Biomedical subjects

H Zola

Publications and source records attributed to H Zola.

At least 73 records · Page 4Linked to original sources

PubMed1995

The effect of IL-4 on antigen expression: a comparison of fluorescence intensity and antigen density.

We have compared the membrane expression of MHC class II antigen and Ig on murine splenic B lymphocytes by two flow cytometric methods. The first method, commonly used, determined the mean immunofluorescence intensity; the second measured antigen density as a ratio of mean fluorescence intensity to cell volume. The data generated by the two methods allow for different interpretations of antigen density in comparison to antigen expression. By mean fluorescence intensity, activated B lymphocytes expressed more MHC class II antigen and membrane Ig than did resting B cells. However, resting B cells had the same density of MHC class II antigen and membrane Ig as activated B cells. Next we examined the effect of IL-4 on antigen expression by the two methods. We found that although IL-4 increased the mean fluorescence intensity and density of MHC class II and membrane Ig on each B cell population, the density of expression of membrane Ig tended to be greater on the resting B cells than on the activated, while the density of MHC class II antigen was essentially the same on each of the IL-4 treated B cell populations.

Animals↗

PubMed1995

Cell surface determinants on typhoid-specific B cells isolated from peripheral blood after oral Ty21a typhoid vaccination in humans.

Antigen specific B cells (ASC) that circulate after oral immunization with the typhoid vaccine Ty21a display cell surface determinants which are potentially involved in B cell differentiation and homing to mucosal sites. These ASC were isolated from peripheral blood after oral Ty21a, and dual labelled for binding of typhoid antigen and expression of various cell surface determinants: alpha 4 integrin (CD49d), CD45RO, CD45RA, L-selectin, CD44 and CD11a. Of particular interest was the finding of CD45RO expression on ASC. A comparison of cell surface determinants on typhoid-specific cells was also made following binding to high endothelial venules on peripheral and mesenteric lymph nodes, and venules in the lamina propria of the small intestine. Generally more typhoid ASC bound to mesenteric compared with peripheral lymph node. More ASC expressing CD45RO and alpha 4 integrin (CD49d) were bound to mesenteric lymph node and small intestine than to peripheral lymph node. When expressed as a fraction of total ASC, the difference was statistically significant only for CD45RO binding to small intestine versus peripheral lymph node. No differences in expression of other homing markers on bound ASC were seen.

Administration, Oral↗

PubMed1995

Cytokine receptor expression in human lymphoid tissue: analysis by fluorescence microscopy.

A highly-sensitive flourescence method, capable of detecting cytokine receptors present at low concentrations (around 100 molecules per cell) by flow cytometry, was adapted for use on tissue sections. This method was used to examine the expression of several cytokine receptors in lymphoid tissues. IL-2 receptors were distributed broadly, with higher concentrations in T cell areas. IL-1 receptor Type 1 was detected in T cell areas and in the follicular mantle, and was strongly expressed on vascular endothelium. IL-6 receptor was found at very low concentration, both within and outside germinal centres. The gp 130 molecule, which is involved in the functional receptor complex for IL-6 and several other cytokines, was present at higher concentrations, particularly in the germinal centre. Analysis of receptor expression in secondary lymphoid tissue provides evidence bearing on the physiological roles of cytokines, as these tissues contain cells at various stages of physiological activation located in well-defined functional zones.

Antibodies, Monoclonal↗

PubMed1995

Expression of cytokine receptors by human cord blood lymphocytes: comparison with adult blood lymphocytes.

The expression of receptors for several cytokines (IL 2, IL-4, IL-6, IL-7, tumor necrosis factor, and interferon-gamma) was examined in human cord blood cells in comparison with adult blood cells. A previously described high sensitivity immunofluorescence procedure was used to render the low levels of receptor measurable. Cord blood lymphocytes expressed measurable levels of most cytokine receptors, but expression tended to be lower than in adult blood cells. Examination of different lymphocyte subpopulations revealed a complex pattern with some cell types expressing particular receptors equivalent to adult levels. Cord and adult blood monocytes expressed similar cytokine receptor profiles. Receptor expression in cord lymphocytes could be regulated by activation. The results provide indications as to the relative activities of different cytokines in the development of the immune system in the neonate.

Adult↗

PubMed1995

Expression of functional insulin-like growth factor-1 receptor on lymphoid cell subsets of rats.

It has become evident that insulin-like growth factor-1 (IGF-1) acts as a growth factor for immune cells, yet the precise regulatory role of IGF-1 in the immune system is unknown. The aim of this study was to examine the distribution of IGF-1 receptors on rat lymphoid cells. A flow cytometric method was used, with a biotinylated and functionally active IGF-1 analogue, namely des(1-3)IGF-1, which binds well to IGF-1 receptor but poorly to IGF binding proteins, followed by phycoerythrin-conjugated streptavidin (PE-SA) staining. Our results showed that IGF-1 receptors were readily detectable on a wide variety of the immune cells, including T cells, B cells and monocytes, but the binding capacity for IGF-1 was monocytes > B cells > T cells, as determined by titration experiments. Furthermore, the level of expression on resting CD4+ T lymphocytes was greater than on CD8+ cells, and the concentration of biotin-des(1-3)IGF-1 required to demonstrate the binding to IGF-1 receptor on CD8+ cells (68 nmol/l) was 200-fold higher than for CD4+ cells (0.34 nmol/l), indicating that most of the IGF-1 receptor on CD8+ cells represented lower affinity sites. The level of IGF-1 receptor expression was increased several-fold after concanavalin A stimulation on both CD4+ and CD8+ T-cell subsets. Kinetic analysis of the expression of IGF-1 receptor and its association with interleukin-2 receptors (IL-2R) following activation showed a similar pattern, with no significant differences in the ratio of IGF-1 receptor: IL-2R per cell during the 3 days of cell culture. Our studies suggest that biological activities of IGF-1 include direct stimulation of immune cells, and that expression of IGF-1 receptor may have a role in regulation of T-cell function.

Animals↗

PubMed1995

Cytokine receptor expression by solid tumours.

The expression of cytokine receptors by a variety of solid tumour tissues was examined, using an immunofluorescence procedure optimized for sensitivity. Several cytokines generally considered as relevant only to the immune and haematopoietic systems were shown to be expressed by solid tumours. For example, breast carcinoma frequently expressed both chains of the IL-3 receptor, the beta chain of the IL-2 receptor, the TNF type two receptor, the signal-transducing chain CD130, and c-kit. The broad expression of cytokine receptors suggests that the receptor profile of individual tumours should be determined before the application of therapy that involves the administration of cytokines.

Breast Neoplasms↗

PubMed1994

Dual staining of lymphocyte membrane antigens with colloidal gold and biotinylated horseradish peroxidase.

Double immunoenzymatic labelling procedures for the localization of antigens on cells in tissue sections using horseradish peroxidase (HRP) and alkaline phosphatase have been described previously, but mainly for detecting antigens on different cells. With this type of staining when two antigens are present on the same cell, an optimal colour combination that shows a high contrast between the basic colour of each enzyme substrate product is difficult to achieve and the interpretation of their mixed colour intermediate is subjective. We present a method for the simultaneous demonstration of two antigens on the same cell. The method can be used to label either single cells in suspension, or cells in paraffin fixed tissue, using a combination of a particulate label, colloidal immunogold-silver, and an enzymatic label HRP-DAB. The method is easy to perform and utilises commercially available staining kits.

Antigens, CD↗

PubMed1994

Cytokine receptor expression in leukaemic cells.

Cytokines have been postulated to play important roles in tumour biology and in the host response to tumours, and a number of therapeutic modalities involving cytokines have been proposed. If patients are to be treated with cytokines, or cytokine inhibitors, it will be important to determine the potential for direct action of the cytokine on the tumour cells. In this study, a high-sensitivity immunofluorescence technique is used to determine the expression of a number of cytokine receptors on a total of 115 leukaemic samples. The results show that many leukaemic samples express low levels of cytokine receptors, and that malignancies of a particular type are heterogeneous with respect to receptor expression. In vitro culture experiments show, as expected, that receptor expression is a necessary but not sufficient requirement for responsiveness to cytokines. The cytokine receptor phenotype may provide useful additional clinical and prognostic information, and should be determined particularly for patients undergoing treatment with cytokines or cytokine inhibitors.

Acute Disease↗

PubMed1994

Rapid detection of immunoglobulin gene somatic hypermutation using heteroduplex formation.

In order to identify somatic hypermutation of rearranged human immunoglobulin genes, we have examined heteroduplex formation between cloned VH6 genes. In test systems, the presence of five or more point mutations could be detected by examining the formation of heteroduplexes between a known germline VH6 gene and other sequenced genes using polyacrylamide gel electrophoresis. If a mutated sequence was used, then the presence of two or more mutations could be detected. The method was used for rapid screening of VH6-D-JH rearrangements for the presence of point mutations before sequencing, and to distinguish between different highly mutated sequences, allowing clones containing the same rearrangement to be identified indirectly.

Base Sequence↗

PubMed1994

A comparison of the sensitivity of immunoperoxidase staining methods with high-sensitivity fluorescence flow cytometry-antibody quantitation on the cell surface.

Surface molecules present in low copy numbers can be detected with high-sensitivity fluorescence flow cytometry. Many cells previously thought not to express certain molecules on their surface can now be shown to have these molecules in very low copy numbers by high-sensitivity fluorescent cytometric methods. Detection of molecules by immunoperoxidase staining methods has not previously been compared with high-sensitivity flow cytometry techniques. Computerized video image analysis (VIA) is a method that allows measurement of area and density of the immunostain chromogen reaction product in a standardized fashion analogous to flow cytometry. In this study, we compared immunoperoxidase reaction products measured by VIA methods with high-sensitivity flow cytometric measurements for cells with 10,000 down to 50 antibody molecules bound to their surfaces. Detection of 100-200 surface molecules was possible with heavy metal-enhanced immunoperoxidase methods, whereas standard immunoperoxidase methods were not as sensitive. The sensitivity of the nickel-enhanced immunoperoxidase staining method was confirmed for detection of an epitope (Tac-IL2 receptor alpha-chain) present in low numbers on the surface of peripheral blood lymphocytes.

Antibodies, Monoclonal↗

PubMed1993

Expression of the Fc receptor for IgG (Fc gamma RII/CDw32) by human circulating T and B lymphocytes.

Tissue-specific isoforms of the human FcR for IgG Fc gamma RII (CDw32) have previously been described by using mAb. These mAb were shown to exhibit different patterns of reactivity with lymphocytes. Among human PBL, Fc gamma RII has been detected on B cells but not T cells when assessed by flow cytometry and microscopy with the use of mAb KB61 and 41H16. Although KB61 and 41H16 were found to react with B cells, the mAb IV.3, CIKM5, and 2E1 did not react with any PBL subset. In this study, we show that KB61 and 41H16 react strongly with the majority (93-96%) of B cells (CD20+), and weakly with a proportion (18-42%) of T cells (CD3+), including 10 to 14% of CD4+ and 27 to 69% of CD8+ cells. In addition, mRNA for Fc gamma RII was detected in purified CD3+CD8high+ lymphocytes by polymerase chain reaction. KB61 and 41H16 also reacted with a majority of CD3-CD16/CD56+ cells, and CD3-CD20- cells. These findings indicate that a subset of T cells and non-T/non-B cells express Fc gamma RII, and are of interest in the light of previous studies which postulate that human Fc gamma R+ cells and Fc gamma RII+ murine T cells suppress the B cell immune response.

Animals↗

PubMed1993

Direct demonstration of membrane IL-1 alpha on the surface of circulating B lymphocytes and monocytes.

A mAb against IL-1 alpha has been used in a high sensitivity immunofluorescence procedure to demonstrate directly the presence of surface IL-1 alpha on human monocytes and B cells from blood and tonsil. IL-1 alpha could be detected on monocytes and a subset (around 30%) of B cells without any in vitro activation. Although the subset of B cells expressing IL-1 alpha was distinct (i.e., the distribution of fluorescence intensity was bimodal), no correlation was seen with any other B cell subset marker in blood. In tonsil, IL-1 alpha was expressed by cells with a phenotype that suggested that they represented a subset of mantle zone B cells. Activation of tonsil B cells with anti-IgM, IL-4, IL-2, and mixtures of these reagents led to small increases in IL-1 alpha expression, as did activation of unfractionated tonsil cells with PHA.

Animals↗

PubMed1993

Coexpression of IL-2 receptor p55 and p75 by circulating blood lymphocytes.

By using high-sensitivity fluorescence and flow cytometry, it is possible to show that 30-40% of lymphocytes from PBL express the p55 chain of the IL-2 receptor, whereas the p75 chain is expressed at low concentrations on most lymphocytes without in vitro activation. The availability of a second fluorochrome capable of high sensitivity allows simultaneous analysis of p55 and p75, albeit with some sacrifice in sensitivity. Two-color analysis shows that a small proportion of cells (1-6%) coexpress measurable concentrations of both chains of the IL-2 receptor, and three-color studies show that these cells are predominantly CD4-positive T cells and express the CD45R0 isoform of the leucocyte-common antigen, i.e., have the phenotype of activated helper T cells. These cells may be a useful indicator of immune activation.

Antibodies, Monoclonal↗

PubMed1993

Expression of IL-4 receptor on human T and B lymphocytes.

The expression of the interleukin-4 receptor on human blood and tonsil lymphocytes has been studied using a monoclonal antibody and high-sensitivity immunofluorescence flow cytometry. While no receptor expression could be detected on circulating or tonsil T cells, a subset of B cells was shown to express the receptor. The IL-4R-positive B cells in tonsil had a phenotype suggesting that they included both germinal centre B cells and B cells outside the germinal centre. The subset of B cells in the blood that expressed the receptor included CD23-positive B cells. Activation of tonsil B cells using anti-IgM, IL-4, IL-2, or combinations of these reagents led to increases in IL-4R expression, but these changes were small compared to changes in the expression of IL-2R p55 (CD25), a known marker of activation. Similarly, activation of T cells led to low-level expression of IL-4R, with IL-4 itself up-regulating IL-4R, especially in CD4 cells. The majority of chronic lymphocytic leukaemia samples were positive for IL-4R expression, whilst most other leukemic samples were negative.

ADP-ribosyl Cyclase↗

PubMed1993

Expression of CD45 isoforms in chronic B-cell leukaemias.

CD45 isoform expression was studied in 204 cases of B-cell lymphoproliferative disease, including 162 chronic lymphocytic leukaemia (CLL). In almost half the samples tested, CD45R0 was co-expressed with CD45RA, unlike normal B-cells, which express only CD45RA, except at terminal stages of differentiation. In a small number of cases CD45R0 was the dominant isoform expressed. No correlation could be discerned either with Binet or RAI staging in CLL or with disease type CLL, prolymphocytic leukaemia (PLL) or hairy cell leukaemia (HCL). Comparison of CD45 isoform expression with other markers showed no correlation with apparent maturational status of the cells involved.

Antigens, CD↗

PubMed1993

An organ fragment culture model to study lymphocyte activation in human lymphoid tissue.

We have established and evaluated an organ fragment culture model for the study of human lymphocyte activation and differentiation. Small fragments of tonsillar tissue were cultured on Gelfoam for periods of up to 7 days. Monoclonal antibody in the medium was able to diffuse into the tissue, as demonstrated by subsequent detection of antibody-coated cells. Phytohaemagglutinin added to the culture medium caused activation of T and B cells, as indicated by changes in expression of a number of markers. Antibody against human IgM (added as a F(ab')2 fragment) together with IL-4 caused B cell activation, detectable by an increased expression of CD23 and other markers. Cell viability fell gradually in culture, but useful data could nevertheless be obtained from culture periods up to 7 days. The organ fragment culture provides a model for the study of T and B cell activation which maintains, at least in part, the intercellular interactions and the native microenvironment of lymphoid tissue.

Antibodies, Monoclonal↗

PubMed1993

Expression of membrane receptor for tumour necrosis factor on human blood lymphocytes.

Using a monoclonal antibody against the human p75 tumour necrosis factor receptor (TNFR-I) combined with a high-sensitivity immunofluorescence flow cytometric procedure, a proportion of peripheral blood lymphocytes can be shown to express TNFR-I constitutively. Approximately 50% of peripheral blood lymphocytes consisting mostly of CD4 cells and including most CD45R0-positive cells, express TNFR-I. Receptor expression is increased by a variety of activation signals. Only a minority (up to 30%) of tonsil B cells express measurable levels of TNFR-I. The tonsil B cells which express TNFR-I include both cells with a germinal centre cell phenotype and cells with the phenotype of the follicular mantle zone. Activation of B cells with anti-immunoglobulin, alone or in combination with interleukin-4 or interleukin-2, increases receptor expression, particularly in cells with the phenotype of mantle zone cells. The functional significance of constitutive expression of TNFR by blood and tissue lymphocytes is discussed.

Antibodies, Monoclonal↗

PubMed1992

Comparative evaluation of CD5 B cells in patients with rheumatoid arthritis and essential mixed cryoimmunoglobulinemia using FACS analyzer and FACSCAN flow cytometer.

In this study, using both a FACSCAN flow cytometer and the FACS analyzer, and two-color fluorescence, CD5 B cells have been enumerated in the peripheral blood (PB) of normal controls (NC) and patients with rheumatoid arthritis (RA) or essential mixed cryoimmunoglobulinemia (EMC). Using a FACSCAN flow cytometer, no significant difference was observed between the percentage of CD5 B cells in the NC (21.24% +/- 3.71) and RA (24.83% +/- 3.85), or EMC (21.03% +/- 6.38). Using the FACS analyzer, however, there was significant difference between the NC (9.14% +/- 3.82) and EMC (2.84% +/- 2.58) (p less than 0.05), but no significant difference between the NC and RA (7.17% +/- 2.55). Further analysis of 10,000 B cells selected by live gating showed that there were three populations of B cells, with the majority of B cells unstained, a small number of cells brightly stained for CD5 in the NC (3.78% +/- 1.09) or RA (2.80% +/- 0.96), and about 6.45% to 9.43% with intermediate staining in patients with RA and the NC, respectively. In addition, serum low molecular weight IgM (LMW IgM) from patients with RA was detected by an enhanced chemiluminescence detection system combined with a modified immunoblot technique. A significant linear correlation was observed between the percentage of CD5 B cells and the height of LMW IgM peak (r = 0.69, p less than 0.05).

Antigens, CD↗