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S Gillis

Publications and source records attributed to S Gillis.

At least 127 records · Page 7Linked to original sources
PubMed1988

In vivo treatment with IL-1 reduces the severity and duration of antigen-induced arthritis in rats.

IL-1 has been implicated in the pathogenesis of arthritis. Although IL-1 injected in vivo into normal joints results in a transient inflammatory reaction, we have shown that three weekly repetitive injections of IL-1 do not produce a progressive inflammatory condition suggestive of chronic arthritis. In fact, priming normal joints with three weekly IL-1 intraarticular injections results in a significant reduction in joint swelling and diminished histopathologic alterations/lesions caused by subsequent methylated BSA-induced arthritis. Similarly, post treatment with IL-1 intraarticular injection after arthritis induction reduced arthritic swelling and joint injury if IL-1 was given during the developing stage of arthritis. Our results suggest that IL-1 might limit arthritic inflammation and progressive cartilage destruction through an, as yet, undetermined mechanism(s). Further in vivo investigations are required to determine the therapeutic utility of IL-1 in reducing early arthritic inflammation.

Animals↗
PubMed1988

IL-4 bypasses the immune suppressive effect of cyclosporin A during the in vitro induction of murine cytotoxic T lymphocytes.

We have analyzed at the clonal level the effect of IL-4 on the immune suppressive action of cyclosporin A (CsA) during the in vitro primary activation of anti-MHC alloantigen-reactive murine CD8+ CTL. Although neither IL-4 nor IL-2 alone were able to overcome the CsA-mediated suppression, the addition of IL-4 in the presence of IL-2 restored in a dose-dependent manner the induction of cytolytic activity. On the other hand, CsA greatly impaired proliferative responses of alloantigen-reactive CD8 T cells, thereby operationally dissecting proliferative responsiveness from acquisition of cytolytic activity during primary activation of alloantigen-reactive CD8+ T cells. The existence of a CsA-resistant induction pathway for Ag-specific CD8+ T cell-mediated cytolytic activity may be of relevance for experimental and clinical organ transplantation.

Animals↗
PubMed1988

Binding, internalization, and intracellular localization of interleukin-1 beta in human diploid fibroblasts.

This study demonstrates internalization of interleukin-1 (IL-1) via its cell surface receptor on human diploid fibroblasts and shows intracellular localization of IL-1 beta. Binding experiments at 8 degrees C using confluent fibroblast monolayers revealed 5,000-15,000 IL-1 receptors/cell that bound both IL-1 alpha and IL-1 beta. Incubation of monolayers with 125I-IL-1 beta (10(-9) M) at 8 degrees C and then at 37 degrees C for various times up to 8 h revealed a t1/2 for internalization of receptor-bound IL-1 beta of about 1.5 h. In addition, it was shown that IL-1 beta internalized via receptors was undegraded and retained binding activity. Electron microscopic autoradiography of monolayers incubated with 125I-IL-1 beta, as above, showed a progressive increase in the ratio of cytoplasmic to cell surface-associated grains. Grains at the cell surface were primarily localized at cell processes or attachment sites, frequently close to intra- and extracellular filamentous material. During incubation at 37 degrees C, most grains were free in the cytoplasm, with few present in lysosomes or vesicles. After 1 h, approximately 15% of the grains were over nuclei. Control cultures incubated at 37 degrees C with 125I-IL-1 beta and 100-fold excess unlabeled IL-1 beta showed increased uptake of label into lysosomes and little into nuclei. This study shows that IL-1 receptors are primarily located at fibroblast processes and that receptor-mediated internalization of the ligand is slow. Nuclear localization apparently requires IL-1 receptor-specific internalization of IL-1 beta, suggesting a possible role for this process in eliciting the IL-1 signal.

Cell Membrane↗
PubMed1988

Stimulation of B-cell progenitors by cloned murine interleukin-7.

The events involved in the commitment and development of lymphoid lineage cells are poorly understood. We have used a recently described long-term culture system to establish a bioassay that can detect a novel growth factor capable of stimulating the proliferation of lymphoid progenitors. Using direct expression in mammalian cells we have isolated a complementary DNA clone encoding this novel haematopoietic growth factor, designated interleukin-7.

Amino Acid Sequence↗
PubMed1988

Soluble CD23 is released by B lymphocytes cycling in response to interleukin 4 and anti-Bp50 (CDw40).

An enzyme-linked immunoassay was developed to quantitate the production of soluble CD23 from cycling B lymphocytes. This molecule has identity both with B cell-derived B cell growth factor and with an IgE-binding factor. B lymphocytes, which had been stimulated for 3 days with phorbol dibutyrate and calcium ionophore, washed and recultured, failed to produce detectable levels of CD23 over the following 3 days. Soluble CD23 was found, however, in the supernatant of cultures where recombinant interleukin 4 had been included. The level of CD23 rose dramatically when anti-Bp50 had also been added. By contrast, anti-Bp50, alone or together with low molecular weight T cell-derived B cell growth factor, failed to promote the release of CD23 in detectable amounts. There was a strong correlation between the appearance of soluble CD23 in culture supernatants and the expression of CD23 on the surface of restimulated cells. The level of CD23 release appeared to relate more to the continued cycling of cells than to their differentiation to immunoglobulin secretion. These findings are discussed with particular emphasis on the role of CD23 as an important multi-functional lymphokine in B lymphocyte physiology.

B-Lymphocytes↗
PubMed1988

Interleukin 4 (BSF-1) induces growth in resting murine CD8 T cells triggered via cross-linking of T3 cell surface structures.

To analyze the role of interleukin 4 (IL4, BSF-1) during primary activation of resting (high-density) murine CD8 T cells, a model system was used which bypasses antigen-presenting cells by the use of anti-T3 monoclonal antibodies immobilized on Sepharose beads. In high, but not in low cell density cultures, IL4 alone induced cell growth. In low cell density cultures, further to T3 cross-linking a soluble macrophage product was required as co-stimulator to induce sensitivity to IL4. This co-stimulator activity was unrelated to recombinant (r)IL1, rIL6 and rTNF-alpha (tumor necrosis factor alpha). In primary CD8 T cell responses rIL4-driven growth was about half of that induced by rIL2, and not inhibitable by anti-IL2 receptor antibodies. Higher concentrations of IL4 down-regulated cell proliferation. In the course of IL4-driven growth, the proliferating cells acquired sensitivity to the growth-promoting effect of IL2. Activated CD4 or CD8 T cells were found to be equally sensitive to the IL4 and IL2-driven growth pathway. Taken together, these results define a physiologic role of IL4 as growth factor during primary activation of resting CD8 T cells and thus extend the spectrum of target cells for IL4.

Animals↗
PubMed1988

Interleukin 4 and soluble CD23 as progression factors for human B lymphocytes: analysis of their interactions with agonists of the phosphoinositide "dual pathway" of signalling.

Human B lymphocytes pre-activated for 24 h with a combination of phorbol dibutyrate [P(Bu)2] and ionomycin were found to provide excellent targets for assessing the detailed action of B cell progression factors. Both recombinant interleukin 4 (IL 4) and affinity-purified 25-kDa fragment of the CD23 molecule (sol-CD23) were shown to be active in this assay. While the progression activity of IL 4 was enhanced by continued co-culture with P(Bu)2, that of sol-CD23 was found to be more strictly dependent upon such a joint application with the phorbol ester. Similar requirements were observed for triggering cell-cycle progression in the pre-activated B cells when using a stimulating CD23 antibody. Ionomycin, in contrast to P(Bu)2, did not augment either IL 4 or sol-CD23 in these assays but did enhance significantly the progression activity of an anti-CDw40 antibody. When added to B cells concomitantly with, or prior to, a high dose of phorbol ester, IL 4 unexpectedly down-regulated the subsequent mitogenic response to this agent whereas, when added 24 h later, IL 4 up-regulated such stimulations. The latter sequence of additions resulted in a particularly dramatic induction of CD23 at the B cell surface, much more so than seen when B cells were incubated with either IL 4 alone or with IL 4 and P(Bu)2 together. This up-regulation of surface CD23 was, in turn, mirrored by the appearance of large amounts of the soluble form of the molecule in such cultures. The findings are discussed with reference to possible mechanisms through which IL 4 and CD23 interact to exert their multiple actions on B cell regulatory pathways.

Antigens, Differentiation, B-Lymphocyte↗
PubMed1988

Human macrophage-colony stimulating factor: alternative RNA and protein processing from a single gene.

Macrophage-colony stimulating factor (M-CSF, CSF-1) has been reported to be required for the proliferation and differentiation of macrophages from hematopoietic progenitor cells. Recently, two human M-CSF cDNA clones were isolated encoding proteins of 256 and 554 amino acids. We report here the isolation of a third M-CSF cDNA that encodes a protein of 438 amino acids. The coding regions for the three cDNA clones share a common amino-terminus of 149 amino acids and a common carboxyl-terminus of 75 amino acids including a membrane spanning region. In addition, we isolated a genomic clone of human M-CSF. When each of the cDNA clones or the genomic clone were transfected into COS-7 monkey kidney cells, biologically active M-CSF was expressed as judged by the ability of transfected cell supernatants to stimulate proliferation and colony formation of murine bone marrow cells, as well as formation of monocytic colonies from human bone marrow cells. Surprisingly, proliferation of human bone marrow cells was not induced by recombinant human M-CSF. Analysis of the M-CSF proteins released by COS-7 cells revealed that monomer subunit proteins of 44 or 28 kDa were produced. In addition, we found that the membrane spanning region, present in all three forms of M-CSF cDNA, was not required for the synthesis of a biologically active protein. However, when the membrane spanning region was present in the three M-CSF cDNAs, cell surface associated forms of M-CSF could be readily detected.

Animals↗
PubMed1988

Bovine GM-CSF: molecular cloning and biological activity of the recombinant protein.

The lymphokine granulocyte-macrophage colony-stimulating factor (GM-CSF) mediates the growth and differentiation of granulocytes and macrophages from bone marrow progenitors, and regulates biological functions expressed by mature cells of these lineages. In order to isolate a bovine GM-CSF cDNA, a cDNA library, generated from the BT2 bovine T cell line, was screened with a human GM-CSF cDNA probe. A cDNA clone was isolated with an insert of 783 bp, that would encode a protein of 143 amino acids, with a predicted mol. wt of 16,160. Bovine GM-CSF exhibits a high degree of sequence homology with mouse and human GM-CSF at both the nucleotide and amino acid levels. Comparison of GM-CSF amino acid sequences from the three species indicates that the bovine GM-CSF precursor contains a putative 17 amino acid signal sequence, cleavage of which would yield a 14,250 mol. wt protein. The cDNA was inserted into a mammalian expression vector and transfected into COS-7 monkey kidney cells. Biologically active bovine GM-CSF was secreted as judged by a bovine bone marrow proliferation assay. Bovine GM-CSF was weakly active in both human and mouse bone marrow proliferation assays. In contrast, human GM-CSF was weakly active on bovine but not murine mouse bone marrow cells and mouse GM-CSF was only active on murine bone marrow cells.

Amino Acid Sequence↗
PubMed1988

Cloning, sequence and expression of bovine interleukin 1 alpha and interleukin 1 beta complementary DNAs.

Interleukin 1 (IL-1) is a cytokine which mediates a variety of immunoregulatory and inflammatory activities. Using human IL-1 alpha and IL-1 beta probes, cDNAs for the corresponding bovine genes were isolated from an alveolar macrophage library. The open reading frames of the bovine IL-1 alpha and IL-1 beta cDNAs encode proteins of 268 and 266 amino acids, respectively, each with a predicted mol. wt of approx. 31,000. Both forms of bovine IL-1 exhibit a high degree of sequence homology with IL-1 gene products from other mammalian species. Based upon comparisons with human IL-1 amino acid sequences, the post-translationally processed, mature forms of bovine IL-1 would occur as 17-18,000 mol. wt proteins. Sequences encoding mature bovine IL-1 alpha and IL-1 beta were inserted into E. coli expression plasmids and biologically active proteins were synthesized as judged by the ability of the recombinant proteins to induce proliferation of bovine thymocytes. Both IL-1 alpha and IL-1 beta exist as single genomic copies. In addition, bovine IL-1 beta mRNA is approx. 10-fold more abundant than IL-1 alpha mRNA in stimulated alveolar macrophages.

Amino Acid Sequence↗
PubMed1988

Expression of functional human interleukin-2 receptors in murine interleukin-3-dependent cells.

A murine recombinant retrovirus containing the cDNA encoding the human p55 interleukin-2 (IL2)-binding protein was used to insert this gene into a murine interleukin-3 (IL3)-dependent cell line, FD.C/1. Virus-infected cells, maintained in medium supplemented with IL3, expressed human p55 on the cell surface and readily adapted to growth using human IL2. In the presence of human IL2, the synthesis of the endogenous murine p55 binding protein was induced in FD.C/1 cells, making it difficult to determine whether the human p55 protein was actively involved in the process of growth signal transduction. A cloned cell line, FD.huIL2R-2, was identified which grew in the presence of human IL2 but which had lost the ability to synthesize murine p55 protein. Growth of this clone was inhibited by the monoclonal antibody 2A3 which specifically blocked binding of human IL2 to the human p55 binding protein. Analysis of restriction enzyme digests of FD.huIL2R-2 cell DNA revealed that a rearrangement of a murine p55 gene had occurred, implying that virus infection had resulted in the integration of retroviral DNA at a site close to or within a murine p55 gene. If IL2 signal transduction involves binding to a surface heterodimeric receptor for IL2, it is argued that FD.huIL2R-2 cells contain an IL2 receptor complex of murine p70 and human p55 IL2-binding proteins. Alternatively, it is possible that integration of human p55 DNA into a site close to a murine p55 gene may lead to a hybrid p55 IL2-binding protein. If FD.huIL2R-2 cells express murine p70 IL2-binding protein as part of the receptor complex, the inability of cells to grow in murine IL2 implies that IL2 binding to p70 protein alone is insufficient for a growth signal in these cells. FD.huIL2R-2 cells grow at rates similar in IL3- or human IL2-dependent states. It is likely therefore that the biochemical pathways that control each of these lymphokine-dependent growth states are very similar.

Animals↗
PubMed1988

Stimulation of neutrophil production in CSF-1-responsive clones.

The hematopoietic growth factor CSF-1 has been considered relatively lineage specific for the production of macrophages, whereas GM-CSF elicits a predominance of neutrophils. It is likely that in vivo, individual clones are stimulated by the two CSFs, although the effect of dual stimulation on progenitors and their progeny has not been completely explored. We found that in cultures initiated with low concentrations of CSF-1 or GM-CSF, alone or in combination, production of macrophages predominated. Maximally stimulatory concentrations of CSF-1 elicited a predominance of macrophages, whereas maximal GM-CSF elicited many more neutrophil/macrophage colonies and pure neutrophil colonies. A combination of maximal CSF-1 and GM-CSF elicited the same differentiation as GM-CSF alone. Delayed addition of GM-CSF to cultures initiated with CSF-1 elicited colonies indistinguishable from GM-CSF alone, suggesting that neutrophil production had been switched on by GM-CSF. In mapping studies, colonies initiated by CSF-1 increased or switched on neutrophil production when GM-CSF was added as a second stimulus. These studies show that individual clones are responsive to both CSFs, and that the differentiating influence of GM-CSF predominates over that of CSF-1. In cultures to which only CSF-1 was added, a population of progenitors was sustained that produced neutrophils only after a GM-CSF stimulus. Thus, CSF-1 may participate in maintaining a reserve of progenitors for neutrophils during periods of increased neutrophil demand.

Animals↗
PubMed1988

Enhanced survival but reduced engraftment in murine recipients of recombinant granulocyte/macrophage colony-stimulating factor following transplantation of T-cell-depleted histoincompatible bone marrow.

In vivo administration of murine recombinant granulocyte/macrophage colony stimulating factor (rGM-CSF) was evaluated for effects on survival and engraftment in an allogeneic murine bone marrow transplantation (BMT) model involving T-cell depletion of donor marrow. The model provides a high incidence of graft failure/rejection. Recipients of continuous subcutaneous infusions of rGM-CSF had a significant survival advantage when compared with untreated controls. However, a significantly lower incidence of donor cell engraftment was noted. Hematological parameters were not substantially affected. When rGM-CSF was administered intraperitoneally (IP), twice daily injections closely approximated the effects of continuous infusion on survival. Single IP injections were without significant effects on survival or engraftment. These results demonstrate that prolonged frequent in vivo exposure to rGM-CSF can significantly improve survival but significantly decreases donor cell repopulation in recipients of T-cell-depleted histoincompatible marrow grafts.

Actuarial Analysis↗
PubMed1988

In vitro and in vivo cytokine-induced facilitation of immunohematopoietic reconstitution in mice undergoing bone marrow transplantation.

The aim of this study was to test whether colony stimulating factors (CSF) and other cytokines facilitate the recovery of a variety of immunohematopoietic functions in lethally irradiated mice undergoing bone marrow transplantation (BMT). Two experimental systems were employed: (a) lethally irradiated mice transplanted with syngeneic or T cell-depleted semi-allogeneic bone marrow (BM) cells (0.1-10 x 10(6)), subsequently treated by multiple doses of cytokines; and (b) lethally irradiated mice transplanted with BM cells that had previously been cultivated with cytokines. The cytokines used were: pure natural mouse interleukin-3 (IL-3); recombinant mouse granulocyte-macrophage CSF (rGM-CSF); recombinant human interleukin-2 (rIL-2); and crude cytokine preparations obtained from the culture supernatants of murine leukemia WEHI-3b cells (containing mainly IL-3), and of phorbol myristate acetate (PMA)-stimulated EL4 leukemia cells and concanavalin A-stimulated rat splenocytes (each containing a multitude of cytokines). For BM cultures (1-9 days), the cytokines were used at a dosage of 1-100 U/ml; for in vivo treatment, 2 x 10(2)-5 x 10(4) units were administered intraperitoneally and subcutaneously at different schedules for varying periods (1-3 weeks). The following parameters were tested 1-10 weeks post-BMT: white blood cell count, colony formation in agar and in the spleen of lethally irradiated mice, proliferative responses to mitogens and alloantigens, allocytotoxicity and antibody production (serum agglutinins and plaque-forming cells) against sheep red blood cells. Under appropriate conditions, cytokine treatment either in vitro or in vivo significantly enhanced (2- to 50-fold compared with controls) most functions tested at 2-8 weeks post-BMT, and shortened the time interval required for full immunohematopoietic recovery by 2-5 weeks. In recipients of semi-allogeneic, T lymphocyte-depleted BM no evidence of graft-versus-host disease was found. It is suggested that judicious application in vitro and/or in vivo of certain pure cytokines (e.g. GM-CSF, IL-3) or cytokine 'cocktails' might be beneficial in enhancing hematopoiesis and in the treatment of immunodeficiency associated with BMT.

Animals↗
PubMed1988

Production of recombinant human colony stimulating factors in yeast.

Efficient yeast expression and purification systems for production of recombinant human GM-CSF, IL-3 and G-CSF have been established. Though yeast-derived production of recombinant CSFs (through the use of secretion based system) allows for generation of native molecules which can then be readily separated from fermentation broth, in many instances, natural cDNAs have had to be altered to allow for efficient expression, as well as production of a less heterogeneous product. In the case of CSFs described herein, beneficial mutations (made through site-directed mutagenesis) have included elimination of potential N-linked glycosylation sites, removal of KexII protease recognition sites (notably alterations in dibasic sequences) and elimination of extraneous cysteine residues which might complicate isolation of a homogeneous product due to intermolecular disulfide bonding.

Amino Acid Sequence↗
PubMed1988

Recombinant granulocyte/macrophage-colony stimulating factor in human and murine bone marrow transplantation.

Recombinant granulocyte/macrophage-colony stimulating factor (rGM-CSF) is a protein that mediates a broad range of biological activities including inducing proliferation of bone marrow progenitor cells and up-regulating the functional capabilities of neutrophils, monocytes, and macrophages. We have administered rGM-CSF in vivo in the context of allogeneic (murine) and autologous (human) bone marrow transplantation (BMT). In a murine allogeneic model of T-cell depletion constructed to favor graft failure/rejection of donor marrow, ex vivo treatment of marrow with rGM-CSF significantly promoted engraftment. However, when rGM-CSF was infused in vivo, engraftment was significantly impaired. We have initiated a phase I clinical trial of rGM-CSF in human autologous BMT for patients with acute lymphoblastic leukemia (ALL). rGM-CSF has been administered for 14-21 days in a dose escalation study (16-128 micrograms/M2/day). At high doses of rGM-CSF, neutrophil recovery has returned earlier and to higher levels than previously observed in historical controls. rGM-CSF related toxicity in this study has been mild.

Adolescent↗