila.

Conditions are described for the heat shock acquisition of thermotolerance, peroxide tolerance and synthesis of heat shock proteins (hsps) in the Antarctic, psychrophilic yeast Candida psychrophila. Cells grown at 15 degrees C and heat shocked at 25 degrees C (3 h) acquired tolerance to heat (35 degrees C) and hydrogen peroxide (100 mM). Novel heat shock inducible proteins at 80 and 110 kDa were observed as well as the presence of hsp 90, 70 and 60. The latter hsps were not significantly heat shock inducible. The absence of hsp 104 was intriguing and it was speculated that the 110 kDa protein may play a role in stress tolerance in psychrophilic yeasts, similar to that of hsp 104 in mesophilic species.

Candida↗

PubMed1997

A comparative electrophoretic analysis of mammalian hair and avian feather proteins.

An improved one-dimensional sodium dodecyl sulphate separation has allowed a detailed electrophoretic comparison of the proteins that constitute hairs and feathers isolated from various representative samples of mammalian, both placental and marsupial and avian species. The proteins from various species were S-carboxymethylated using labelled (14C)-iodoacetic acid and detected both by Coomassie stain and fluorography. Proteins from mammalian species were compared with respect to their molecular masses and relative cysteine content using laser densitometry of both Coomassie stained gels and fluorographs of 14C labelled gels. These results may be quite useful forensically as an adjunct to fibre analysis.

Animals↗

PubMed1997

Stress co-tolerance and trehalose content in baking strains of Saccharomyces cerevisiae.

Fourteen wild-type baking strains of Saccharomyces cerevisiae were grown in batch culture to true stationary phase (exogenous carbon source exhausted) and tested for their trehalose content and their tolerance to heat (52 degrees C for 4.5 min), ethanol (20% v/v for 30 min), H2O2 (0.3 M for 60 min), rapid freezing (-196 degrees C for 20 min, cooling rate 200 degrees C min-1), slow freezing (-20 degrees C for 24 h, cooling rate 3 degrees C min(-1)), salt (growth in 1.5 M NaCl agar) or acetic acid (growth in 0.4% w/v acetic acid agar) stresses. Stress tolerance among the strains was highly variable and up to 1000-fold differences existed between strains for some types of stress. Compared with previously published reports, all strains were tolerant to H2O2 stress. Correlation analysis of stress tolerance results demonstrated relationships between tolerance to H2O2 and tolerance to all stresses except ethanol. This may imply that oxidative processes are associated with a wide variety of cellular stresses and also indicate that the general robustness associated with industrial yeast may be a result of their oxidative stress tolerance. In addition, H2O2 tolerance might be a suitable marker for the general assessment of stress tolerance in yeast strains. Trehalose content failed to correlate with tolerance to any stress except acetic acid. This may indicate that the contribution of trehalose to tolerance to other stresses is either small or inconsistent and that trehalose may not be used as a general predictor of stress tolerance in true stationary phase yeast.

Acetic Acid↗

PubMed1997

Efficacy and tolerance of a 6-month treatment course of daily interferon-alpha 2a for chronic hepatitis C with cirrhosis. The Australian Hepatitis C Study Group.

Response to interferon-alpha (IFN-alpha) treatment in hepatitis C is poorer when cirrhosis is present. In the third Australian multicentre hepatitis C trial, Aushep-3, we examined the efficacy and tolerability of an intensive 24-week course of interferon-alpha 2a in Child-Pugh grade A patients with chronic hepatitis C and cirrhosis. This was an open uncontrolled trial of 4.5 million units (MU) of IFN-alpha 2a daily for 24 weeks; follow-up was 48 weeks. Chronic hepatitis C and cirrhosis were confirmed histologically. HCV RNA was determined in serum by reverse transcriptase polymerase chain reaction (PCR), and viral genotyping was by line-probe assay. Treatment response was defined as a reduction of alanine aminotransferase (ALT) to less than 1.5 times the upper limit of normal (and by at least 50% of pretreatment values) at weeks 20 and 24. Sustained response was defined as normal serum ALT after treatment from trial week 28 until week 48. Among the 56 patients, a treatment response occurred in 18 (32% by intention-to-treat; 42% of those who completed treatment) and eight (14%) had a sustained response. At 24 weeks, HCV RNA was not detectable in 12 of 17 treatment responders, and remained negative at 48 weeks in six of eight sustained responders. Treatment response by genotype occurred in 75% of patients with HCV type 2, in 38% with HCV type 3a and in 12% with HCV genotype 1. Sustained response occurred in only one (4%) patient with HCV genotype 1 but in five (20%) with genotypes 2 or 3a. Among 13 patients withdrawn, nine were for adverse effects, most often haematological; 10 others underwent dose reduction for adverse effects. It is concluded that a sustained biochemical and viral response to treatment with IFN-alpha 2a can be obtained in some patients with hepatitis C and cirrhosis, particularly those with genotypes 2 or 3a. Therefore, patients with cirrhosis should be considered for interferon treatment on an individual basis. Genotyping may improve case selection, but vigilance is required for haematological complications.

Adult↗

PubMed1997

Treatment of cirrhotic hepatitis C virus patients with daily doses of interferon-alpha 2a.

In patients with hepatitis C who have cirrhosis the rate of sustained response following interferon therapy is less than half that of patients without cirrhosis. It has been suggested, however, that a higher dose regime in patients with cirrhosis may improve response. The results of a recent Australian study of cirrhotic patients who were given an intense interferon programme of 4.5 MIU daily for 24 weeks were compared with previous studies of patients with hepatitis C. In the Australian study, 14% of patients had a sustained response at 6 months after end of therapy. Of 11 studies of interferon response in chronic hepatitis C comparison of pretreatment variables showed considerable differences. Identification of predictors of response by univariate and multivariate analysis regularly indicated the importance of age and fibrosis. Analysis of six studies with either a poor (5% or less) or a reasonable (14-19%) sustained response rate to interferon in patients with cirrhosis suggested that a higher dose or longer duration of therapy was associated with better results. The experience of the Australian study, where 14% of patients had a sustained biochemical response to interferon and side-effects were reasonably tolerated with careful monitoring, suggests that future studies in cirrhosis should be carried out exploring higher doses and longer durations of therapy.

Australia↗

PubMed1997

Membrane fatty acid composition and membrane fluidity as parameters of stress tolerance in yeast.

The relationship among membrane fatty acid composition, membrane fluidity, and stress tolerance was investigated in yeast cells. Several strains were examined for their ability to survive heat, ethanol, and hydrogen peroxide stresses. Membrane fluidity was determined by measuring fluorescence anisotropy using diphenylhexatriene as a probe. There was no obvious relationship among membrane fatty acyl composition, membrane fluidity, and stress tolerance in the strains examined. A consistent trend in the present study was an observed decrease in membrane fluidity following thermal treatment, which coincided with a reduction in cell viability. We suggest that protein denaturation may be responsible for the observed effect of elevated temperature on membrane fluidity and viability. This was implied by observations on the irreversible nature of thermal transitions, as measured by breaks in Arrhenius plots, in which stationary phase cells were shown to exhibit higher transition temperatures (53.9-55.5 degrees C) than exponential phase cells (49.5-51 degrees C). Furthermore, the thermal transition temperature was shown to increase in exponential phase cells following heat shock, which was associated with an increase in thermotolerance. We suggest that the thermotolerant state of heat-shocked cells and cells entering stationary phase may be associated with increased protein stability. However, despite the relatively good correlation between thermal transition temperature and stress tolerance, the thermal transition temperature did not predict the stress tolerance of a given strain, as stress-sensitive strains had similar transition temperatures to those of stress-resistant strains.

Cell Membrane↗

PubMed1997

Prevention of hepatitis A and B.

The vaccine preventable hepatitides, hepatitis A and hepatitis B, cause a great burden of illness in both developing and industrialized nations. There is increased community awareness of the problems of viral hepatitis, and it is important for general practitioners to be aware of current recommendations for prevention of hepatitis A and B.

Adolescent↗

PubMed1997

Progress in local anaesthesia.

Research into improving local anaesthetic agents and methods of delivering them in dental practice has been very slow. This article describes some of the progress that is being made in these areas.

Anesthesia, Dental↗

PubMed1996

Heat shock protein synthesis and trehalose accumulation are not required for induced thermotolerance in depressed Saccharomyces cerevisiae.

Intrinsic and heat shock induced thermotolerance of Saccharomyces cerevisiae was investigated in cells grown on glucose and acetate supplemented media. Heat shocked cells (37 degrees C/30 min), in either medium, exhibited induced synthesis of heat shock proteins (hsp) and trehalose. In all cases, with the notable exception of repressed cells of a relatively thermosensitive strain, heat shock acquisition of thermotolerance also occurred in the absence of protein synthesis and coincident decrease in trehalose accumulation. Results indicted that the marked increase in thermotolerance exhibited by non-fermenting (acetate) cells compared with fermenting (glucose) cells was not closely correlated with levels of hsp or trehalose. It was concluded that mechanisms for intrinsic and induced thermotolerance appear to be different and that growth on acetate endows cells with a biochemical predisposition, other than hsp or trehalose, that confers intrinsic tolerance, a factor which may be subject to heat induced modification.

Acclimatization↗

PubMed1996

Peptide mapping of S-carboxymethylated hair and feather proteins using two-dimensional electrophoresis.

The aim of this work was to develop a two-dimensional electrophoretic method for the multiple simultaneous peptide mapping of a broad spectrum of human and animal hair and avian feather proteins. The 14C labelled S-carboxymethylated proteins of hair and feather were separated in one dimension by sodium dodecyl sulphate-polyacrylamide slab gel electrophoresis and stained with Coomassie Blue. Each of the gel lanes containing the separated hair or feather proteins from one individual was cut and transferred at right angles on to a second slab gel. A solution of trypsin (5 micrograms/ml) in stacking gel buffer was poured on to the gel lane. Partial proteolysis of the hair or feather proteins proceeded in situ while the stacking gel buffer set. The second dimension of electrophoresis followed by staining and/or fluorography showed a characteristic pattern of peptides of proteins in the form of spots derived from each individual protein, leaving undigested proteins well separated on the diagonal. The protease digestion pattern was reproducible and characteristic of each sample. We were able to establish a finer peptide signature, for individual samples containing a mixture of many proteins, than was hitherto possible with previously published two-dimensional electrophoretic techniques. It was concluded that this technique may be of use in future evolutionary, ontogenetic and forensic studies using hair and feathers as a biological source material.

Animals↗

PubMed1996

Role of molecular regulation in vascular calcification.

Calcium deposits account for most of the dry weight of atherosclerotic lesions. Previously considered uncommon, vascular calcification is now known to be present in 80% of significant lesions and in at least 90% of patients with coronary artery disease. Previously considered a passive process, it is increasingly recognized as an active, regulated process. Previously considered benign, it is now becoming recognized as a major risk factor for cardiovascular events, and a major contributor to systolic hypertension, heart failure, plaque rupture and stenosis. To confirm the similarity of vascular calcification with embryonic osteogenesis, we demonstrated the expression of bone morphogenetic protein in calcified human lesions, and we developed an in vitro model of vascular calcification that provides a useful experimental system for elucidating the molecular regulation of this process, which we have shown to include alkaline phosphatase induction and expression of bone matrix proteins and differentiation factors. Understanding the regulatory mechanisms of vascular calcification will allow future therapeutic approaches to prevent and possibly reverse this disease and its clinical consequences.

Animals↗

PubMed1996

Acute toxicity of selected pesticides to the estuarine shrimp Leander tenuicornis (Decapoda:Palaemonidae).

The shrimp Leander tenuicornis is abundant in southeastern Queensland intertidal marsh pools and was chosen as an indicator species for toxicological studies with pesticides. Acute toxicity to this crustacean of temephos and 3 pesticide compounds under evaluation for registration in Australia (Bacillus thuringiensis var. israelensis, s-methoprene, and pyriproxyfen) was tested in 96-h laboratory trials. Temephos was the most toxic compound, with a median lethal concentration (LC50) of 0.01 ppm (0.33 times the estimated field concentration [EFC] for a 15-cm-deep pool). s-Methoprene was the least toxic compound, with an LC50 of 14.32 ppm (1,790 times the EFC). Bacillus thuringiensis var. israelensis and pyriproxyfen produced LC50 values of 60.9 x 10(6) ITU (176 times the EFC) and 0.098 ppm (12.25 times the EFC), respectively.

Animals↗

PubMed1995

Extra-nuclear location of histones in activated human peripheral blood lymphocytes and cultured T-cells.

Dextrin-2-sulphate (D2S) is a sulphated polysaccharide which inhibits human immunodeficiency virus type 1 infection of T-cells by binding to the cell surface. During our investigations of the nature of this interaction, a cell membrane fraction was prepared by ultracentrifugation from the T-cell line, HPB-ALL. Separation of membrane proteins by sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis and analysis for binding proteins using ligand blotting showed that 3H-D2S bound, in a saturable and displaceable manner, to two regions corresponding to molecular weights of 14,000-18,000 and 28,000-32,000. The N-terminal sequences of two of the major protein components in the 14,000-18,000 region were consistent with those of histones H2B and H3. The presence of histone H2B in the cell membrane preparation was confirmed by immunoblotting and enzyme-linked immunosorbent assay using a specific antibody. Histone standards were used to determine the level of each histone in the cell membrane fraction. In addition, the binding of 3H-D2S to purified histone standards was quantified. These results show that all of the binding of 3H-D2S to proteins in the 14,000-18,000 region of the cell membrane preparation can be attributed to the histones present. In contrast to HPB-ALL cells, a cell membrane fraction from freshly isolated human peripheral blood lymphocytes contained very low levels of histones. However, after culture with phytohaemagglutinin for 3 days the cell membrane fraction contained greatly increased levels of histones. To exclude the possibility of contamination of the cell membrane preparation with histones derived from the nucleus, cell membranes were also prepared using an affinity-based method using polyethyleneimine-cellulose. Immunoblotting of adsorbed plasma membranes showed the presence of histone H2B. SDS-polyacrylamide gels stained for protein also indicated that the preparation contained histones H1, H2A, H3 and H4. In further experiments whole cells were used to avoid contamination from nuclear proteins. Lactoperoxidase mediated 125I labelling, a method specific for radiolabelling cell surface proteins, confirmed the presence of histones H2B, H3 and H4 on the surface of HPB-ALL cells. Also, incubation of HPB-ALL cells or phytohaemagglutinin-activated peripheral blood lymphocytes with D2S caused displacement of histones from the cell surface into the supernatant without altering cell viability. In addition, immunocytochemistry of freshly isolated peripheral blood lymphocytes showed that histone H2B was located predominantly in the nucleus. However, in phytohaemagglutinin-activated peripheral blood lymphocytes immunoreactive material was also prominent in the endoplasmic reticulum and on the plasma membrane.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗

PubMed1995

High resolution one-dimensional electrophoretic separation and partial characterisation of human head hair proteins.

A reproducible, rapid procedure for the extraction, labelling and separation of human hair proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) has been developed. Hair proteins were extracted in 8 M urea, containing 0.2 M mercaptoethanol, followed by sonication. Extracts were neutralised with Tris and incubated with either labelled (14C) or unlabelled iodoacetic acid to S-carboxymethylate cysteine groups. Proteins were separated on 12.5% SDS-polyacrylamide gels and gels stained with Coomassie Brilliant Blue and/or silver nitrate to reveal major protein bands. Gels were then treated with a fluorographic agent, dried and autoradiographed to reveal major sites of S-carboxymethylation. A given gel was scanned by laser densitometry after Coomassie and/or silver stain to quantitate the protein content of each major protein zone. An autoradiogram of the same gel was scanned to estimate the cysteine content of each major zone. In this way it was possible to partially characterise rapidly and reproducibly many different protein zones in different individual samples on one gel at the same time. By calculating the ratio of autoradiograph absorbance to Coomassie Blue absorbance, protein zones could be assigned to four different categories, viz: very high cysteine (VHC) proteins, high cysteine (HC) proteins, low cysteine (LC) proteins and very low cysteine (VLC) proteins. The method described is reproducible, rapid and inexpensive enough to be suitable for mass screening. Overall the results were more informative than previously reported one-dimensional separations and indeed this technique may well be more suited to forensic and/or population investigations than the much more laborious and time-consuming two-dimensional techniques.

Autoradiography↗