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Biomedical subjects

M R Alderson

Publications and source records attributed to M R Alderson.

At least 19 recordsLinked to original sources
PubMed1995

A metalloprotease inhibitor blocks shedding of the IL-6 receptor and the p60 TNF receptor.

Many cytokines and soluble cytokine receptors are generated by limited proteolysis of membrane-bound precursors. We have examined the ability of the recently described metalloprotease inhibitor, TNF-alpha protease inhibitor (TAPI), and other protease inhibitors to modulate shedding. The membrane-bound forms of the ligands TNF-alpha and CSF-1, the p60 TNFR and the IL-6R, were expressed in COS-7 cells. As expected, TAPI blocked the spontaneous and PMA-induced release of TNF-alpha from transfected cells. Interestingly, TAPI also inhibited the release of soluble forms of p60 TNFR and IL-6R in COS-7 cells. However, the processing of CSF-1, which also requires proteolytic cleavage of a membrane protein, was not affected. The ability of TAPI to inhibit shedding was unique, since several other classes of protease inhibitors, including three other metalloprotease inhibitors, did not inhibit shedding of IL-6R. To determine whether TAPI would prevent shedding under more physiologic conditions, we demonstrated that TAPI was able to prevent unstimulated and PMA-induced release of the soluble forms of TNF-alpha, p60 TNFR, and IL-6R from the monocytic cell line, THP-1, and from human peripheral blood monocytes. In addition, TAPI was able to inhibit LPS-induced shedding of the p60 TNFR and TNF-alpha from monocytes. In summary, our results indicate that a metalloprotease or group of related metalloproteases is responsible for the proteolytic cleavage of several cell surface proteins.

Antigens, CD↗
PubMed1995

Fas ligand mediates activation-induced cell death in human T lymphocytes.

A significant proportion of previously activated human T cells undergo apoptosis when triggered through the CD3/T cell receptor complex, a process termed activation-induced cell death (AICD). Ligation of Fas on activated T cells by either Fas antibodies or recombinant human Fas-ligand (Fas-L) also results in cytolysis. We demonstrate that these two pathways of apoptosis are causally related. Stimulation of previously activated T cells resulted in the expression of Fas-L mRNA and lysis of Fas-positive target cells. Fas-L antagonists inhibited AICD of T cell clones and staphylococcus enterotoxin B (SEB)-specific T cell lines. The data indicate AICD in previously stimulated T cells is mediated by Fas/Fas-L interactions.

Antigens, Surface↗
PubMed1995

B-cell stimulation.

Recent studies have identified CD40 ligand (CD40L) as the critical membrane-expressed molecule responsible for T cell dependent B-cell activation. CD40L co-operates with various cytokines to induce B-cell activation, proliferation, and immunoglobulin isotype switching. Some antigens, however, can also stimulate B-cell activation and isotype switching in the absence of CD40L or T cells. Recent studies have suggested that cytokines derived from non-T cells, such as natural killer cells, macrophages and mast cells, are responsible for isotype switching in T cell independent responses.

Animals↗
PubMed1994

Recombinant CD40 ligand exerts potent biologic effects on T cells.

Murine and human CD40 ligand (CD40L) were recently cloned, expressed, and shown to possess potent activity on human and murine B cells, including stimulation of proliferation and Ig secretion in the presence of cytokines. In addition to its action on B lymphocytes, this report demonstrates that CD40L induced both CD4+ and CD8+ T cells isolated from murine lymphoid tissues to proliferate in the presence of submitogenic dosages of Con A, PHA, CD3 mAb, and TCR-alpha beta mAb. The presence of CD40L during suboptimal TCR stimulation resulted in increased expression of the activation Ags IL-2R alpha and CD69 and increased IL-2 production. Taken together, these results show that CD40L is a potent activator of murine T cells and suggest that CD40L is involved in the regulation of T cell function mediated through T:T cell interaction.

Animals↗
PubMed1994

gld/gld mice are unable to express a functional ligand for Fas.

Mice homozygous for either the lpr or gld genes develop phenotypically identical autoimmune disorders. The gene responsible for the pathology in lpr/lpr mice encodes the Fas antigen, a protein associated with the induction of programmed cell death. To determine if the defect associated with gld represents a mutation in the ligand for Fas, we have assessed the ability of lymphoid cells from homozygous gld/gld mice to lyse target cells in a Fas-dependent manner. Using an antagonistic antibody to Fas, we demonstrate that activated T cells from normal and lpr mice are capable of inducing Fas-mediated lysis of tumor target cells. In contrast, activated T cells from gld/gld mice fail to induce lysis of tumor targets, although cells from gld mice are able to lyse specific allogeneic targets following mixed lymphocyte culture. In addition, activated T cells from gld/gld homozygous animals are not capable of binding to a Fas.Fc fusion protein at high levels, whereas activated T cells from normal and lpr/lpr animals bind Fas.Fc efficiently. These data indicate that mice homozygous for gld are unable to express a functional ligand for Fas.

Animals↗
PubMed1994

Molecular and biological characterization of human 4-1BB and its ligand.

4-1BB was originally described as a cDNA expressed by activated murine T cells and subsequently demonstrated to encode a member of the tumor necrosis factor receptor family of integral membrane proteins. Recently, we identified and cloned a murine ligand for 4-1BB (mu4-1BB-L) and demonstrated it to be a member of an emerging family of ligands with structural homology to tumor necrosis factor. To characterize further the role of 4-1BB in the immune response we undertook to clone the human homologue of 4-1BB-L. However, attempts to isolate a cDNA encoding the human 4-1BB-L by cross-hybridization with the murine cDNA were unsuccessful. Therefore we first utilized cross-species hybridization to isolate a cDNA encoding human 4-1BB (hu4-1BB). A fusion protein consisting of the extracellular portion of hu4-1BB coupled to the Fc region of human immunoglobulin G1 (hu4-1BB.Fc) was then used to identify and clone a gene for human 4-1BB-L from an activated CD4+ T cell clone using a direct expression cloning strategy. Human 4-1BB-L shows 36% amino acid identity with its murine counterpart and maps to chromosome 19p13.3. Scatchard analysis demonstrated high-affinity binding of hu4-1BB.Fc to either native or recombinant human 4-1BB-L. Both monoclonal antibody to hu4-1BB and cells transfected with hu4-1BB-L induced a strong proliferative response in mitogen co-stimulated primary T cells. In contrast, ligation of 4-1BB on T cell clones enhanced activation-induced cell death when triggered by engagement of the TCR/CD3 complex.

Amino Acid Sequence↗
PubMed1994

The activation antigen CD69.

One of the earliest cell surface antigens expressed by T cells following activation is CD69, which is detectable within one h of ligation of the T cell receptor/CD3 complex. Once expressed, CD69 acts as a costimulatory molecule for T cell activation and proliferation. In addition to mature T cells, CD69 is inducibly expressed by immature thymocytes, B cells, natural killer (NK) cells, monocytes, neutrophils and eosinophils, and is constitutively expressed by mature thymocytes and platelets. Recently, cDNA clones encoding human and mouse CD69 were isolated and showed CD69 to be a member of the C-type lectin superfamily. Gene mapping studies have placed CD69 on distal mouse chromosome 6 and human chromosome 12p13, close to, if not in, the NK gene complex. The structure, chromosomal localization, expression and function of CD69 suggest that it is likely a pleiotropic immune regulator, potentially important not only in NK cell function but also in the activation and differentiation of a wide variety of hematopoietic cells.

Animals↗
PubMed1994

Synergistic effects of IL-4 and either GM-CSF or IL-3 on the induction of CD23 expression by human monocytes: regulatory effects of IFN-alpha and IFN-gamma.

CD23 expression by B cells and monocytes can be regulated by cytokines including IL-4, IFN-gamma and IFN-alpha. We recently reported that GM-CSF and IL-3 are also capable of regulating CD23 expression, with GM-CSF enhancing CD23 expression by monocytes and IL-3 enhancing CD23 expression by both monocytes and B cells. In this study, we have assessed the effect of combinations of cytokines on monocyte CD23 expression. IL-4 acted in a synergistic manner with either GM-CSF or IL-3 to induce monocyte CD23 expression. Interestingly, culture of monocytes with GM-CSF, IL-3 or IL-4 resulted in two subpopulations of cells, one negative or dull for surface CD23 and the other expressing relatively high levels of CD23. In contrast, culturing monocytes in the combination of IL-4 with either GM-CSF or IL-3 resulted in a single population of cells expressing very high levels of CD23. The synergy between IL-4 and either GM-CSF or IL-3 was also reflected at the level of release of soluble CD23 and CD23 mRNA expression and was seen at both sub-optimal and optimal cytokine concentrations. GM-CSF, IL-3 and IL-4 all enhanced monocyte expression of class II MHC, though no additive or synergistic effects were seen with cytokine combinations. IFN-gamma failed to induce monocyte CD23 expression when acting alone, though it enhanced both surface and soluble CD23 when acting in the presence of GM-CSF, IL-3 or IL-4. In contrast, IFN-alpha was a potent inhibitor of monocyte CD23 expression induced by these three cytokines.(ABSTRACT TRUNCATED AT 250 WORDS)

Cells, Cultured↗
PubMed1994

The mouse Fas-ligand gene is mutated in gld mice and is part of a TNF family gene cluster.

The gene for the mouse Fas ligand was cloned and its chromosomal position determined. Fasl was tightly linked to gld (no crossovers in 567 meiotic events) on mouse chromosome 1 and closely linked with a novel member of the same TNF family of ligands, the Ox40 ligand (Ox40l, 1 crossover in 567 meiotic events). Southern blot analysis did not reveal any difference between the Fasl gene from gld and wild-type mice and levels of Fasl mRNA transcripts were similar in PMA and ionomycin induced wild-type and coisogenic gld T cells. Sequence analysis of the gld gene indicated a single amino acid change (Phe Leu) in the COOH terminal portion of this type II transmembrane protein, and COS cells transfected with Fasl cDNA from gld mice failed to induce apoptosis of Fas-expressing target cells. Thus, the data demonstrate that the gld phenotype is the result of a point mutation in the Fasl gene and that Fasl is part of a complex of ligands structurally related to TNF mapping within a small region of mouse chromosome 1.

Amino Acid Sequence↗
PubMed1994

Regulation of apoptosis and T cell activation by Fas-specific mAb.

Fas was initially described as a molecule expressed on the surface of certain cell lines that could mediate programmed cell death (apoptosis) subsequent to ligation by specific mAb. To determine whether mAb to other epitopes on the Fas molecule might mediate other functions, we generated a panel of mAb to the extracellular portion of human Fas. Significant lysis of Fas-expressing target cells was only observed when the new mAb were first bound to a solid-phase support and not when the mAb were added in solution. However, several of these mAb inhibited the killing of target cells induced by the prototypic Fas-specific mAb, CH-11. Those mAb that inhibited apoptosis of target cells mediated by the CH-11 mAb also blocked lysis of target cells mediated by cells expressing Fas ligand. Finally, some of the Fas-specific mAb were found to co-stimulate proliferation of peripheral blood T cells in the presence of immobilized CD3 mAb. Thus, the data indicate the existence of a complex set of interactions mediated by Fas in both normal and transformed lymphoid cells that may have important implications regarding the role(s) of this molecule in regulation of immune responses.

Animals↗
PubMed1994

Regulation of immune responses by the ligands for CD27, CD30, and 4-1BB.

Activated T cells express a number of cell surface proteins that play an important role in cell contact-dependent interactions with cells in the immune system, including B cells, macrophages and other T cells. Among these T cell-expressed proteins are members of the TNF ligand family, which includes TNF, lymphotoxin (LT)-alpha and -beta, and the ligands for CD27, CD30, CD40, 4-1BB, OX-40, and Fas. The recent cloning of a number of these ligands has paved the way for a detailed analysis of the role of these molecules in immune responses. Initial studies have suggested that most, if not all, of these ligands co-stimulate T-cell proliferation. It is becoming increasingly apparent, however, that members of the TNF ligand family play unique and critical roles in both the development of the immune system and in immune responses to specific pathogens.

Antigens, CD↗
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

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↗