Biomedical subjects

J D Sedgwick

Publications and source records attributed to J D Sedgwick.

69 records · Page 4Linked to original sources

PubMed1988

Long-term depletion of CD8+ T cells in vivo in the rat: no observed role for CD8+ (cytotoxic/suppressor) cells in the immunoregulation of experimental allergic encephalomyelitis.

A method is described for the long-term depletion of CD8+ T cells in the rat. Employing Lewis rats depleted of this cell type it is shown that such animals acquire, and subsequently recover from both passively and actively induced experimental allergic encephalomyelitis (EAE) in a comparable fashion to normal animals. Furthermore, after spontaneous recovery from the initial bout of disease, CD8+ T cell-depleted rats do not show any signs of relapse and, like normal rats following recovery from active EAE, are still resistant to further attempts at disease induction. Overall, these data do not support a role for the classic cytotoxic/suppressor (CD8+) T cell in the immunoregulation of EAE in the Lewis rat.

Animals↗

PubMed1987

Suppression of IgE responses by antigen inhalation: studies on the role of genetic and environmental factors.

Repeated inhalation of low levels of ovalbumin (OVA) by mice or rats preferentially induces tolerance in the IgE antibody class, and this process may represent an important protective mechanism that normally prevents allergic sensitization to air-borne antigens. Dose-response experiments involving exposure of a number of inbred rat strains to graded doses of aerosolized OVA confirmed the inverse relationship between sensitivity to tolerogenesis and IgE-responder phenotype. These experiments additionally demonstrated that F1 hybrids derived from low X high responder crosses co-inherited high sensitivity to tolerance induction, together with the low IgE-responder phenotype. Sensitivity to tolerance induction in low versus high IgE-responder strains was found to be independent of the route of administration of OVA, indicating that the relevant genetically determined control mechanism(s) operated systemically. However, pre-exposure of animals to a variety of exogenous agents, noteably inhaled irritants (NO2 and histamine), the inflammatory adjuvants pertussigen and aluminium hydroxide injected at sites that stimulate the regional lymph nodes draining the respiratory tract, or a single subcutaneous injection of the reticuloendothelial system stimulator oestradiol, were shown to partially abrogate this natural tolerance process and promote allergic sensitization.

Administration, Inhalation↗

PubMed1986

The ELISA-plaque assay for the detection and enumeration of antibody-secreting cells. An overview.

A solid-phase immunoenzymatic technique has been developed which allows the ready detection and enumeration of total- and antigen-specific immunoglobulin-secreting cells (ISC). The procedure involves the addition of putative ISC to plastic wells pre-coated with specific antigen or antisera. During incubation, the product of a single cell is immobilized by the solid phase at the point of release providing an immunological 'finger print' of the ISC which is subsequently developed by the application of appropriate enzyme-anti-Ig conjugates and an enzyme substrate which yields an insoluble product after incubation. Blue spots or 'ELISA plaques' are thus produced and can be counted macroscopically. This technique has been employed in rat, mouse and human systems and in each case appears to be of equivalent or greater sensitivity to existing haemolytic plaque techniques. The assay is particularly suited to the enumeration of antigen-specific ISC in which the antigen is difficult to couple to red cells or where a high degree of discriminating power is necessary as is required for example in the enumeration of IgE-ISC.

Antibody Affinity↗

PubMed1986

The mechanism of inhibition of experimental allergic encephalomyelitis in the rat by monoclonal antibody against CD4.

Lewis rats with actively induced or passively transferred experimental allergic encephalomyelitis (EAE) were treated with a monoclonal antibody (MAb) which binds to the CD4 antigen of rat helper/inducer T cells. Actively immunized animals treated at the first onset of clinical signs experienced only a mild form of the disease and rapidly recovered while the majority of those treated prophylactically never showed clinical signs of EAE. Passively transferred EAE was also completely inhibited with anti-CD4 MAb. In treated animals which exhibited only mild clinical signs of EAE, spinal cord and cerebellar leukocyte infiltrates were quite similar to those in untreated rats but where anti-CD4 MAb treatment completely prevented clinical EAE, histological signs were minimal or absent. Like Lewis rats which have recovered naturally from EAE, those treated with anti-CD4 MAb were both resistant to a secondary challenge with myelin basic protein and harboured potential encephalitogenic cells which were capable of transferring disease to recipient rats. Disease in these recipients was, however, of much greater severity than that experienced by animals receiving cells from naturally recovered (untreated) donors. These data demonstrate that administration of anti-CD4 MAb to rats can prevent EAE by a mechanism which does not ablate the encephalitogenic CD4+ cells or prevent the development of resistance to EAE but which may inhibit the disease by preventing the function of already activated effector cells.

Animals↗

PubMed1986

Induction of IgE-secreting cells in the lymphatic drainage of the lungs of rats following passive antigen inhalation.

Repeated challenge of high-IgE-responder BN rats with an antigen aerosol in the absence of adjuvant, elicited both specific IgE and IgG responses in serum. Employing the recently developed ELISA plaque assay to detect individual IgE and IgG secreting cells, the lymph nodes draining the lower respiratory tract were pinpointed as the site for initiation of this response, and the sole location of specific IgE-secreting cells. Subsequent analysis of the tissue distribution of total IgE-secreting cells revealed that greater than or equal to 50% were located in the lymph nodes draining the upper and lower respiratory tracts.

Aerosols↗

PubMed1985

Induction of IgE-secreting cells and IgE isotype-specific suppressor T cells in the respiratory lymph nodes of rats in response to antigen inhalation.

Repeated exposure of high-IgE-responder Brown Norway (BN) rats to an aerosol of ovalbumin (OVA) once weekly triggered progressively increasing levels of OVA-specific IgG in serum. In contrast, responses in the IgE class were transient, declining from peak titers during the third week to background levels by Week 5, despite continuing aerosol exposure. Subsequent parenteral challenge of these animals revealed a state of antigen- and IgE isotype-specific tolerance. Adoptive transfer of splenocytes or pooled respiratory tract lymph node (RTLN) cells from aerosol-exposed animals to naive rats abrogated subsequent OVA-specific primary IgE responses in the recipients, but did not affect specific IgG responses, and kinetic studies indicated that these suppressor cells arose first in the RTLN. Transfer studies employing individual lymph node groups which constituted the RTLN pool pinpointed the superficial cervical nodes, which drain the uppermost portion of the respiratory tract, as the major source of suppressor cells. Fractionation of cell populations before adoptive transfer employing monoclonal antibodies directed against T-cell markers, defined a population of suppressor cells generated by aerosol exposure which expressed both the W3/13 (pan T-cell) and OX8 (cytotoxic/suppressor T-cell) antigens, but which was negative for the W3/25 (helper T-cell) marker. Analysis of the IgE and IgG responses induced by OVA inhalation was performed employing the ELISA plaque technique, recently developed in this laboratory. These studies revealed the parathymic and posterior mediastinal nodes draining the lower lung, as the major sites of specific IgE and IgG production; smaller numbers of OVA-specific IgG-secreting cells (but none secreting specific IgE) were detected in the nodes draining the upper respiratory tract, while antibody secretion outside the respiratory tract was restricted to comparatively few cells in the spleen. The ELISA plaque assay was also employed to enumerate total numbers of cells secreting the IgE isotype in aerosol-exposed and control rats, employing samples from 10 different lymphoid organs. Approximately 50% of the IgE-secreting cells in these animals were localized in RTLN, as opposed to 25% in gut-associated lymphoid tissues. These data are discussed in relation to the pivotal role of respiratory-tract associated lymphoid tissues in regulation of IgE responses to aeroallergens.

Aerosols↗

PubMed1985

Down-regulation of immune responses to inhaled antigen: studies on the mechanism of induced suppression.

Repeated exposure of rats to an aerosol of ovalbumin (OVA) or its dinitrophenylated derivative (DNP-OVA) induced carrier-specific tolerance to subsequent challenge with the same haptenated antigen. Following parenteral challenge with DNP-OVA, both anti-DNP and anti-OVA IgE titres were reduced relative to controls, whereas anti-DNP responses following challenge with DNP-Ascaris were normal. Stimulation of tolerant rats with OVA, together with the polyclonal B-cell mitogen LPS, restored their capacity to respond to the antigen. In contrast to WAG rats, which have previously been shown to develop equivalent tolerance in the IgE an IgG antibody classes (Sedgwick & Holt, 1984), BN rats exposed to an OVA aerosol developed high serum titres of anti-OVA IgG. Following parenteral challenge with DNP-OVA, however, anti-DNP IgG responses in the BNs were markedly reduced relative to unexposed controls, while anti-OVA IgG titres were maintained at a high level. Further strain-dependent differences in T-cell function in tolerized rats appeared in in vivo assays of DTH reactivity and in in vitro antigen-driven lymphocyte proliferation. Both BN and WAG rats displayed diminished in vitro responses, whereas DTH reactions were only suppressed in the latter strain.

Aerosols↗

PubMed1984

ELISA plaque assay for the detection of antibody secreting cells: observations on the nature of the solid phase and on variations in plaque diameter.

This report examines the basis for variations in the size and number of ELISA plaques detected in vitro. The nature and concentration of antigen used to coat the solid phase is shown to be critical, high concentrations of polymerized antigen being optimal for low molecular weight proteins, while coating conditions for high molecular weight multideterminant antigens are similar to conventional liquid ELISA. Variations in plaque diameter are shown to reflect maturational changes in the immune response, and probably indicate differences in the affinity of antibody being secreted within a population of plasma cells.

Animals↗

PubMed1984

Suppression of IgE responses in inbred rats by repeated respiratory tract exposure to antigen: responder phenotype influences isotype specificity of induced tolerance.

Repeated exposure of rats to an aerosol of ovalbumin (OVA) induced tolerance to subsequent parenteral challenge with the same antigen. In the low-IgE responder WAG strain, responses in both the IgE and IgG classes were affected, whereas rats of the moderate (Lou/M) and high-IgE responder BN strain developed high titers of anti-OVA IgG in serum during exposure with concomitant tolerance in the IgE class. Repeated parenteral challenge, however, failed to elicit significant secondary anti-OVA IgG responses in the Lou/M and BN strains, suggesting that the isotype specificity of induced tolerance in these strains was not absolute. Spleen and respiratory tract lymph node cells, but not serum from aerosol-exposed BN rats, were capable of transferring IgE isotype- and antigen-specific tolerance. Dose response experiments demonstrated that the low-IgE responder WAG strain was exquisitely sensitive to tolerance induction in response to antigen inhalation, being susceptible to dosages in the nanogram range; at least 1000 times more antigen was required in the high-IgE responder BN to induce comparable tolerance in the IgE class. It was also apparent that the IgE isotype was more readily suppressed than the IgG isotype in both high- and low-IgE responder strains.

Aerosols↗

PubMed1984

Long-lived IgE- and IgG-secreting cells in rodents manifesting persistent antibody responses.

BALB/c mice and BN rats manifesting persistent IgE and IgG responses were examined up to 1 year after immunization. A significant proportion of the ongoing antibody response in these animals survived lethal X-irradiation employing dosages sufficient to deplete B memory cells. The persistent IgE responses in both species were refractory to exogenous isotype-specific suppressor cells taken from tolerant syngeneic animals, which were shown to abrogate primary IgE responses in parallel tests. Employing a novel ELISA-based assay for plaque forming cells, long-lived radioresistant IgE- and IgG-secreting cells were identified in differing ratios in lymph nodes, spleen, and bone marrow of both species. These long-lived cells were shown to arise following maximum antigenic challenge with antigen plus adjuvant, and after repeated low-grade stimulation by antigen alone, including passive inhalation of dilute antigen aerosols.

Animals↗

PubMed1984

Enumeration of human immunoglobulin-secreting cells by the ELISA-plaque method: IgE and IgG isotypes.

A new system is described for the enumeration of human immunoglobulin-secreting cells (ISC), based upon the ELISA methodology. In principle, putative ISC are incubated over a solid phase containing bound anti-Ig of the isotype being tested. Secreted Ig is immobilized at or near the point of release from the ISC, and the resulting Ig fingerprint of the ISC is then visualized by the sequential application of an anti-Ig-alkaline phosphatase conjugate, followed by a substrate-agarose overlay. The system is capable of detecting IgE-secreting cells, and pokeweed mitogen-stimulated IgG-secreting cells with sensitivity at least equivalent to the protein A hemolytic plaque assay.

Animals↗

PubMed1983

Kinetics and distribution of antigen-specific IgE-secreting cells during the primary antibody response in the rat.

A new assay system is described for the enumeration of antigen-specific IgE immunoglobulin-secreting cells (ISC) based on an enzyme-linked immunoabsorbent assay. Using this technique to monitor the organ distribution of OVA-specific ISC after primary immunization of rats, approximately 12,000 specific IgE ISC were detected at the peak of the response in the draining lymph nodes compared with 117,000 IgG ISC; the splenic anti-OVA response was restricted to the IgM class. Using plates precoated with anti-rat IgE instead of antigen, total IgE ISC were enumerated in normal and helminth-parasitized rats. The assay system detected up to 5 X 10(5) IgE ISC in mesenteric lymph nodes from parasitized animals compared with less than 50 in controls.

Animals↗

PubMed1983

A solid-phase immunoenzymatic technique for the enumeration of specific antibody-secreting cells.

A sensitive immunoassay is described for the detection of idiotype- and isotype-specific antibody-secreting cells (ASC), based upon the well established principles of ELISA. Single cell suspensions containing ASC are incubated on solid phase to which specific antigen has been chemically conjugated. Antibody attaches to the latter within the immediate microenvironment of the ASC, producing localized zones of bound antibody which are subsequently developed as visual 'spots' in the ELISA. This assay system has sensitivity and specificity at least equivalent to haemolytic plaque assays.

Animals↗

PubMed1983

Induction of IgE-isotype specific tolerance by passive antigenic stimulation of the respiratory mucosa.

Repeated exposure of high IgE-responder BN rats to an aerosol of ovalbumin (OVA) induced tolerance which was specific both for the antigen and the IgE antibody class. Radiotracer studies with 125I-OVA indicated that inhaled antigen doses in the low microgram range were tolerogenic, the bulk of the inhaled antigen being distributed 2:1 between the gastro-intestinal and respiratory tracts. Subsequent gastric intubation studies showed that the gastric component did not contribute to tolerance induction, suggesting that antigenic stimulation of the respiratory mucosa was central to this process.

Aerosols↗

PubMed1981

Production of a histamine-releasing lymphokine by antigen- or mitogen-stimulated human peripheral T cells.

Human peripheral blood leucocytes (PBL), cultured in the presence of mitogen or antigen, yielded supernatants which contained a factor with histamine-releasing activity (HRA). This factor, analogous to the putative lymphokine HRA described by Thueson et al. (1979a), was shown to promote the rapid release of histamine from PBL preparations containing basophils. HRA production by PBL in response to co-culture with the skin test antigens Candida and SK-SD was shown to reflect the in vivo immune status of the PBL donor. Experiments with isolated cell populations from PBL implicated T cells as the prime source of HRA, production of the factor being macrophage-dependent. HRA activity failed to synergize anti-IgE-mediated histamine release, and was not species-restricted.

Adult↗