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S Gillis

Publications and source records attributed to S Gillis.

At least 235 records · Page 13Linked to original sources
PubMed1981

The role of interleukin-2 (IL-2) in the differentiatin of cytotoxic T cells: the effect of monoclonal anti-IL-2 antibody and absorption with IL-2 dependent T cell lines.

A variety of approaches, including the use of a monoclonal antibody, have indicated that the lymphokine Interleukin 2 is a necessary and sufficient mediator for initiating cytotoxic T cell differentiation in vitro. Mitogen (Con A, PHA) stimulation of spleen cells and K/D or I region stimulation in primary MLC all led to IL-2 production, as measured by the growth of an IL-2 dependent T cell clone, and to the production of a "helper" factor necessary for cytotoxic T lymphocyte (CTL) differentiation against metabolically inactivated stimulator spleen cells. Several lines of evidence suggested that the IL-2 activity and CTL "helper" activity were due to a single molecular entity. These included the observations that: i) There was a linear relationship (R = 0.99) between IL-2 activity and relative CTL "helper" activity regardless of the preparation used (purified IL-2; MLC, or mitogen-induced supernatants). ii) Absorption of MLC supernatants with T cell "blasts" or with IL-2 dependent T cell lines concomitantly removed both IL-2 and CTL "helper" activities. iii) IL-2 activity purified by isoelectric focusing, or by polyacrylamide gel electrophoresis, displayed CTL "helper" activity. iv) Both CTL "helper" and IL-2 activities were neutralized by a monoclonal antibody directed against IL-2.

Absorption↗
PubMed1981

Interleukin-2 induction of hapten-specific cytolytic T cells in nude mice.

In light of i) increasing evidence that the thymus guides the development and antigen recognition repertoire of cytotoxic T cells (CTL) and ii) the capacity of purified interleukin 2 (IL-2) to allow for the in vitro generation of CTL from nude mice, we investigated the ability of IL-2 to influence the in vitro generation of hapten-specific cytotoxicity from nude mouse spleen cell cultures. We found that stimulation of nude mouse splenocytes with trinitrophenol (TNP)-derivatized autologous or allogeneic spleen cells in the presence of IL-2 generated hapten-specific CTL. EFfector cells harvested from IL-2-supplemented cultures killed only TNP-derivatized target cells; however, the cytotoxicity observed was not restricted to a particular target histocompatibility type. This study confirms our previous observations that IL-2 has the capacity to induce growth of CTL from pre-T cell populations in the total absence of thymic influence. However, it is clear that the generation of H-2 restricted killing (recognition and lysis of cells bearing antigen presented solely in association with "self" H-2) probably requires further processing of pre-killer T cell populations by a thymus-derived differentiation signal. In addition, these studies provide further evidence that a major role of the thymus, in addition to programming effector T cell antigen recognition repertoire, may be its function to provide essential differentiation of the IL-2 producer T cell.

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PubMed1981

Constitutive production of interleukin 2 activity by a T cell hybridoma.

Several laboratories have recently confirmed the importance that interleukin 2 (IL-2, formerly referred to as T cell growth factor) plays not only in the maintenance of cultured functional T cell lines but also as a potent regulator of immune responses. Despite the development of preparative IL-2 purification procedures, as well as identification of IL-2 producer tumor cell lines, simple sources of lectin-free IL-2 containing conditioned medium remain undefined. In hopes of establishing such a source of mitogen-free IL-2, we elected to fuse the high titer IL-2 producer tumor cell line, LBRM-33, with a drugmarked T cell lymphoma. In this communication we report the successful isolation of a T cell hybridoma that constitutively synthesizes and secretes biologically active IL-2. Hybridoma conditioned medium contained approximately 10 to 100 times the amount of IL-2 produced by conventional T cell mitogen-stimulated murine splenocyte preparations. In addition to providing a simple means for producing IL-2 containing conditioned medium without the presence of contaminating lectin, we hope that constitutive IL-2 producer hybrid cell lines will prove interesting as vehicles for dissection of the molecular events involved in IL-2 biosynthesis.

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PubMed1981

The biochemical and biological characterization of lymphocyte regulatory molecules. VI. Generation of a B cell hybridoma whose antibody product inhibits interleukin 2 activity.

In attempts to generate monoclonal antibodies with reactivity directed against the lymphokine Interleukin 2 (IL-2, T cell growth factor), spleen cells harvested from BALB/c mice previously immunized with rat IL-2 were fused with the BALB/c myeloma SP2. Several of the resultant hybrid cell lines secreted a product that significantly neutralized (greater than 50%) IL-2--dependent T cell proliferation. Several lines of evidence suggested that the inhibitory activity was associated with a monoclonal IgG antibody directed against IL-2 determinants. First, passage of cloned hybrid cell culture supernatants through a protein A-coupled Sepharose column yielded purified immunoglobulin G fractions that inhibited mouse, rat, and human IL-2 activity. Secondly, hybridoma-derived IgG, in concert with lyophilized Staphylococcus aureus, was capable of precipitating both "cold" and intrinsically labeled IL-2 activity. Finally, Sepharose conjugated with purified IgG fractions provided on extremely reactive IL-2 absorption matrix. These results suggest that monoclonal antibodies directed against IL-2 determinants may eventually provide new detection assays for IL-2 and allow affinity chromatography to be employed for the isolation of this lymphokine.

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PubMed1981

Biochemical and biologic characterization of lymphocyte regulatory molecules. VIII. Purification of interleukin 2 from a human T cell leukemia.

The ability of interleukin 2 (IL 2) to stimulate and maintain growth in IL 2-dependent cell lines distinguishes this substance from interleukin 1 and other T cell lymphokines. Jurkat-FHCRC, a human T cell leukemia cell line, has been shown to produce relatively high quantities of IL 2 after stimulation with the T cell mitogens phytohemagglutinin or concanavalin A. In this paper we describe techniques used to isolate human IL 2 from Jurkat supernatants and the biochemical properties of the resulting material. IL 2 produced by this cell line is eluted from a DEAE-Sephacel ion-exchange column with 0.05 M NaCl and has a m.w. determined by AcA54 gel chromatography in the range of 9000 to 18,000. Isoelectric focusing yields active material with a pI value of 7.75. Elution of IL 2 from a SDS-polyacrylamide gel shows it to have a m.w. of approximately 13,500. This material is capable of stimulating mouse as well as human IL 2-dependent cell lines.

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PubMed1981

Purification of murine helper T cell-replacing factors.

Humoral factors secreted by T lymphocytes and macrophages appear to play a role in cell communication leading to the triggering of immune responses. A factor has been purified from the culture supernatants of concanavalin A(Con A)-activated murine spleen cells with lymphokine activity in four assay systems. 1) stimulation of antibody responses to erythrocyte antigens in BALB/c.nu spleen cultures 2) amplification of production of cytotoxic T cells in thymocyte cultures, 3) stimulation of mitogenic responses to Con A in thymocyte cultures where the cell density is too low to support responses to Con A alone, and 4) maintenance of continuous T cell growth. The biologic activity has been sequentially purified by salt precipitation, gel filtration, chromatography, ion-exchange chromatography, and isoelectric focusing (IEF). These four lymphokine activities appear to be due to one class of molecules, termed Interleukin 2 (IL-2), with an apparent molecular weight of 30,000 daltons. A T cell lymphoma, LBRM-33 has been found to produce high titers of IL-2 activity upon mitogen stimulation. LBRM-derived IL-2 possesses similar chemical and biological properties to splenic-derived IL-2. The mode of action of IL-2 appears to be the clonal expansion of antigen- or mitogen-activated T cells. While mitogen-activated LBRM cells secrete IL-2, which appears to exert helper T cell-replacing activity via the expansion of pre-T cells in nude spleens, mitogen-activated spleen cells appear to secrete both IL-2 and another class of T cell-replacing factor (TRF). These findings raise the question of whether there exist multiple classes of factors with TRF activity. The identification of these molecules may lead to an understanding of the mechanism that regulates the induction of antigen-sensitive cells.

Animals↗
PubMed1980

Biochemical and biological characterization of lymphocyte regulatory molecules. V. Identification of an interleukin 2-producing human leukemia T cell line.

To isolate a stable tumor cell line capable of producing human interleukin 2 (IL-2; formerly referred to as T cell growth factor), 16 human T and B leukemia cell lines were screened for constitutive and mitogen-stimulated IL-2 production. We found that the T cell leukemia line designated Jurkat-FHCRC produced > 200 U/ml of IL-2 activity after a 24-h stimulation with T cell mitogens. Peak mitogen-induced IL-2 activity was found in supernates harvested from 24-h Jurkat-FHCRC cell cultures stimulated with either 1% phytohemagglutinin or 20 microgram/ml concanavalin A. Addition of the fatty acid derivative phorbol myristate acetate to mitogen-stimulated cultures increased Jurkat-FHCRC IL-2 production to concentrations > 400 U/ml. IL-2 activity observed in such cases represented between 100--300 times that produced in conventional cultures of mitogen- or alloantigen-stimulated normal human peripheral blood or splenic lymphocytes. Jurkat-FHCRC-derived conditioned medium demonstrated equal capacity to promote the sustained in vitro proliferation of either murine or human activated T cell lines confirming the ability of Jurkat-FHCRC cells to produce human IL-2. These studies identify a new source of human IL-2 and establish a valuable reagent for the isolation and further molecular characterization of this immunoregulatory molecule.

Animals↗
PubMed1980

Biochemical separation of interleukin 2.

Interleukin 2 (IL-2) is a class of T cell growth factors produced by T cells of a number of species. The unique growth-promoting properties of these molecules allow the development of antigen-specific effector T cell lines which can be used to analyze the molecular basis of lymphocyte interactions. A murine T cell tumor line has been identified as a source of IL-2. The purification and biochemical properties of murine IL-2 are described, and compared with rat and human Il-2.

Animals↗
PubMed1980

Effect of chronic sulpiride on striatal spiperone binding.

Increased 3H-spiperone binding after chronic neuroleptic treatment has been proposed as a molecular model of tardive dyskinesia. Sulpiride has been claimed to be an atypical neuroleptic that might not produce tardive dyskinesia. The effect of chronic sulpiride was, therefore, compared to that of chronic haloperidol on striatal 3H-spiperone binding. 3 weeks of haloperidol feeding caused a 28% increase in 3H-spiperone binding, whereas even very high dose sulpiride had no effect on spiperone binding. These findings support the concept that sulpiride may be a unique neuroleptic with regard to long-term effects on dopamine receptors.

Animals↗
PubMed1980

Biochemical characterization of lymphocyte regulatory molecules. II. Purification of a class of rat and human lymphokines.

Rat spleen cells activated in vitro by concanavalin A produce lymphokine molecules that possess biologic activity in a number of murine lymphocyte response assays. A single class of lymphokine most adequately described as T cell growth factor (TCGF, Interleukin-2) with a m.w. of 15,000 as estimated from gel filtration studies and with an isoelectric range of 5.4 to 5.6 stimulates i) the growth of established T cell lines in culture, ii) the proliferation of thymocytes in the presence of Con A under culture conditions where Con A alone is non-mitogenic, iii) the induction of antibody responses to heterologous erythrocyte antigens in athymic (nude) mouse spleen cell cultures, and iv) the generation of cytotoxic T lymphocytes (CTL) in thymocyte cultures and in nude mouse spleen cell cultures. We suggest that in each of the assay systems tested, this class of rat lymphokine acts directly on activated T cells. Nonactivated T cells must be stimulated by either mitogen or antigen before becoming responsive to lymphokine, but do not require antigen or mitogen for continued lymphokine-dependent proliferation. Similarly, human peripheral blood mononuclear cells activated by phytohemagglutinin (PHA) produce a class of lymphokines of identical size with an isoelectric point of 6.0 to 6.5 that possess the same biologic properties as measured in murine lymphocyte response systems.

Animals↗
PubMed1980

Biochemical and biologic characterization of lymphocyte regulatory molecules. III. The isolation and phenotypic characterization of Interleukin-2 producing T cell lymphomas.

To isolate a stable tumor cell line source of Interleukin 2 (IL-2 formerly referred to as T cell growth factor), over 40 murine leukemia and lymphoma cells as well as 9 clonal helper and killer IL-2-driven T cell lines were screened for both constitutive and mitogen-stimulated IL-2 production. A radiation-induced splenic lymphoma from the B10.BR mouse, the LBRM-33 cell line, could be stimulated to produce over 1000 units/ml of IL-2 after 24 hr exposure to T cell mitogens. Peak IL-2 activity was found in supernatants harvested from 24-hr cultures of either 1% PHA or 20 micrograms/ml Con A-stimulated LBRM-33 cells (10(6) cells/ml). IL-2 production observed in both serum-free and serum-containing cultures represented between 1000 and 5000 times the quantity of IL-2 produced in conventional cultures of mitogen-activated rat or mouse spleen cells. Peak IL-2 production by LBRM-33 cultures (stimulated at either optimal Con A or PHA concentrations or co-stimulated with suboptimal amounts of mitogen and phorbol myristate acetate) was consistently accompanied by LBRM-33 cell death. Phenotypic characterization of the producer cell revealed LBRM-33 cells to be Thy 1+, Ly 1+, Ly 2+, Ly 3+, Qa 2-3+, Qa 3.2+, Qat 4+, and Ly 5+. These studies provide further evidence that IL-2 is a T cell product and establish a source of IL-2 that will be a valuable reagent for the isolation and further molecular characterization of this immunoregulatory molecule.

Animals↗
PubMed1980

Biochemical and biologic characterization of lymphocyte regulatory molecules. IV. Purification of Interleukin 2 from a murine T cell lymphoma.

A cloned murine T cell line, LBRM-33 5A4, growing in culture can be activated by phytohemagglutinin (PHA) to secrete Interleukin 2 (IL-2) activity. In the absence of PHA, LBRM cells do not produce IL-2. The chemical and biologic properties of LBRM-derived IL-2 are compared to those of murine spleen-derived IL-2. The fractionation of culture supernatants from PHA-activated LBRM cells by ion-exchange chromatography, gel filtration, and isoelectric focusing (IEF) reveals that IL-2 activity is contained in a class of molecules indistinguishable in size (30,000) and charge (pI 4.3-4.7 and 4.9-5.1) from murine spleen IL-2. LBRM-derived IL-2 also exhibits the entire spectrum of biologic activities exhibited by spleen IL-2, stimulating the growth of established T cell lines in culture, the generation of cytotoxic T lymphocytes in thymocyte cultures and the induction of antibody responses in nude spleen cultures. Both LBRM and splenic-derived IL-2 activities exhibit similar properties after digestion with proteases, and various chemical and temperature treatments. Since IL-2 is derived from a cloned cell line, it is likely these lymphocyte assays are detecting a single class of molecules that act directly on activated T cells. The use of LBRM cells as a source of IL-2 should greatly facilitate studies on the molecular characterization and biologic activity of this class of lymphocyte regulatory molecules.

Animals↗
PubMed1979

Biochemical and biological characterization of lymphocyte regulatory molecules. I. Purification of a class of murine lymphokines.

Murine spleen cells activated by concanavalin A (Con A) in culture produce a class of lymphokine molecules which possess biological activity in a number of lymphocyte response assays. Lymphokines with a mol wt of 30,000, as estimated from gel filtration studies, can be resolved into two components which differ by charge, with isoelectric point (pI) values of 4.3 and 4.9, respectively. Both components stimulate (a) the growth of established T-cell lines in culture, (b) the proliferation of thymocytes in the presence of Con A under culture conditions where Con A alone is nonmitogenic, (c) the induction of antibody responses to heterologous erythrocyte antigens in athymic (nude) spleen cultures, (d) the generation of cytotoxic T lymphocytes (CTL) in thymocyte cultures, and (e) the generation of CTL in nude spleen cultures. In each of these culture systems we suggest that the assays are detecting a single class of lymphokine which acts directly on activated T cells. Nonactivated T cells must be stimulated by either antigen or mitogen before becoming responsive to lymphokine, but do not require antigen or mitogen for continued growth with lymphokine. The two molecular species, separable by isoelectric focusing are referred to as the T-cell growth factor (TCGF). A lymphokine, similar in size (30,000 daltons) to TCGF but heterogeneous in charge (pI 3.0--4.0), stimulates immune responses to erythrocyte antigens in T-cell-depleted spleen cultures but has no stimulatory activity in the other lymphocyte assay systems described. The data have been interpreted as showing the two molecular forms of murine TCGF (pI 4.3 and 4.9) are responsible for many of the lymphokine activities described elsewhere as thymocyte mitogenic factor, nonspecific T-cell-replacing factor and killer helper factor or costimulator. The other lymphokine, separable from TCGF by charge, appears to have true T-cell-replacing activity.

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