Biomedical subjects

E A Havell

Publications and source records attributed to E A Havell.

At least 37 records · Page 2Linked to original sources

PubMed1991

Tumor necrosis factor-independent IL-6 production during murine listeriosis.

We report that TNF, IL-6, and IFN-alpha/beta are produced by mice during either sublethal or lethal Listeria monocytogenes infections. The quantities of these cytokines in infected spleens increase and decrease in concordance with bacterial numbers in these organs. While all of these cytokines were present in Listeria-infected spleens, only IL-6 and IFN-alpha/beta were found in the peripheral circulation. Inasmuch as TNF has been reported to be responsible for the production of IL-6 in vivo following the inoculation of a lethal dose of the Gram-negative bacterium, Escherichia coli (Fong et al., 1989. J. Exp. Med. 170: 1627), experiments were undertaken to determine whether IL-6 production elicited by the Gram-positive bacterium, L. monocytogenes, was also TNF-dependent. It was found that the passive immunization of mice with neutralizing antibodies specific for TNF shortly before i.v. injection of a lethal or sublethal Listeria inoculum resulted in the complete neutralization of endogenously produced TNF, and in the progressive multiplication of bacteria in infected organs. It was also found that the anti-TNF IgG treatment resulted in a progressive increase in the amounts of Listeria-induced IL-6 present in spleen and blood, until the death of the host. These findings indicate that Listeria-induced IL-6 production in mice occurs primarily through a TNF-independent pathway, and correlates directly with the severity of the infection.

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PubMed1991

A role for tumor necrosis factor in poly(I:C)-induced hemorrhagic necrosis and T-cell-dependent regression of a murine sarcoma.

It was found that intravenous injection of the synthetic double-stranded ribonucleic acid, polyinosinic-polycytidylic acid [poly(I:C)], which is a well-studied interferon (IFN)-inducing agent, can result in extensive hemorrhagic necrosis of the center of an established murine sarcoma and in subsequent complete regression of the surviving rim of the tumor. The poly(I:C)-induced intratumor hemorrhagic reaction was associated with production of appreciable quantities of tumor necrosis factor (TNF) by the host. Neutralization of TNF by treatment with anti-rTNF immunoglobulin G (IgG) caused substantial inhibition of hemorrhagic necrosis and prevented tumor regression from proceeding. Tumor regression was prevented in all mice by depleting them of CD8+ T cells 1 day before poly(I:C) was given. Taken as a whole, the results indicate that the antitumor action of poly(I:C), like that of endotoxin, is based on its capacity to induce the host to make enough TNF to cause a hemorrhagic reaction extensive enough to reduce the tumor burden to a size capable of being dealt with by an underlying host antitumor immune response.

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PubMed1990

Roles of tumor necrosis factor and macrophages in lipopolysaccharide-induced accumulation of neutrophils in cutaneous air pouches.

The role of tumor necrosis factor (TNF) in macrophage-dependent neutrophil accumulation induced by lipopolysaccharide (LPS) was examined through the use of cutaneous air pouches formed on the backs of mice. To investigate the possibility that TNF functions in LPS-induced neutrophil accumulation, we injected LPS into newly formed air pouches (containing relatively few endogenous macrophages), 48-h-old air pouches (containing large numbers of endogenous macrophages), or newly formed air pouches instilled with 10(6) alveolar macrophages (AM). Six hours after LPS injection, air pouches possessing either AM or endogenous macrophages contained large numbers of neutrophils. Infusion of anti-TNF immunoglobulin G into the air pouches inhibited LPS-induced neutrophil accumulation by 84% in air pouches containing AM and 71% in air pouches containing large numbers of endogenous macrophages. TNF was also capable of including neutrophil accumulation when injected into air pouches containing relatively large numbers of either endogenous or exogenous macrophages but not when injected into air pouches containing small numbers of macrophages. In addition, incubation of AM in vitro with TNF induced the AM to cause neutrophil accumulation upon injection into newly formed air pouches. These results indicate that TNF functions in LPS-induced neutrophil accumulation. Furthermore, the results indicate that TNF functions by enhancing the ability of macrophages to cause neutrophil emigration. This is consistent with the possibility that LPS induces TNF production and that TNF, in turn, induces macrophages to produce cytokines with inflammatory activities.

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PubMed1990

Differential inhibition of lipopolysaccharide-induced phenomena by anti-tumor necrosis factor alpha antibody.

Tumor necrosis factor alpha (TNF alpha) has been implicated as a major mediator of lipopolysaccharide (LPS)-induced phenomena. Administration to mice of a polyclonal, monospecific antibody prepared against recombinant murine TNF alpha abolished detection of LPS-induced TNF alpha activity and significantly reduced levels of LPS-induced colony-stimulating factor but failed to reduce the production of LPS-induced interferon, corticosterone, or LPS-induced hypoglycemia.

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PubMed1989

Evidence that tumor necrosis factor has an important role in antibacterial resistance.

TNF is produced in the spleens of Listeria-infected mice during the first 3 days of a sublethal immunizing infection. The production of Listeria-induced TNF coincides with the time when peak numbers of bacteria are present in the liver and spleen. Evidence suggesting that TNF produced in Listeria-infected organs functions in a T cell-independent resistance mechanism comes from results showing that listeriosis is exacerbated in both T cell-intact mice and T cell-deficient (athymic nude) mice treated with a monospecific anti-murine TNF IgG. Listeriosis is exacerbated, however, only if the anti-TNF IgG is given during the first 3 days of infection, i.e., at the time when TNF is being produced in the spleen. Anti-TNF IgG administered on the first day of infection neutralized all the cytotoxic activity of endogenously produced TNF in the spleen. Additional evidence that TNF plays an important role in antibacterial defenses was obtained by showing that administration of pure murine rTNF protects mice against a normally lethal Listeria challenge given 1 to 24 h later.

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PubMed1989

Glucocorticoid-mediated inhibition of endotoxin-induced intratumor tumor necrosis factor production and tumor hemorrhagic necrosis and regression.

Intravenous injection of 25 micrograms of bacterial endotoxin on day 9 of growth of the SA1 sarcoma results in extensive necrosis of the core of this tumor and in its subsequent complete regression. Tumor hemorrhagic necrosis and regression failed to occur in mice that were given a subcutaneous injection of cortisone acetate or dexamethasone 12 h before being giving endotoxin. Inhibition of tumor hemorrhagic necrosis and regression by glucocorticoids was associated with inhibition of endotoxin-induced intratumor TNF production that normally takes place several h after endotoxin is given. In contrast, glucocorticoids had no effect on the ability of intravenously injected rTNF to cause tumor hemorrhagic necrosis and regression. The results lend further support to the belief that TNF is the predominant mediator of endotoxin-induced hemorrhagic necrosis of established murine tumors, and that hemorrhagic necrosis is a prerequisite for the immunologically mediated regression that follows.

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PubMed1988

The antitumor function of tumor necrosis factor (TNF), I. Therapeutic action of TNF against an established murine sarcoma is indirect, immunologically dependent, and limited by severe toxicity.

The ability of murine recombinant tumor necrosis factor (rTNF) and natural TNF in tumor-necrotizing serum (TNS) to cause regression of the SA1 sarcoma was investigated. We found that to cause regression of a 9-d SA1 sarcoma, near lethal quantities of rTNF and TNS had to be given to the host. However, even at these highly toxic doses, rTNF was not reliable at causing complete tumor regression. On the other hand, both types of TNF were reliable at causing a tumor hemorrhagic reaction that resulted in the destruction of greater than 75% of the tumor's center in 24 h. The TNF-induced hemorrhagic reaction involved the development of numerous petechial hemorrhages in the tumor's vascular bed, which apparently resulted from destruction of the tumor's blood vessels. It was possible to follow the development of the hemorrhagic reaction against time after giving rTNF or TNS by measuring the intratumor extravasation of 51Cr-labeled syngeneic red cells. According to this method, TNF-induced intratumor hemorrhaging was in progress within 1 h of giving TNF and continued for about a 6-h period. However, the hemorrhagic reaction was greatly reduced and complete regression of the rim of the living tumor tissue that survived hemorrhagic necrosis failed to occur, if SA1 sarcoma was growing in T cell-deficient (TXB) mice. This indicates that the TNF-induced hemorrhagic reaction is partly dependent, and the tumor regression that follows is completely dependent on host immunocompetence. This suggests in turn, that rTNF does not directly destroy SA1 tumor cells in vivo, even though it was shown that it can destroy SA1 tumor cells in vitro. This interpretation is supported by the additional findings that rTNF was no more therapeutic against a 3-d (3-mm) SA1 than against a 9-d (8-mm) SA1, and was no more therapeutic when injected directly into the tumor than when injected intravenously. Lastly it was possible to completely inhibit the ability of rTNF and TNS to cause tumor hemorrhagic necrosis and regression by infusing the host with a monospecific, polyvalent anti-rTNF antibody that neutralized the cytotoxic action of rTNF in vitro.

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PubMed1988

The antitumor function of tumor necrosis factor (TNF) II. Analysis of the role of endogenous TNF in endotoxin-induced hemorrhagic necrosis and regression of an established sarcoma.

In agreement with the results of previous studies (1), it was shown that intravenous injection of endotoxin into mice bearing 9-d SA1 sarcoma resulted in a tumor hemorrhagic reaction that rapidly caused necrosis of most of the center of the tumor, and then the complete regression of the rim of living tumor tissue that survived the hemorrhagic reaction. The tumor hemorrhagic reaction was confined to the vascular bed of the tumor, and its rate and extent of development were measured in terms of the intratumor extravasation of 51Cr-labeled syngeneic red cells. The development of the hemorrhagic reaction was associated with the presence in the tumor over a 6-h period of endogenous TNF that was measured in terms of its capacity to kill L929B cells in vitro and identified by its susceptibility to neutralization with a monospecific, polyvalent anti-rTNF antibody. The same antibody was capable in vivo of inhibiting the endotoxin-induced tumor hemorrhagic reaction by only approximately 50%, even when present in the tumor in excess. However, it was capable when given in the same quantity of inhibiting the ability of endotoxin to cause complete tumor regression. The fact that TNF was generated in the tumor during the tumor hemorrhagic reaction, and that infusion of a sufficient quantity of anti-rTNF antibody severely interfered with hemorrhagic necrosis and prevented tumor regression represents convincing evidence that TNF is an essential participant in endotoxin-induced regression of an established SA1 sarcoma. Moreover, because tumor regression, as opposed to hemorrhagic necrosis, failed to occur if the tumor was growing in immunoincompetent mice, but did so if the mice were infused with tumor-sensitized T cells, it can be concluded that an adequate level of T cell-mediated immunity is also an essential requirement for endotoxin-induced tumor regression. The participation of other endotoxin-induced mediators in tumor regression cannot be ruled out.

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PubMed1987

Production of tumor necrosis factor during murine listeriosis.

During a lethal murine infection with the gram-positive bacterium, Listeria monocytogenes, a factor appears in the serum that is capable of lysing certain tumor cells in vitro. The levels of this serum cytotoxic factor increase with the progression of morbidity. Neutralization of Listeria-induced cytotoxic factor activity with a monospecific antiserum to recombinant murine tumor necrosis factor (TNF) revealed that the cytotoxic factor was antigenically indistinguishable from natural TNF present in endotoxin-induced tumor necrosis serum. In contrast to a lethal infection, no TNF activity was detected in sera of mice throughout a sublethal infection. However, shortly after initiation of a sublethal infection, mice acquire a greatly enhanced capacity to produce serum TNF in response to an intravenous injection of endotoxin. Cultures of unfractionated spleen cells from mice with ongoing Listeria infections produced TNF when incubated in the absence, but not in the presence, of antibiotics. However, such antibiotic-treated cultures could be stimulated with endotoxin to produce substantially higher TNF yields than spleen cells of uninfected mice. It is also shown that intravenous infusion of anti-recombinant murine TNF IgG into sublethally infected mice results in increased Listeria proliferation in spleens and livers, and ultimately in death from an overwhelming infection.

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PubMed1986

Monoclonal antibody-mediated inhibition of interferon-gamma-induced macrophage antiviral resistance and surface antigen expression.

A monoclonal antibody (MAb) with specificity for murine interferon-gamma (IFN-gamma) was used as a probe for studying the effect of recombinant IFN-gamma (rIFN-gamma) on antiviral activity, Fc receptor expression, and Ia antigen induction in macrophages. Cultures of C3H/HeJ peritoneal exudate macrophages were used to allow direct comparison of all three functions in the same target cell system. Our data provide two major findings: the efficacy of the MAb is very different depending on whether murine fibroblasts or macrophages are used as the target cell in the antiviral assay, i.e., greater than 20 to 100 times more MAb was required to block antiviral activity in macrophage cultures; and 10 to 50 times more MAb was required to inhibit Fc receptor vs Ia antigen expression in response to rIFN-gamma. These latter findings confirm and extend previous observations, which indicate that the induction pathways of two important differentiation markers by IFN-gamma may be dissociable.

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PubMed1986

Purification and further characterization of an anti-murine interferon-gamma monoclonal neutralizing antibody.

The R4-6A2 rat-mouse hybridoma, which secretes a rat IgG1 monoclonal antibody (MAb) capable of neutralizing murine interferon-gamma (MuIFN-gamma), was grown as ascites in athymic nude (nu/nu) mice. The neutralizing titers of ascitic fluids were 400 times higher than those of supernatants from R4-6A2 cultures. Moreover, the specific activities (MAb) neutralizing units/mg protein) of the ascitic fluids were much greater than those of R4-6A2 culture supernatants. Ascitic fluid MAb was purified by anion-exchange chromatography, with excellent recovery of applied neutralizing activity. The ability of the R4-6A2 anti-MuIFN-gamma MAb to neutralize several different activities of MuIFN-gamma was studied. The MAb neutralized the ability of MuIFN-gamma to inhibit the growth of cells (antiproliferative activity). The ability of MuIFN-gamma to render cells of a heterologous species (rat) resistant to viral replication was also neutralized by this MAb. The MAb-neutralizing titers for MuIFN-gamma antiviral activities, on both homologous (murine L929) and heterologous (rat fibroblast) cells, were inversely proportional to the antiviral activity titers on each cell type.

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PubMed1986

Augmented induction of interferons during Listeria monocytogenes infection.

Mice infected iv with an immunizing dose of the gram-positive bacterium, Listeria monocytogenes, produced circulating interferon (IFN) during the inductive phase of the immune response to Listeria. Listeria infection also dramatically altered the host's responsiveness to IFN-inducing agents. Within 24 hr of infection, mice acquired a 50-fold greater than normal capacity to produce the IFN alpha and/or IFN beta classes (IFN alpha/beta) following iv injection of endotoxin. Serum levels of IFN alpha/beta peaked by 2 hr, after which high levels of IFN gamma were detected in the sera of Listeria-infected mice given the B cell mitogen. Similar studies carried out with the interferon-inducing agent polyinosinic-polycytidylic acid (poly(I).poly(C) revealed that mice infected for 24 hr produced only 4-8 times more IFN alpha/beta than did noninfected mice. Unlike endotoxin, however, poly(I).poly(C) did not induce IFN gamma in Listeria-infected animals.

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PubMed1986

Synthesis and secretion of interferon by murine fibroblasts in response to intracellular Listeria monocytogenes.

Listeria monocytogenes, a gram-positive facultative intracellular bacterium, was shown to be capable of infecting and proliferating in murine embryo fibroblasts. During exponential proliferation, the doubling time of the bacterium was determined to be 2.5 h intracellularly, compared with 25 min extracellularly. Progressive intracellular growth of listeriae ultimately resulted in the destruction of initially infected cells and the spread of infection to neighboring cells. Listeria infection induced fibroblasts to synthesize considerable quantities of an acid-stable interferon that proved to be antigenically indistinguishable from both polyinosinic-polycytidylic acid-induced and virus-induced interferon.

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PubMed1984

Monoclonal antibody to murine gamma interferon inhibits lymphokine-induced antiviral and macrophage tumoricidal activities.

Fusion of rat immune spleen cells with mouse myeloma cells resulted in the formation of a stable hybridoma that secretes monoclonal antibody (MAb) directed against murine gamma interferon ( MuIFN -gamma). This MAb specifically neutralized the antiviral activity of a variety of MuIFN -gamma preparations, including a sample produced by recombinant DNA technologies. In contrast, the antiviral activities of a mixture of MuIFN -alpha plus MuIFN -beta, as well as those of rat or human IFN-gamma, were not neutralized by this antibody. The ability of the MAb to inhibit lymphokine-induced macrophage activation was also tested. It was found that in relation to the quantity of antibody needed to completely neutralize antiviral activity, much higher concentrations of MAb were required to abolish the capacity of lymphokine preparations to induce macrophage tumoricidal activity in vitro. The MAb was also coupled to cyanogen bromide-activated Sepharose beads and used as an immunoadsorbent. By reacting lymphokines with MAb coupled to an insoluble matrix, it was possible to show that this immobilized antibody completely and specifically removed from the lymphokine preparations the ability both to invoke macrophage tumoricidal activity and to mediate antiviral activity.

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PubMed1984

The glycoprotein nature of murine gamma interferon.

Synthesis of murine gamma interferon (MuIFN gamma) by phytohemagglutinin (PHA)-stimulated spleen cells was inhibited in a dose-dependent manner by graded concentrations of tunicamycin or 2-deoxy-glucose, both of which inhibit glycosylation. The homologous (murine) and heterologous (rat) antiviral activities of the MuIFN gamma preparations secreted in the presence of the various concentrations of either glycosylation inhibitor were reduced to similar degrees. MuIFN gamma synthesized in the presence of tunicamycin (tunicamycin-MuIFN gamma) exhibited a lower molecular weight (MW), a lack of binding to immobilized Concanavalin-A (Con A), and different charge properties when compared to MuIFN gamma produced in absence of inhibitor (control-MuIFN gamma). Potent rabbit and rat neutralizing antibodies raised against a control-MuIFN gamma subcomponent, isolated by its specific binding to a Con A affinity column, neutralized the antiviral activity of tunicamycin-MuIFN gamma to the same degree as the immunogen.

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PubMed1983

Two molecular weight species of murine gamma interferon.

Concanavalin A (Con A) affinity chromatography was used to concentrate and partially purify a murine gamma interferon (MuIFN gamma) from large volumes of crude MuIFN gamma (specific activity (SA), 10(4) units/mg of protein) produced by mitogen-induced T lymphocytes. Elution of the MuIFN gamma from the immobilized Con A with alpha-methyl-D-mannoside resulted in a 13-fold purification. Further purification of the Con A-bound MuIFN gamma was accomplished by gel filtration chromatography (SA 10(6.7) units/mg of protein) which also demonstrated the molecular weight (MW) to be 38,000 +/- 3000. Molecular weight determinations by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, under both reducing and nonreducing conditions, revealed that the Con A-bound MuIFN gamma may exist as a dimer, consisting of monomers of 20,500 MW. Specific rabbit antiserum raised against the Con A-bound MuIFN gamma neutralized the antiviral activities of crude, Con A-bound and unbound MuIFN gamma fractions, as well as the 20,500 MW MuIFN gamma.

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PubMed1983

Production and characterization of anti-murine interferon-gamma sera.

A partially purified phytohaemagglutinin (PHA)-induced spleen cell murine interferon-gamma (MuIFN-gamma) proved a potent immunogen in a rabbit and rats. Neutralizing anti-MuIFN-gamma activity was detected in rabbit serum following two biweekly subcutaneous inoculations of MuIFN-gamma emulsified in Freund's complete adjuvant. Further biweekly injections of immunogen in Freund's incomplete adjuvant, greatly elevated the serum anti-MuIFN-gamma neutralizing titer. Inbred DA rats also responded with serum neutralizing antibodies for MuIFN-gamma. Rabbit and rat anti-MuIFN-gamma sera neutralized the antiviral activities of MuIFN-gamma preparations produced by T-cells in response to different polyclonal T-cell mitogens or by sensitized T-cells stimulated with specific antigen(s). In addition, the antiviral activity of a rat IFN-gamma on murine cells was neutralized by rabbit anti-MuIFN-gamma serum, but not by the rat anti-MuIFN-gamma serum.

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PubMed1983

Effect of interferon on the growth of Chlamydia trachomatis in mouse fibroblasts (L cells).

The effect of murine interferon on the growth of the lymphogranuloma venereum biotype of Chlamydia trachomatis (strain 440L) in murine fibroblasts (L cells) was examined. Treatment of infected cell cultures with interferon caused a reduction in the number of inclusion-bearing cells as seen by light and electron microscopy and a decrease in yields of chlamydiae as determined by infectivity assays. Interferon also inhibited cycloheximide-resistant (chlamydia-specific) protein synthesis in infected cells. The interferon effect was dose dependent, with 80 to 90% inhibition occurring at concentrations of greater than 200 IU/ml. The inhibitory effect was neutralized by anti-murine interferon globulin. Interferon did not inactivate extracellular chlamydiae, and both host cell RNA and protein synthesis were required for the development of the interferon-induced antichlamydial state. Inhibition of chlamydial growth by interferon was demonstrable in cells treated 18 h before infection or up to 4 h after infection. Cells infected after interferon was removed exhibited an antichlamydial activity decline which was complete by 30 h after interferon removal. We show that interferon treatment did not affect either entry of chlamydiae into host cells or chlamydial conversion to reticulate bodies but rather caused a reduction in the rate of reticulate body replication.

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