Biomedical subjects
J D Fraser
Publications and source records attributed to J D Fraser.
Enterotoxin residues determining T-cell receptor V beta binding specificity.
Superantigens such as the staphylococcal enterotoxins bind to major histocompatibility complex (MHC) class II molecules and activate T cells through a specific interaction between the V beta region of the T-cell antigen receptor (TCR) and the toxin. The TCR beta-chain alone is sufficient to produce the interaction with the enterotoxin-class II complex. Identification of the regions of enterotoxins that interact with TCR has so far proved equivocal because of difficulties in distinguishing between direct effects on T-cell recognition and indirect effects resulting from alteration of binding to class II. For example, amino-terminal truncations of SEB abrogated T-cell stimulation whereas carboxy-terminal truncation of SEA stopped its mitogenic activity. The most comprehensive study to date, accounting for both enterotoxin binding to class II and enterotoxin interactions with the TCR, identified two functionally important regions for SEB binding to TCR. Although the amino-acid sequences of staphylococcal enterotoxins A and E are 82% identical, they activate T cells bearing different V beta elements. We have assayed the binding of cells coated with these enterotoxins to soluble secreted TCR beta-chain protein and find that V beta 3 binds enterotoxin A but not E, whereas V beta 11 binds enterotoxin but not A. To map the amino-acid residues responsible for these different binding specificities, we prepared a series of hybrids between the two staphylococcal enterotoxins. We report that just two amino-acid residues near the carboxy terminus of the enterotoxins are responsible for the discrimination between these molecules by V beta 3 and V beta 11.(ABSTRACT TRUNCATED AT 250 WORDS)
Zinc regulates the function of two superantigens.
Staphylococcal enterotoxins bind with high affinity to class II major histocompatibility complex proteins and subsequently stimulate large numbers of T cells via the V beta portion of the T-cell receptor. Binding of enterotoxin A and enterotoxin E to HLA-DR was completely abolished by low levels of EDTA, whereas binding of toxic shock toxin was unaffected. Addition of Zn2+ to as little as 2 microM excess over EDTA completely reconstituted binding, but Ca2+, Mg2+, Cu2+, Fe2+, and Mn2+ had no effect. The dissociation constant (Kd) of 65Zn2+ binding to a single site on purified enterotoxin A was 2 microM, and addition of purified HLA-DR1 did not alter the Kd, indicating that the binding site was exclusive to enterotoxin A. In the presence of saturating levels of zinc the Kd for enterotoxin A binding to purified HLA-DR1 was 25 nM. Thus, zinc binding is an essential first step in the formation of the major histocompatibility complex binding domain of at least two bacterial superantigens. Given the measured Kd of zinc binding to enterotoxin A, serum levels of free zinc (0.2-1.0 microM) may well regulate the toxic sequelae by these two superantigens.
CD28 and T cell antigen receptor signal transduction coordinately regulate interleukin 2 gene expression in response to superantigen stimulation.
Activation of an immune response requires intercellular contact between T lymphocytes and antigen-presenting cells (APC). Interaction of the T cell antigen receptor (TCR) with antigen in the context of major histocompatibility molecules mediates signal transduction, but T cell activation appears to require the induction of a second costimulatory signal transduction pathway. Recent studies suggest that interaction of CD28 with B7 on APC might deliver such a costimulatory signal. To investigate the role of CD28 signal transduction during APC-dependent T cell activation, we have used Staphylococcal enterotoxins (SEs) presented by a B7-positive APC. We used anti-B7 monoclonal antibodies and a mutant interleukin 2 (IL-2) promoter construct, unresponsive to CD28-generated signals, in transient transfection assays to examine the contribution of the CD28-B7 interaction to IL-2 gene activation. These studies indicate that the CD28-regulated signal transduction pathway is activated during SE stimulation of T cells and plays an important role in SE induction of IL-2 gene expression through its influence upon the CD28-responsive element contained within the IL-2 gene promoter. This effect is particularly profound in the activation of the IL-2 gene in peripheral blood T cells.
Regulation of T-cell lymphokine gene transcription by the accessory molecule CD28.
T-cell activation results in the production of multiple lymphokines. Efficient lymphokine gene expression appears to require both T-cell antigen receptor (TCR) signal transduction and an uncharacterized second or costimulatory signal. CD28 is a T-cell differentiation antigen that can generate intracellular signals that synergize with those of the TCR to increase T-cell activation and interleukin-2 (IL-2) gene expression. In these studies, we have examined the effect of CD28 signal transduction on granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukin 3 (IL-3), and gamma interferon (IFN-gamma) promoter activity. Stimulation of CD28 in the presence of TCR-like signals increases the activity of the GM-CSF, IL-3, and IFN-gamma promoters by three- to sixfold. As previously demonstrated for the IL-2 promoter, the IL-3 and GM-CSF promoters contain distinct elements of similar sequence which specifically bind a CD28-induced nuclear complex. Mutation of the CD28 response elements in the IL-3 and GM-CSF promoters abrogates the CD28-induced activity without affecting phorbol ester- and calcium ionophore-induced activity. UV cross-linking indicates that the CD28-induced nuclear complex contains polypeptides of approximately 35, 36, and 44 kDa. These studies indicate that the TCR and CD28-regulated signal transduction pathways coordinately regulate the transcription of several lymphokines and that the influence of CD28 signals on transcription is mediated by a common complex.
The zeta chain is associated with a tyrosine kinase and upon T-cell antigen receptor stimulation associates with ZAP-70, a 70-kDa tyrosine phosphoprotein.
Stimulation of the T-cell antigen receptor (TCR) leads to tyrosine phosphorylation of a number of cellular proteins, including phospholipase C (PLC) gamma 1 and the TCR zeta chain. We describe here a 70-kDa tyrosine phosphoprotein (ZAP-70) that associates with zeta within 15 sec following TCR stimulation. The phosphorylation of ZAP-70 and its association with zeta is independent of the other TCR chains since stimulation of a functional CD8/zeta chimeric receptor in a TCR-negative T cell leads to coprecipitation of ZAP-70 with the chimeric protein. In a Jurkat cell expressing the TCR and the CD8/zeta chimeric protein, tyrosine phosphorylation and association of ZAP-70 occurs exclusively with the stimulated receptor complex. In addition, a tyrosine kinase that does not appear to be fyn associates with the cytoplasmic domain of zeta and phosphorylates zeta and ZAP-70 in vitro.
Regulation of interleukin-2 gene enhancer activity by the T cell accessory molecule CD28.
The mechanism by which cell surface molecules regulate T cell production of lymphokines is poorly understood. Production of interleukin-2 (IL-2) can be regulated by signal transduction pathways distinct from those induced by the T cell antigen receptor. Stimulation of CD28, a molecule expressed on most human T cells, induced the formation of a protein complex that bound to a site on the IL-2 gene distinct from previously described binding sites and increased IL-2 enhancer activity fivefold. The CD28-responsive complex bound to the IL-2 gene between -164 and -154 base pairs from the transcription start site. The sequence of this element is similar to regions conserved in the 5' flanking regions of several other lymphokine genes.
CT findings in lipoma of the flexor retinaculum of the palm.
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Survey of cardiac pacing in Canada.
UNLABELLED: The status of cardiac pacing in Canada in 1989 was determined from data provided by 62 of 128 physicians surveyed (48% response) and four major manufacturer/distributors. A questionnaire designed for the IXth World Symposium on Cardiac Pacing was used. DEMOGRAPHICS: There were five implant hospitals per million population, 65% community based and 35% university affiliated; 63% of implanters were surgeons. There were 279 new implants and 46 replacement procedures per million population. INDICATIONS: Sinus node disorders accounted for 44.6% of implants, atrioventricular block 43.2% (fixed 24.4%, intermittent 12.0%, incomplete 6.8%), tachycardias 2.9%, drug-induced bradycardia 3.1%, and other (including automatic implantable cardioverter defibrillators) 6.2%. TECHNOLOGY: Single chamber units were implanted in 78.6% of patients, and dual chamber in 22.7%, and 19.5% of the total were rate-adaptive. Unipolar leads were used in 57.1% of atrial and 53.2% of ventricular insertions; 40.4% of atrial and 5.8% of ventricular leads were active fixation. The pervenous sheath introducer technique was used in 64.9% of lead insertions. PERIOPERATIVE: Major complications occurred in 2.6% of single and 6.8% of dual chamber primary implants, but mortality was less than 0.1%; 8.4% of replacements were unanticipated; there was no known death from malfunction. Mean hospital stay was 2.7 days for primary implants and 1.4 days for replacement/revisions. CONCLUSIONS: Comparison with prior surveys (1979, 1981, 1985) reveals: increased physician response to the survey questionnaire; relatively stable electrocardiographic indications for implant; a modest increase of new implants per million; continued decrease in replacements; an increase in dual chamber and rate-adaptive pacing; and increased use of active fixation and bipolar electrodes in both atrium and ventricle.
Induction of matrix Gla protein synthesis during prolonged 1,25-dihydroxyvitamin D3 treatment of osteosarcoma cells.
The synthesis of matrix Gla protein (MGP) and bone Gla protein (BGP) have been shown to be mutually exclusive in all osteosarcoma cell lines investigated. In the cell lines that produce the respective proteins, synthesis is stimulated by 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) within the first several hours of hormone treatment. In the present studies we have investigated the effects of longer-term treatment with 1,25(OH)2D3 in the ROS 17/2 cell line, a cell line that synthesizes BGP constitutively but does not synthesize MGP. In agreement with earlier studies, the rate of BGP synthesis increases within 8 hours of hormone treatment, is maximal by 24 hours, and remains at the maximal rate through 48 hours of 1,25(OH)2D3 treatment. The present study is the first to report that the rate of BGP secretion at times beyond 48 hours declines to that of control cultures despite the continued administration of 1,25(OH)2D3, and that MGP synthesis is induced in ROS 17/2 cells by 48 hours of 1,25(OH)2D3 treatment. At this time, MGP mRNA could be detected by northern blot analysis and MGP secretion could be demonstrated by radioimmunoassay of culture medium. Both the level of MGP message per unit total RNA and the rate of MGP secretion into culture medium increased steadily between 2 and 6 days of 1,25(OH)2D3 treatment. The MGP synthesized by the 1,25(OH)2D3-treated ROS 17/2 cells was identical to that found in bone by northern blot analysis of message and by western blot analysis of the media antigen. Half-maximal induction of MGP synthesis was obtained with 0.3 nM 1,25(OH)2D3, a 60-fold higher dosage than was required for the half maximal stimulation of BGP synthesis in these cells. Treatment of ROS 17/2 cells with 24,24-F21,25(OH)2D3 suggests that the observed difference in dose dependence is not due to an increased rate of hormone catabolism.
Treatment of early breast cancer.
The early follow-up of patients treated by simple mastectomy alone or simple mastectomy combined with radical radiotherapy is presented. Both groups were well matched for age, menopausal status, duration of symptoms, size of tumour, and lymph-node involvement. There was no significant difference in survival of patients in the two groups at three years, but local recurrence was significantly more frequent (28%) in the mastectomy-alone patients. Early survival was not adversely affected by radiotherapy.
Autoantibodies in early breast cancer: a stage-related phenomenon?
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Host responses to cryoablation of normal kidney and liver tissue.
Following cryoablation of one kidney in Strain-2 guinea-pigs, the viable opposite kidney was assessed by functional, histological and immunopathological techniques for evidence of immunologically mediated damage. The autoantibody content of postoperative sera was also examined. Control operations involved ligation, surgical nephrectomy and the production of hepatic cryolesions. The results indicate that autoantibody production and glomerular deposition of immune complexes can occur and may cause abnormal passage of hippuran through the kidney. There is no evidence for autoallergy against highly tissue-specific determinants; the specificity, strength and time course of the reactions obtained was dependent on the site of the lesion, whether or not freezing was accompanied by ligation.