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M J Howell

Publications and source records attributed to M J Howell.

At least 37 records · Page 2Linked to original sources
PubMed1985

In vitro translation of mRNA from Toxocara canis larvae.

300 micrograms of total RNA was extracted from 1 ml of packed Toxocara canis larvae by centrifugation through a 5.7 M cesium chloride cushion. 60 micrograms of polyadenylated messenger RNA was separated from 300 micrograms of total RNA in an oligothymidylic acid-cellulose gel column. The in vitro translation of the mRNA, isolated from T. canis larvae, was carried out using the rabbit reticulocyte cell-free translation system. Incorporation of [35S]methionine into trichloroacetic acid precipitable material in the lysate containing mRNA was 4-5 times greater than that of control. Translation products were analysed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) followed by autoradiography. Many polypeptides ranging in molecular weight from 10000 to 100000 were synthesised in the lysate. A T. canis positive human serum was mixed with translation products to form antigen-antibody complexes, which were then absorbed by Staphylococcus aureus Cowan 1 strain and analysed by the autoradiography of SDS-PAGE. Three antigenic polypeptides with molecular weights of 49000, 27000 and 22000 which reacted specifically with IgG antibody in T. canis positive human serum, were demonstrated. The 27000 MW polypeptide reacted particularly strongly with the IgG antibody.

Animals↗
PubMed1985

Biosynthetic labelling of the excretory and secretory antigens of Toxocara canis larvae.

Toxocara canis larvae were cultured in vitro in medium containing [35S-]methionine for six days. The medium and the larval tissues were analysed for biosynthetically labelled polypeptides by sodium dodecyl sulphate polyacrylamide gel electrophoresis and autoradiography. Immunoprecipitates with positive and negative human antiserum were similarly analysed, using Staphylococcus aureus to absorb immunocomplexes. The larvae secrete biosynthetically labelled polypeptides into the medium, with three major polypeptides of molecular weights between 99 and 110 X 10(3) the major constituents. Both of these react strongly with human IgG in human positive sera. Many polypeptides become labelled in the larval tissue, but only one polypeptide with similar molecular weight to the ES antigens, strongly reacted with human IgG.

Animals↗
PubMed1985

Contrast agents and enzyme inhibition. II. Mechanisms.

The inhibition of enzymes by radiological contrast agents has been invoked to explain some of their clinical effects. In a previous paper, in-vitro studies of the inhibition of the enzyme acetylcholinesterase were presented and some clinical correlations were discussed. More detailed studies, described here, have now shed new light on the mechanisms involved in this important phenomenon. Because the findings restore the iodine atoms to a central role, they have important general implications for contrast agent pharmacology.

Cholinesterase Inhibitors↗
PubMed1984

Killing of Taenia hydatigena oncospheres by sheep neutrophils.

Neutrophils collected from the mammary glands of uninfected sheep or from sheep infected with Taenia hydatigena, attached to and killed T. hydatigena oncospheres in vitro in the presence of serum from infected sheep. Infected sheep serum alone was not deleterious to the parasite in vitro. Fc receptors for antibody were detected on both normal and immune neutrophils; they were present at a greater density on the latter. Immune neutrophils were more reactive towards oncospheres than normal neutrophils and formed extensive capsules around the parasite. Fc receptors were not detected on oncospheres. It is hypothesised that neutrophils may kill the parasite by producing hydrogen peroxide and the superoxide anion, both of which are toxic to a variety of cell types and protozoa. The function of antibody may be to facilitate attachment of neutrophils to oncospheres by way of their Fc receptors.

Animals↗
PubMed1984

Contrast-medium-induced acute renal failure and Tamm-Horsfall proteinuria.

An acute obstructive nephropathy due to precipitation of urinary glycoprotein-contrast complexes in the renal tubules has been postulated to explain the episodes of renal failure occasionally seen following intravascular contrast medium administration. In an in vitro study we were unable to produce any precipitation of contrast-glycoprotein complexes over a wide range of concentrations, temperatures and pH values in urine with any urographic contrast agent, conventional or new. Meglumine ioglycamide (Biligram: Schering) alone, the strongly protein-binding agent, exhibited the phenomenon, and only in high concentration. It is concluded that such a mechanism is unlikely to play a role in contrast medium nephrotoxicity, which therefore remains unexplained.

Acute Kidney Injury↗
PubMed1984

Cyclohexanone extraction: an improvement in the thiobarbituric acid method for the determination of nonenzymatic glycosylation of hair and epidermal keratin.

Colorimetric techniques such as the thiobarbituric acid assay are widely used for the determination of nonenzymatic glycosylation of proteins. One of the major problems associated with this technique is the high nonspecific background absorbance which, due to its variability, results in loss of sensitivity. This report describes a method of removing the nonspecific absorbance by extracting the final chromogen into cyclohexanone. Using this method for the determination of nonenzymatic glycosylation of hair and epidermal keratin, at least 90% of the background absorbance is removed, increasing the sensitivity of the technique. This allows more effective discrimination of the level of glycolsylation of proteins from control and diabetic patients, by reducing the degree of overlap. The requirement for individual borohydride reduced samples is also avoided, thus simplifying the technique. Cyclohexanone extraction provides a simple addition to the standard thiobarbituric acid technique with significant improvement in results.

Aged↗
PubMed1984

Early lymphocyte trapping in malaria infections: a particulate antigen mediated phenomenon.

During the course of rodent malaria a marked decrease in the numbers of circulating lymphocytes within the peripheral blood occurred 2-4 days post-infection. Monocytes and polymorphs did not show the same degree of decline. For both avirulent Plasmodium yoelii and lethal Plasmodium berghei infections lymphocyte numbers returned to control levels by day 6-8 post-infection. While these levels were maintained until clearance of P. yoelii infection, a sustained and abnormal increase occurred during P. berghei infection. Early lymphocyte depletion was also observed following Babesia microti and Plasmodium vinckei petteri infections, and could be induced by freeze-thawed P. yoelii infected blood and its particulate, but not soluble, fraction. Corynebacterium parvum and sheep red blood cells had no depressant effect on peripheral blood lymphocyte counts. Cell trapping experiments indicated that peripheral blood lymphocytes were preferentially recruited to the spleen in the initial stages of infection. Cell surface marker tests showed that the major cell type involved was Thy-1.2+ T-lymphocytes.

Animals↗
PubMed1982

Characterization of excretory-secretory antigens of Fasciola hepatica.

Twenty-one day old Fasciola hepatica were recovered from the livers of infected mice and cultured for 5-7 days in a serum-free medium containing either [14C]leucine, [14C]isoleucine or [35S]methionine, or in a medium containing [14C]leucine and serum from sheep vaccinated with excretory-- secretory (ES) antigens of juvenile F. hepatica. All three labelled amino acids were incorporated into fluke proteins. Labelled proteins also appeared in the culture medium. Three major polypeptides detected in the culture media had apparent molecular weights of 26,000, 24,000 and 23,000. All were immunoprecipitated from [14C]leucine-labelled culture medium using antisera against fluke somatic antigens raised in rabbits or from sheep vaccinated with ES antigens of juvenile F. hepatica. A polypeptide of molecular weight 27,000 was also prominent in the culture medium when [14C]isoleucine was used. This polypeptide was present was a minor component when [14C]leucine and [35S]methionine were included in the culture media; it did not appear to be immunoprecipitated by the above antisera from [14C]leucine-labelled culture medium. In the presence of serum from vaccinated sheep, the ES antigens formed immune complexes which contained the polypeptides mentioned above, together with several higher molecular weight polypeptides. Additionally, a number of minor bands of varying molecular weight were present. After micro-Ouchterlony gel immunodiffusion, 2 precipitin lines formed between the labelled ES antigens and antisera. Electrophoresis of these indicated that the 23,000, 24,000 and 26,000 Dalton labelled polypeptides were present in each. The higher molecular weight and the 27,000 Dalton labelled polypeptides were also present in one of the lines.

Animals↗
PubMed1981

Preparation and in vitro translation of mRNA from Fasciola hepatica.

Total RNA was extracted from mature and juvenile Fasciola hepatica by homogenizing in 5.0 M guanidine thiocyanate and centrifugation through a 5.7 M CsCl cushion. Yields of 2 mg/g and 1 mg/g wet weight starting material were obtained, respectively. Messenger RNA was separated from the bulk extracted RNA by binding to oligo(dT)-cellulose. About 25% of the extracted RNA bound in both adult and juvenile cases. This material was subsequently tested in a rabbit reticulocyte cell-free translation system. Up to a 12-fold stimulation of incorporation of [35S]methionine into trichloroacetic acid-precipitable material was observed over that where no message was added. When the in vitro translation products were analysed by autoradiography of SDS-polyacrylamide gradient gels, polypeptides ranging in apparent molecular weight from about 10,000 to 100,000 were observed. Several minor differences in the electrophoretic patterns obtained from juvenile and adult mRNA were observed.

Animals↗
PubMed1981

Precipitating antibodies against excretory/secretory antigens of Fasciola hepatica in sheep serum.

Gel diffusion techniques were used to study antigen-antibody reactions in precipitates forming around juvenile, semi-mature and adult Fasciola hepatica cultured in serum from infected sheep. The number of reactions was analysed in both primary and challenge infections. The number of antigens shared by the various developmental stages of the fluke was also examined. At least 2 antigens were involved in precipitate formation around juvenile flukes. These antigens were also produced by the later stages of development. Two additional antigens were produced by semi-mature flukes and these, together with two others, produced by adult flukes. Challenge infections had little effect on the number of antigens or antibodies in the precipitate.

Animals↗
PubMed1981

Response of sheep to challenge infection with Fasciola hepatica.

Sheep were infected with 100 metacercariae of Fasciola hepatica and reinfected 16 weeks later with a further 100 metacercariae. Serum samples were taken weekly for 36 weeks after primary infection. Serum was assayed for the presence of the enzymes glutamate dehydrogenase (GLDH) and gamma-glutamyl transferase (gamma-GT), as indicators of liver and bile duct damage respectively, and for levels of precipitating antibody. Antibody and GLDH levels rose following the primary infection but fell after patency had been reached . A peak in gamma-GT activity was associated with the onset of patency. After the challenge infection levels of both enzymes rose substantially and there were persistent fluctuations in activity. Antibody levels did not rise markedly following challenge but fluctuated at low levels until autopsy, 20 weeks after challenge. There was no resistance to challenge judged by worm recoveries at autopsy. It is suggested that the presence of adult flukes in the bile ducts suppresses the antibody response to challenge infection. Tissue damage, which is shown by fluctuations in GLDH and gamma-GT levels after adult flukes have become established in the bile ducts, is considered to be due to the feeding activity of adult flukes and the deposition of immune complexes in the liver parenchyma.

Animals↗
PubMed1980

An attempt to vaccinate sheep against Fasciola hepatica using a juvenile fluke antigen sheep antibody complex.

Sheep were injected intramuscularly with a mixture of Freund's complete adjuvant and a precipitated fluke antigen-antibody complex isolated from in vitro cultures of excysted metacercariae of Fasciola hepatica in serum from infected sheep. Control groups of sheep were given Freund's complete adjuvant or were untreated. All sheep were then challenged with 200 metacercariae of F hepatica. Assays of antibody and liver enzymes in serum collected during the experiment indicated that although vaccination with the complex resulted in a pronounced antibody response to its antigenic component(s), there was no apparent effect on the migratory activity of the juvenile flukes in these animals. Failure to confer protection on vaccinated sheep was confirmed at autopsy when no significant difference was found between the worm burden of these animals and the controls.

Animals↗