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Biomedical subjects

M R Alderson

Publications and source records attributed to M R Alderson.

At least 55 records · Page 3Linked to original sources
PubMed1993

Fas transduces activation signals in normal human T lymphocytes.

The Fas gene encodes a cell surface molecule that is a member of the the nerve growth factor/tumor necrosis factor receptor family of proteins and can mediate programmed cell death (apoptosis) in certain transformed cell lines. To characterize further the biological function of Fas, particularly with regard to its function in normal cells, a panel of monoclonal antibodies (mAbs) was generated against the extracellular portion of human Fas. Some of these mAbs induced apoptosis in transformed cell lines expressing Fas, but only when immobilized on the culture vessel. One of the new Fas mAbs (M38) was used for studies on normal lymphoid cells and found to stimulate the proliferation of purified human T cells and thymocytes when immobilized on culture wells along with CD3 antibody. T cell proliferation induced by Fas mAb was largely interleukin 2 independent and was demonstrated to be due to a direct effect on the precursor T cell. Thus, the data demonstrate that in addition to a role in the induction of apoptosis in certain transformed cell lines, the Fas protein may also play an important role in the activation and proliferation of normal T cells.

Antibodies, Monoclonal↗
PubMed1993

CD40 expression by human monocytes: regulation by cytokines and activation of monocytes by the ligand for CD40.

CD40 is a member of the tumor necrosis factor (TNF) receptor family of cell surface proteins and was originally described as a B cell restricted antigen. Treatment of primary human monocytes with granulocyte/macrophage colony-stimulating factor (GM-CSF), interleukin 3 (IL-3), or interferon gamma (IFN-gamma) resulted in the induction of CD40 mRNA and enhancement of cell surface protein expression. CD40 was found to mediate monocyte adhesion to cells expressing recombinant CD40 ligand. CD40 ligand-transfected cells provided a potent costimulus for monocyte TNF-alpha and IL-6 production in the presence of GM-CSF, IL-3, or IFN-gamma, and enhanced IL-8 production stimulated by GM-CSF or IL-3. In addition, CD40 ligand-transfected cells acting in the absence of a costimulus induced monocytes to become tumoricidal against a human melanoma cell target. Collectively, these data indicate that CD40 ligand is pleiotropic with potent biological activity on monocytes.

Animals↗
PubMed1993

Interleukin 1 signaling occurs exclusively via the type I receptor.

Two receptors for the proinflammatory cytokine interleukin 1 (IL-1) have been cloned and characterized biochemically. While it has been well established that the type I (80-kDa) IL-1 receptor can mediate responses to IL-1, the function of the type II (60-kDa) IL-1 receptor has been unknown. In this manuscript we describe experiments designed to ask whether the type II receptor is capable of delivering a biological signal. We have examined two types of experimental situation: responses to IL-1 in cells which express predominantly the type II receptor, and responses to IL-1 which have been suggested previously in the literature to be mediated by type II receptors. In both situations we find that the responses instead are mediated via type I receptors. A blocking antibody against the type II receptor never inhibits, and in fact sometimes enhances, the responses. We conclude that a very small number of type I receptors is sufficient to mediate all of the actions of IL-1 which we have examined here and that the function of the type II receptor may not be to transduce signals.

Animals↗
PubMed1993

Molecular and biological characterization of a ligand for CD27 defines a new family of cytokines with homology to tumor necrosis factor.

CD27 is a surface antigen found on T and B cells that has homology to a family of molecules including the receptors for tumor necrosis factor (TNF) and nerve growth factor. A cDNA encoding a ligand for CD27 was isolated by a direct-expression cloning strategy using a fusion protein composed of the extracellular domain of CD27 linked to the constant domain of a human immunoglobulin G1 molecule as a probe. The predicted protein product is a type II transmembrane protein whose gene maps to 19p13 and that shows homology to TNF and the ligand for CD40. Biological characterization indicates that the cloned ligand induces the proliferation of costimulated T cells and enhances the generation of cytolytic T cells.

Amino Acid Sequence↗
PubMed1993

Effect of granulocyte-macrophage colony-stimulating factor and interleukin-3 on interleukin-8 production by human neutrophils and monocytes.

Interleukin-8 (IL-8) is a major neutrophil chemoattractant and functional stimulant that is induced by IL-1, tumor necrosis factor alpha (TNF alpha), and lipopolysaccharide (LPS). We report that recombinant human (rh) granulocyte-macrophage colony-stimulating factor (GM-CSF) and rhIL-3 are also potent inducers of IL-8 messenger RNA (mRNA) accumulation and protein secretion by normal peripheral blood monocytes. Neutrophils produce IL-8 in response to GM-CSF but not to IL-3. In contrast, recombinant human granulocyte-CSF (rhG-CSF), at concentrations as high as 100 ng/mL, does not induce IL-8 in either cell type. rhGM-CSF also induces IL-8 mRNA expression and IL-8 protein in the promonocytic cell line, U-937, whereas rhG-CSF does not. IL-8 secretion by monocytes was stimulated within 2 hours after incubation with rhGM-CSF or rhIL-3. Stimulation of neutrophils with rhGM-CSF resulted in an increase in cell-associated IL-8 at 4 hours. At 24 hours, cell-associated IL-8 levels declined, whereas secreted IL-8 levels increased. In contrast, virtually all IL-8 induced in monocytes appeared as secreted protein. Neither rhGM-CSF nor rhIL-3 induced detectable secretion of IL-1, TNF alpha, or IL-6 protein by monocytes. rhGM-CSF, and to a lesser degree rhIL-3, potently stimulated IL-8 secretion in cultures of heparinized whole blood, whereas rhG-CSF had no significant effect on IL-8 secretion. Induction of IL-8 by GM-CSF may be physiologically important in enhancing the acute inflammatory response.

Blotting, Northern↗
PubMed1993

Molecular characterization of the early activation antigen CD69: a type II membrane glycoprotein related to a family of natural killer cell activation antigens.

CD69 is a disulfide-linked homo-dimer expressed on the surface of activated T cells, B cells, natural killer cells, neutrophils and platelets. Antibody crosslinking of CD69 in the presence of phorbol ester results in cellular activation events including proliferation and the induction of specific genes. Using an expression cloning strategy we have isolated cDNA encoding human CD69 from a CD4+ T cell clone. Transfection of the cDNA clone in CV-1/EBNA cells results in the expression of a covalently linked homodimer. The cDNA insert hybridizes to a 1.7-kb mRNA in phorbol 12-myristate 13-acetate- or phytohemoagglutinin-stimulated human T cells. Using the human clone we have isolated cDNA encoding mouse CD69, which, when expressed in human T cells allowed those cells to respond to anti-mouse CD69 antibodies by secreting interleukin-2 and interferon-gamma. Sequence analysis showed that both mouse and human CD69 are type II membrane glycoproteins related to the NKR-P1 and Ly-49 families of natural killer cell activation molecules.

Amino Acid Sequence↗
PubMed1993

CD40 ligand is a T cell growth factor.

CD40 ligand (CD40L) is a 33-kDa type II membrane glycoprotein induced on T cells upon activation. CD40L has previously been shown to induce proliferation of resting B cells, immunoglobulin (Ig) secretion from B cells cultured with cytokines and cytokine secretion and tumoricidal activity from monocytes. In this report CD40L is shown to be stimulatory for human T cells, inducing CD25 (p55 IL-2R) and CD40L expression on resting peripheral blood T cells, enhanced expression of these molecules and CD69 on CD3-activated cells and secretion of interferon-gamma, tumor necrosis factor-alpha and interleukin (IL)-2 from T cells cultured in the presence of a sub-mitogenic concentration of phytohemagglutinin A (PHA). Furthermore, stimulation with CD40L induces proliferation of CD3- or PHA-activated T cells of blood, tonsillar or thymic origin. A similar proliferative response is observed with CD4+ and CD8+ T cells and this effect is largely IL-2 independent. A soluble construct of the extracellular domain of the CD40L has similar activity to that of membrane-expressed ligand in the induction of T cell surface antigens and proliferation. The results presented here taken together with the various activities ascribed for CD40L on B cells and monocytes demonstrate that CD40L has pleiotropic biological activity for cells of the hemopoietic lineage.

Antibodies, Monoclonal↗
PubMed1993

CD40L: a multi-functional ligand.

B cells can be stimulated to proliferate and differentiate in response to cell-contact dependent signals provided by activated, but not resting, T cells. In the human system, antibodies specific for the surface antigen CD40 induce similar B cell responses. The cloning of a ligand for CD40, and the generation of reagents which can block the interaction of this ligand with its receptor, have demonstrated that the major component of the contact-dependent signal leading to B cell activation is CD40 ligand. Studies of individuals lacking functional CD40 ligand have indicated that signaling through CD40 is essential for immunoglobulin (Ig) heavy chain switching and the production of all isotypes other than IgM. In addition to its activities on B cells, CD40 ligand is stimulatory for cells of monocyte and T lineages suggesting a pleiotropic role for CD40 ligand in vivo.

Animals↗
PubMed1993

Induction by interleukin-7 of lymphokine-activated killer activity in lymphocytes from autologous and syngeneic marrow transplant recipients before and after systemic interleukin-2 therapy.

Therapy with recombinant lymphokines after autologous bone marrow transplantation (ABMT) is being explored as a way to prevent relapse. Lymphokine therapy may exert an antitumor effect through a variety of mechanisms, including the induction of lymphokine-activated killer (LAK) cell cytotoxicity. We tested the ability of interleukin-7 (IL-7) to induce LAK cytotoxicity in peripheral blood mononuclear cells (PBMC) from healthy subjects and from patients early after ABMT. LAK activity was defined as lysis of Daudi by PBMC after incubation with IL-7 at 10 to 100 ng/mL or IL-2 at 1000 U/mL. PBMC from four healthy subjects were cultured with either IL-7 or IL-2. IL-7 induced LAK activity in two of the four, whereas IL-2 induced LAK activity in all four. The median percent lysis (effector-to-target ratio [E:T] 40:1) with IL-7 (23%) was lower than with IL-2 (67%). PBMC were obtained from 15 patients 27 to 84 days after autologous (n = 13) or syngeneic (n = 2) bone marrow transplantation (BMT) and tested for IL-7-induced LAK activity. Eleven exhibited significant activity (10% to 77% lysis at E:T 40:1). In contrast to the results in PBMC from normal subjects, in PBMC from ABMT patients IL-7 induced LAK activity of a magnitude similar to that induced by IL-2. Studies were also performed on PBMC from eight patients who had received IL-2 after ABMT (3.0 x 10(6) U/m2/d) for 4 days by continuous intravenous (IV) infusion. In seven of the eight patients, IL-7 induced significant LAK activity, which was higher than that seen in PBMC from ABMT patients who had not received IL-2. Thus, IL-7 reproducibly induced significant LAK activity in cells obtained early after autologous or syngeneic BMT. Indeed, such LAK activity was comparable quantitatively to that induced by IL-2. Finally, IL-7 induced an even greater LAK activity in vitro in PBMC obtained after ABMT and preactivated in vivo by IL-2 therapy. The results suggest that IL-7 may have a potential immunotherapeutic role, alone or with IL-2, after ABMT.

Adult↗
PubMed1992

Regulation of human monocyte cell-surface and soluble CD23 (Fc epsilon RII) by granulocyte-macrophage colony-stimulating factor and IL-3.

We have investigated the regulation of expression of cell-surface and soluble CD23 (sCD23) by purified human peripheral blood monocytes and in cultures of human whole blood. IL-3, IL-4, and GM-CSF were found to markedly enhance the expression of CD23 on the surface of elutriated monocytes and to increase levels of sCD23 in monocyte-culture supernatants. The induction of CD23 expression by monocytes was confirmed at the mRNA level by Northern blot analysis. The ability of GM-CSF, IL-3, or IL-4 to induce cell-surface CD23 on monocytes was inhibited by specific neutralizing antibodies to the corresponding cytokine. IL-3 and GM-CSF induced maximal surface CD23 expression on monocytes by 24 to 48 h, followed by a slight decline at 72 and 96 h. In contrast, IL-4 induced a progressive increase in monocyte CD23 expression that reached a maximum at approximately 72 h. IL-4, GM-CSF, and IFN-gamma increased both surface and soluble CD23 expression by the monocytic cell line U937, whereas IL-3 had no effect. The plasma from fresh human whole blood or nonstimulated whole blood cultured for 24 to 48 h contained detectable sCD23, and addition of IL-3, IL-4, or GM-CSF to these cultures resulted in increased levels of this molecule. Two-color flow cytometry revealed that IL-3, but not GM-CSF, also enhanced CD23 expression by B cells enriched from PBMC, although the effect of IL-3 was weak in comparison with that of IL-4. These findings may have important implications for the in vivo therapeutic use of these cytokines.

Antigens, Differentiation, B-Lymphocyte↗
PubMed1991

Induction of macrophage inflammatory protein-1 beta gene expression in human monocytes by lipopolysaccharide and IL-7.

HuMIP-1 beta is a member of a gene family of inflammatory cytokines whose expression is induced by proinflammatory and mitogenic stimuli. The gene was rapidly induced in human peripheral blood monocytes by either LPS or IL-7. However, IL-7 was unable to induce HuMIP-1 beta mRNA in peripheral blood T cells. The induction of HuMIP-1 beta mRNA in monocytes by IL-7 or LPS was inhibited by IL-4. The 5'-regulatory region of the HuMIP-1 beta gene was cloned and sequenced. An analysis of this sequence revealed three consensus-binding sites for the nuclear factor PU.1 and three potential glucocorticoid response elements. However, the HuMIP-1 beta gene appears to be unresponsive to dexamethasone. Also present in the 5'-regulatory region was a LPS-responsive element located within 455 bp 5' to the start of transcription.

Base Sequence↗
PubMed1991

Interleukin 7 induces cytokine secretion and tumoricidal activity by human peripheral blood monocytes.

Peripheral blood monocytes can be induced by stimuli such as bacterial lipopolysaccharide (LPS) to secrete an array of cytokines. We have studied the effects of interleukin 7 (IL-7) on human peripheral blood mononuclear cells (PBMC) and found that IL-7 is a relatively potent inducer of IL-6 secretion IL-6 protein levels were determined either by the B9 hybridoma growth factor assay or by enzyme-linked immunosorbent assay, and mRNA for IL-6 was analyzed by Northern hybridization. Detailed examination revealed that, among PBMC, monocytes, rather than lymphocytes, were secreting IL-6 in response to IL-7. In contrast to the low concentrations of IL-7 required to stimulate T cell growth and differentiation (as low as 0.1 ng/ml), relatively high concentrations of IL-7 were necessary to induce IL-6 secretion by monocytes (at least 10 ng/ml). An optimal concentration of IL-7 (100 ng/ml) induced monocytes to secrete 10-fold more IL-6 than an optimal concentration of IL-1 beta (10 ng/ml), and almost as much as LPS. However, significantly more IL-7 than IL-1 beta was required to induce detectable levels of IL-6. The kinetics of IL-6 secretion by monocytes were identical in response to IL-7, IL-1 beta, or LPS, with IL-6 protein detectable in culture supernatants as early as 2 h after the initiation of culture. IL-4 was found to markedly inhibit the ability of IL-7 or LPS to induce IL-6 mRNA and IL-6 secretion. In addition to promoting IL-6 production, IL-7 induced the secretion of immunoreactive IL-1 alpha, IL-1 beta, and tumor necrosis factor alpha (TNF-alpha) by monocytes. IL-7 also induced monocyte/macrophage tumoricidal activity against a human melanoma cell target, an activity that may be related to the secretion of IL-1 alpha, IL-1 beta, and TNF-alpha. Finally, we used a whole blood culture system as a bridge to in vivo analysis to demonstrate that IL-7 induces cytokine secretion in the absence of culture medium, fetal calf serum, and adherence to plastic. Our data suggest that IL-7, in addition to regulating lymphocyte growth and differentiation, has potent effects on cells of the monocytic lineage. Thus, IL-7 may be an important mediator in inflammation and in the macrophage immune response to tumors.

Cell Survival↗
PubMed1990

Interleukin 7 enhances cytolytic T lymphocyte generation and induces lymphokine-activated killer cells from human peripheral blood.

The effects of purified recombinant interleukin 7 (IL-7) on the generation of cytolytic T lymphocytes (CTL) in mixed lymphocyte culture (MLC) and on the induction of lymphokine-activated killer (LAK) cells in autologous cultures of human peripheral blood mononuclear cells were investigated. IL-7 was found to induce the generation of both CTL and LAK cells in bulk cultures. The appearance of peak CTL activity in MLC established with exogenous IL-7 was delayed in comparison with replicate cultures containing exogenous IL-2, but both cytokines stimulated quantitatively similar levels of antigen-specific lytic activity. An IL-2-neutralizing antiserum inhibited substantially, but not completely, the effect of IL-7 on CTL generation, implying the existence of both an indirect component of IL-7 activity via IL-2 utilization, as well as an IL-2-independent component. Cell surface phenotypic analysis of IL-2- or IL-7-generated CTL effector cells revealed that CD8+ cells were responsible for the vast majority of lytic activity. Limiting dilution analysis (LDA) revealed that essentially identical frequencies of CTL precursors (CTL-P) were capable of clonal expansion and/or differentiation in the presence of exogenous IL-2, IL-4, or IL-7, supporting the concept that all three of these cytokines are capable of exerting a major influence on T cell growth and differentiation. Approximately half of the CTL-P that responded in IL-7-supplemented LDA cultures did so in an IL-2-independent manner. IL-7 stimulated the development of LAK cells in autologous bulk cultures, but only weakly in comparison with IL-2. In contrast to its effects on CTL generation, the induction of LAK cells by IL-7 was virtually independent of IL-2. LAK cells induced by IL-7, like those induced by IL-2, were phenotypically heterogeneous and included CD8+, CD56+, and gamma/delta+ cells. Limiting dilution analysis indicated that IL-2 stimulated fivefold more LAK-P than IL-7 and 220-fold more than IL-4. Collectively, these data suggest that IL-7 has potent regulatory effects on human cytolytic cell populations and, either alone or in combination with other cytokines, could be important for the in vitro expansion of cells for adoptive immunotherapy.

Antigens, CD↗
PubMed1989

Recombinant human interleukin 6 (B cell stimulatory factor 2) enhances immunoglobulin secretion by single murine hapten-specific B cells in the absence of cell division.

We have assessed the role of recombinant human IL-6 (r-hu-IL-6) in promotion of early activation, proliferation, and immunoglobulin (Ig) secretion amongst single hapten-specific murine splenic B cells in vitro. It was found that r-hu-IL-6 acting alone was able to induce early B cell activation in a proportion of B cells, as measured by a significant increase in cell diameter within 24 h. An enhanced effect was seen in the concomitant presence of a 'T-independent' antigen. None of the B cells activated by r-hu-IL-6 appeared to divide, as the frequencies of proliferating clones induced by either medium alone or antigen alone were virtually identical whether r-hu-IL-6 was present or absent. However, assay of the culture supernatants for the presence of Ig by ELISA revealed that r-hu-IL-6 effected a significant 2-fold increase in the frequency of B cells secreting Ig. Thus, the prime effect of r-hu-IL-6 appears to be to recruit more precursor B cells into Ig secretion, rather than to promote proliferation or to enhance the amount of Ig secreted by pre-committed but non-cycling B cells. Delayed addition experiments showed that r-hu-IL-6 enhanced Ig secretion late in the activation pathway. Kinetics studies demonstrated detectable Ig secretion as early as day 2, and when taken with its apparent ability to induce early activation, these findings suggest that IL-6 is not exclusively a late-acting interleukin. Studies with size fractionated hapten-specific B cells showed the larger B cells to be preferentially responsive to r-hu-IL-6.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Recombinant T cell replacing factor (interleukin 5) acts with antigen to promote the growth and differentiation of single hapten-specific B lymphocytes.

The role of murine recombinant T cell replacing factor (rTRF) (interleukin 5) in the early activation, proliferation, and antibody-forming cell (AFC) clone formation of single fluorescein (FLU)-specific B cells was examined in vitro. FLU-specific B cells were selected by their adherence to FLU-gelatin and then cultured in 10-microliters wells with or without rTRF in the presence or absence of the T-independent antigen FLU-polymerized flagellin (FLU-POL). rTRF acting alone was unable to induce early B cell activation as assessed by significant cell enlargement after 24 hr in culture. When acting in the presence of FLU-POL, however, a greater number of B cells were induced to enlarge than with FLU-POL alone. When FLU-specific B cells were cultured in the presence of FLU-POL, the addition of rTRF markedly increased the frequencies of both proliferating clones and AFC clones above that induced by FLU-POL alone. Furthermore, in the presence of FLU-POL, the activity of rTRF was comparable to that seen with the mixture of B cell growth and differentiation factors contained within the supernatant from concanavalin A-stimulated EL4 cells. However, rTRF exerted little activity when acting alone in contrast to the medium conditioned by concanavalin A-stimulated EL4 cells which showed some activity in the absence of FLU-POL. rTRF acting with FLU-POL also promoted AFC clone development among single B cells stimulated in the presence of 3T3 fibroblast filler cells. Thus rTRF can be added to the list of B cell active factors (including recombinant murine interleukin 1 and recombinant human interleukin 2) that act in the concomitant presence of antigen to induce both growth and differentiation among single hapten-specific murine splenic B cells. This stands in contrast to the activity seen with interleukin 4 (formerly termed B cell stimulatory factor 1) which acts to promote early activation and proliferation but not IgM secretion.

Animals↗
PubMed1987

Effects of antigens and lymphokines on early activation of single hapten-specific B lymphocytes.

An assay was developed to monitor early activation of single fluorescein-specific B cells obtained from the spleens of nonimmunized adult mice by prefractionation on hapten gelatin. Early activation was assessed as a significant increase in the diameter of individual B cells after 24 hr in vitro. Significant enlargement of the single B cells was induced within 24 hr by either T-independent antigens acting alone or a crude source of B cell growth and differentiation factors (EL-BGDF-pik) acting alone. In contrast, T-dependent antigens acting alone were ineffective. When selected T-independent antigens and EL-BGDF-pik acted together, a greater number of B cells were induced to enlarge. B cell stimulatory factor 1 (BSF 1) behaved in a similar manner as EL-BGDF-pik, inducing early B cell enlargement both in the absence and more so in the presence of antigen. Both EL-BGDF-pik and BSF 1 enhanced the survival of single hapten-specific B cells during the 24-hr period. Interleukin 1 was unable to cause B cell enlargement when acting alone, although it was able to augment B cell enlargement induced by antigen. Interleukin 2 did not induce cell enlargement in either the presence or absence of antigen. Activation was demonstrated among cells of all sizes, regardless of the stimulus, although a greater response was demonstrated amongst the larger cell population. The addition of 3T3 filler cells enhanced early B cell activation and cell survival during the 24-hr period. The 24-hr B cell enlargement assay as applied to isolated single cells provides an unequivocal approach to the analysis of early B cell activation, adding a further parameter for the dissection of the precise roles of antigen and the various factors in the B cell differentiation pathway.

Animals↗
PubMed1987

Single cell studies on the role of B-cell stimulatory factor 1 in B-cell activation.

The role of the T-cell-derived lymphokine B-cell stimulatory factor 1 (BSF-1) in the early activation, proliferation, and antibody-forming cell (AFC) clone formation of single fluorescein-specific B lymphocytes isolated from normal mouse spleens by hapten-gelatin adherence has been studied in vitro. BSF-1 acting alone induced early B-cell activation, as assessed by a significant increase in cell diameter of single B cells cultured for 24 hr. A small but significant number of these B cells formed proliferating clones, some of which secreted antibody. When acting with the specific antigen fluorescein-polymerized flagellin, BSF-1 augmented early cell enlargement and markedly enhanced proliferation, but it did not increase the frequency of AFC clones stimulated by fluorescein-polymerized flagellin alone. The further addition of recombinant murine interleukin 1 (IL-1) marginally enhanced proliferation caused by antigen plus BSF-1. No synergy was observed between BSF-1 and IL-1 for antibody formation. In the presence of fibroblast filler cells, BSF-1 substantially inhibited AFC clone development achieved by antigen plus IL-1. BSF-1 was also found to be inhibitory to AFC clone development stimulated by specific antigen acting with either recombinant human interleukin 2 (IL-2) or with IL-2 plus IL-1, both in the presence or absence of filler cells. The results suggest that BSF-1 plays a complex role in the regulation of the B-cell activation pathway by enhancing early activation and antigen-specific proliferation as well as inhibiting the effects of other B-cell factors on antibody formation. BSF-1 is the only cytokine so far tested in the single B-cell system that acts with antigen to promote proliferation without concomitant antibody production.

Animals↗