Biomedical subjects

C R Stokes

Publications and source records attributed to C R Stokes.

At least 55 records · Page 3Linked to original sources

PubMed1994

Workshop studies on monoclonal antibodies reactive against porcine myeloid cells.

Investigators from eight laboratories analyzed the reactivity of 22 monoclonal antibodies (mAb) against porcine myeloid cells. Based on binding data, clustering analysis and inhibition studies, workshop mAb 74-22-15 (003) and 6F3 (007) were assigned a swine workshop cluster number 3 (SWC3). These mAb recognized macrophages and neutrophils; neutrophils; a monocyte/macrophage-specific mAb was not identified by this workshop.

Animals↗

PubMed1994

The effect of delayed weaning on the development of oral tolerance to soya-bean protein in pigs.

The antibody response to a dietary antigen (soya-bean protein) and the development of oral tolerance was studied in pigs in a family pen system where the piglets are left with their mothers and gradually wean themselves onto a soya-bean-based diet over a 12 week period. In the first experiment three groups of pigs (eight pigs/group) aged either 2, 8 or 13 weeks were immunized with soya-bean protein or ovalbumin (OvA; controls) intra-peritoneally (i.p.) in Quill A adjuvant and subsequently boosted 2 weeks later. All groups showed an IgG response to the injected antigens indicating lack of tolerance induction to the dietary antigen. Interestingly the groups injected with OvA showed an almost identical response to soya-bean protein as the groups injected with soya-bean protein. In a second experiment with a similar protocol, soya bean was withdrawn from the feed before immunization which resulted in lack of response to soya-bean protein in the groups injected with OvA and a lack of response to injected soya-bean protein in the 14-week-old group, indicating that systemic tolerance was established by 12 weeks of age. The results from the two experiments suggest a compartmentalized response to soya-bean protein i.e. local antibody production to dietary soya bean along with systemic tolerance to injected soya-bean protein. The work also suggests that delayed 'natural' weaning may avoid damaging hypersensitivity reactions associated with early weaning.

Aging↗

PubMed1994

Altered immune response to proteins fed after neonatal exposure of piglets to the antigen.

The weaning of piglets onto soya proteins at 3 weeks old normally results in an active response to the fed protein, as determined by the appearance of serum IgG antisoya antibody. This system thus allows the effects of manipulation on the response to a fed protein to be studied. In animals previously given 1 g of soya protein at birth, the magnitude of the antibody response to soya fed at 3 weeks was decreased, although similar amounts of the fed protein could be detected in serum. In addition, the relative affinity of the dominant interaction between antigen and antibody was reduced in these piglets by almost an order of magnitude. By comparison, the ability of piglets given soya at birth to respond to injected soya was not significantly reduced. These results indicate that the regulation of responses to fed and systemic antigens is largely separate. Very early oral exposure to antigen may affect the ability of neonatal animals to mount immune responses to, specifically, fed proteins while leaving the response to systemic antigen largely intact.

Animals↗

PubMed1994

Cytokine-induced major histocompatibility complex class II antigens on cultured bovine mammary gland epithelial cells.

The aim of this work was to establish a simple but reliable method of collecting epithelial cells from tissue lining the gland cistern of the bovine mammary gland and to investigate the ability of cultured epithelial cells to express major histocompatibility complex class II antigens. Bovine mammary gland epithelial cells were isolated and grown as monolayers on five substrates in vitro. The cells were identified as epithelial in origin by immunofluorescent staining with anticytokeratin monoclonal antibodies. Major histocompatibility complex class II antigens were induced on epithelial monolayers by incubation with supernatants from bovine peripheral blood mononuclear cells stimulated with concanavalin A or incubation with recombinant bovine interferon-gamma. These results suggest that, in bovine mammary gland epithelium, recombinant bovine interferon-gamma, acting alone, can induce class II expression. This observation permits future investigation of the putative role of bovine mammary gland epithelial cells as accessory cells in the initiation of local immune responses and their involvement in peptide transport.

Animals↗

PubMed1994

Production of cytokines by lymphocytes from spleen, mesenteric lymph node and intestinal lamina propria of pigs.

Large numbers of cells can be recovered from pig intestine with phenotypes suggesting lamina propria rather than intraepithelial origin. Following activation with concanavalin A these cells produced a T-cell growth factor (TCGF) activity which was not inhibited in the presence of a monoclonal antibody recognizing pig interleukin-2 receptors (IL-2R). In contrast, the activity of recombinant human IL-2 and of supernatants from activated spleen cells was almost entirely inhibited by anti-IL-2R. The failure of anti-IL-2R to inhibit the activity of lamina propria-derived TCGF was not apparently owing to interference by soluble receptor with binding of monoclonal to target blast cells as no effect of supernatants on binding was observed. The results suggest that cells derived from the pig intestinal lamina propria fail to produce IL-2 following polyclonal activation in vitro. Consistent with this finding, IL-2 transcripts could be detected by polymerase chain reaction (PCR) following reverse transcription of mRNA derived from spleen and from mesenteric lymph node but not lamina propria lymphocytes, while IL-4 cDNA could be detected from all three sources.

Animals↗

PubMed1993

Epidemiological and diagnostic features of canine and feline dermatophytosis in the United Kingdom from 1956 to 1991.

Between 1956 and 1991, 8349 samples from dogs and cats were received for investigation of suspected dermatophytosis, and 1368 (16 per cent) yielded positive cultures. Cats had a significantly higher proportion of positive cultures (26 per cent) than dogs (10 per cent), and of these Microsporum canis accounted for 92 per cent in cats and 65 per cent in dogs. The other isolates were diverse but mainly sylvatic dermatophytes, and M gypseum was isolated on only four occasions. Different breeds of dog and cat had significantly different prevalences of infection, with pedigree and long-haired cats, and Jack Russell and Yorkshire terrier dogs having a particularly high proportion of positive cultures. Animals less than one year old appeared to be predisposed to infection, but there was no apparent sex predisposition and no conclusive evidence of any seasonal variation in the incidence of the disease. In comparison with the results of dermatophyte culture, examination under Wood's lamp had a positive predictive value of 90 per cent and a negative predictive value of 94 per cent in determining M canis infection, and direct microscopy had positive and negative predictive values of 93 per cent in determining the presence of dermatophytosis. However, cultural examination alone was insufficient for the diagnosis of dermatophytosis owing to the occurrence of false positive and false negative results.

Age Factors↗

PubMed1993

The separation and identification by monoclonal antibodies of dog IgG fractions.

Four fractions of IgG from normal dog serum have been successfully isolated by gel filtration followed by protein A and protein G affinity chromatography using the fast protein liquid chromatography (FPLC) system. Protein A chromatography produced three peaks: peak 1 was fallthrough material consisting of components which did not bind to protein A, peak 2 consisted of bound material eluting at pH 6, and peak 3 contained bound material eluting at pH 3.5. The three peaks were then subjected individually to protein G affinity chromatography. Peak 1 from protein A chromatography produced a fallthrough peak followed by a weakly binding component which eluted at pH 8, and was called peak w. Peak 2 from protein A chromatography bound to protein G and eluted as a single peak at pH 3.8, and was called peak x. Peak 3 from protein A chromatography emerged as two separate peaks (y and z) off the protein G column; peak y bound and eluted at pH 4.1, and peak z bound weakly to protein G and emerged as a broad band at pH 8. Peaks w, x, y and z have been named gamma w, gamma x, gamma y and gamma z, respectively, and there purified IgG fractions were used to immunize mice for the preparation of monoclonal antibodies (McAbs). To date, two sets of McAbs have been produced: one which recognizes an epitope present in both gamma w and gamma z fractions and another set of McAbs which recognizes an epitope in the gamma x and gamma y fractions.

Animals↗

PubMed1993

Nucleotide and deduced amino acid sequence of porcine interleukin 4 cDNA derived from lamina propria lymphocytes.

Total RNA was isolated from in vitro activated lamina propria lymphocytes and used to direct the synthesis of cDNA. Interleukin 4 transcripts were then specifically amplified by PCR. Comparison of the nucleotide sequence with its human homologue demonstrates deletion within the coding region of pig interleukin 4 centred around amino acid residue 70 in the mature human protein.

Amino Acid Sequence↗

PubMed1993

Maturation of cellular defence in the respiratory tract of young calves.

The maturation of respiratory tract defence was investigated in a longitudinal study of calves during the first 100 days of life. From day 7, the proportions of the cell types identified in bronchoalveolar lavage (BAL) fluid were similar to those found in adults, with a predominance of alveolar macrophages over polymorphonuclear neutrophils (PMNs) and lymphocytes. Functionally, bactericidal activity of BAL cells was defective and for the first 21 days they supported intracellular bacterial growth. At 24 hours of life, the movement of peripheral blood neutrophils to a chemotactic source was poor, but this increased rapidly during the first week of life. Like BAL cells, peripheral blood PMNs supported intracellular bacterial growth for the first two weeks of life. These studies suggest that cellular defence mechanisms may be compromised during the first week of life.

Animals↗

PubMed1993

Immune cell distribution in the small intestine of the pig: immunohistological evidence for an organized compartmentalization in the lamina propria.

Using monoclonal antibodies in immunohistochemistry, the distribution of the cells with the following surface antigens was studied in samples of proximal and distal small intestine of five 6-month-old pigs: CD2, CD4 (helper/inducer T-cells), CD8 (suppressor/cytotoxic T cells), accessory cell marker (monocyte/granulocyte), MHC Class II (DRw), and interleukin 2 (IL-2) receptor. CD2+ cells were found in high numbers in both the epithelium and the lamina propria. More cells were demonstrated in villis than in crypts (proportion approximately 4:1). At least two subpopulations of intraepithelial lymphocytes were identified: apically in the epithelium there were CD2+CD4-CD8- (double negative) cells, whereas cells expressing CD8 marker were concentrated around the basement membrane. CD4+ cells were localized in the lamina propria towards the villus core. Accessory cells were distributed in crypts and the villus base and more cells were found in ileum than in duodenum. In contrast, MHC Class II+ cells were located predominantly in villi, just underneath the basement membrane, forming a sheath of cells between the CD8+ and the CD4+ cells. Cells expressing IL-2 receptor were sparse but widely distributed in both the lamina propria and the epithelium. This organized cell distribution may be related to the physiology of the mucosal immune system in the gut.

Animals↗

PubMed1993

Expression of major histocompatibility complex (MHC) class II antigens in the murine mammary gland.

Major histocompatibility complex (MHC) class II antigens were identified on cells within mammary gland connective tissue of lactating mice using a paraformaldehyde-lysine-periodate-gluteraldehyde fixative and an immunoperoxidase staining method. The distribution of class II expressing cells within interalveolar and interlobular connective tissue was similar both throughout lactation and in successive lactations. Epithelial cells within secretory alveoli and mammary ducts did not express class II antigens.

Animals↗

PubMed1993

Adjuvant action of cholera toxin and pertussis toxin in the induction of IgA antibody response to orally administered antigen.

The ability of cholera toxin B-subunit to bind to intestinal epithelium and in particular the dome epithelium of the Peyer's patch can account for its potency as an immunogen. However, pure B-subunit is a less effective immunogen than whole cholera toxin. The immunogenicity of B-subunit may be restored by the addition of either traces of whole toxin or by pertussis toxin. Cholera toxin and pertussis toxin were both able to stimulate a response when fed in conjunction with keyhole limpet haemocyanin, whereas recombinant B- subunit of heat-labile toxin from Escherichia coli had no effect, demonstrating that the adjuvant action is a property of the enzymically active A-subunits. The adjuvant activity of both pertussis toxin and cholera toxin may be due to their ability to cause an increase in the activity of adenylate cyclase via their action on GTP-binding regulatory proteins. However, feeding of forskolin, a direct activator of adenylate cyclase, had no effect on the mucosal immune response, indicating a role for cholera and pertussis toxin which is independent of enhancement of adenylate cyclase activity in the regulation of the immune response. Antibody to pertussis toxin was not detected, which was attributed to inadequate absorption of pertussis toxin.

Adenylate Cyclase Toxin↗

PubMed1993

Polymerase chain reaction and partial sequencing of a British isolate (T637) of feline immunodeficiency virus.

We have used the polymerase chain reaction (PCR) and direct sequencing of the amplified products to obtain information of the molecular nature of an FIV isolate, T637. Cats experimentally infected with T637 have progressed to clinical immunodeficiency disease. The 5' long terminal repeat (LTR), most of the genes coding for internal proteins (GAG) and surface proteins (ENV), and part of the polymerase (POL) gene have been sequenced. The LTR of T637 has 92% nucleic acid identity with the prototype strain, FIV-Petaluma and the Glasgow isolate, FIV-14, 89% with a Swiss isolate, FIVZ2, and 95% with the PPR isolate. Both GAG and POL genes of T637 share extensive homology with Petaluma and PPR. In the ENV gene, T637 has 91% nucleic acid homology with Petaluma and 86% with PPR, and an overall amino acid homology of between 81-87%. For the surface (SU) region of the ENV gene product, T637 has 89% amino acid homology with Petaluma and FIVZ2 and 86% with PPR.

Amino Acid Sequence↗

PubMed1993

Programmed cell death (apoptosis) as a mechanism of cell death in peripheral blood mononuclear cells from cats infected with feline immunodeficiency virus (FIV).

FIV is a lentivirus infection of cats which induces an immunodeficiency syndrome associated with early qualitative defects in antigen-specific T cell function and with late quantitative defects in CD4+ T lymphocytes. We have observed that peripheral blood mononuclear cells (PBMC) from FIV-infected cats have impaired survival in culture. The mechanism of this in vitro dysfunction and depletion is not known. We have proposed that inappropriate induction of programmed cell death (apoptosis) could account for these in vitro defects. Here, we report that PBMC from FIV-infected cats, with impaired T cell blastogenesis and impaired survival in vitro, undergo an active cell death upon culture, which has the morphological and biochemical characteristics of programmed cell death (PCD). Apoptosis occurred in all six asymptomatic FIV-infected cats, and in none of the nine uninfected cats, which were studied. Changes in cell morphology under both light and electron microscopy, and fragmentation of genomic DNA were characteristic for apoptosing cells. Cell death was spontaneous and occurred in the absence of any stimuli, and culture with the T cell mitogen, concanavalin A (Con A), did not significantly enhance cell death. Activation-induced cell death was inhibited, in a dose-dependent manner, by addition to the incubation medium of zinc, which has been shown to inhibit the action of endonuclease responsible for the characteristic fragmentation of DNA. Since apoptosis has recently been implicated in AIDS pathogenesis, FIV infection may prove useful to study this aspect of retroviral, in particular HIV, infection.

Animals↗

PubMed1993

Specific immunological unresponsiveness following active primary responses to proteins in the weaning diet of piglets.

Young piglets weaned onto soya diets frequently develop diarrhoea which may have a dietary and/or immunological component. Piglets abruptly weaned onto soya at 3 weeks of age developed levels of serum IgG anti-soya antibodies almost comparable to those induced by injection with soya protein in adjuvant at 7 weeks. In the piglets primed by feeding, no significant further increase in antibody occurred after subsequent systemic injection. In contrast, secondary responses were observed in age-matched animals, previously primed by injection, and primary responses were obtained in previously naive piglets. The results demonstrate the development of specific unresponsiveness to soya proteins in neonates fed soya, despite the occurrence of an initial vigorous immune response to the fed protein.

Administration, Oral↗

PubMed1993

Humoral immune responses in cats with dermatophytosis.

The IgG and IgM classes of antibodies to a water-soluble antigen preparation derived from Microsporum canis were determined by ELISA in the sera of 79 cats with dermatophytosis confirmed by results of fungal culture, and of 46 specific-pathogen-free-derived, barrier-maintained cats with no previous exposure to dermatophytes. Of the 79 cats with dermatophytosis, the species isolated were: M canis from 72, M gypseum from 6, and Trichophyton mentagrophytes from 1. Concentrations of soluble M canis antigen-specific IgG and IgM were significantly (P < 0.05) higher in the cats with dermatophytosis than in the control cats. The IgG concentration was > or = 2.0 ELISA units/ml ml in 71% of the cats with dermatophytosis and in 9% of the control cats, whereas IgM concentration was > or = 4.0 ELISA units/ml in 38% of the cats with dermatophytosis and in 11% of control cats. There was no significant difference in either IgG or IgM values between the cats with M canis infection and those with other non-M canis dermatophyte infections.

Animals↗

PubMed1992

A monoclonal antibody recognising an epitope associated with pig interleukin-2 receptors.

A monoclonal antibody is described which recognises an epitope associated with a receptor for interleukin-2 (IL-2) on pig lymphocytes. The monoclonal antibody inhibits high affinity binding of radiolabelled recombinant human IL-2 (rhIL-2) by pig lymphoblasts and also non-competitively inhibits both pig-TCGF and rhIL-2 maintained proliferation. By flow cytometry the antigen is apparently not present on freshly isolated blood lymphocytes but is detectable on small cells between 6 and 12 h after activation and on large cells by 24-h. These findings are comparable with those obtained using monoclonal antibodies recognising the 55 kDa alpha chain of the human and mouse IL-2 receptor (p55, TAC) expressed on activated cells in vivo and in vitro. However, the molecular weight of the porcine antigen is between 65 and 70 kDa.

Animals↗

PubMed1992

PROBIT: weighted probit regression analysis for estimation of biological activity.

PROBIT estimates biologic activity by weighted probit regression analysis and comparison with a known standard. The program is intended to speed up calculation of the concentration of cytokines in large numbers of cell supernatants. Data are input from either sequential or free-form text files created with a spreadsheet or text editor. This allows transfer of data from a beta-counter equipped with a suitable terminal without retyping. Results are displayed and/or printed as relative potency (% of standard) and 50% effective dose (ED50).

Cytokines↗