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P Puccetti

Publications and source records attributed to P Puccetti.

At least 73 records · Page 4Linked to original sources
PubMed1993

Natural killer cells do not play a dominant role in CD4+ subset differentiation in Candida albicans-infected mice.

The effects of in vivo administration of monoclonal antibodies against NK-1.1-bearing cells on the early production of gamma interferon (IFN-gamma) in vitro and development of Th1-associated immunity were studied in mice infected with a live vaccine strain of Candida albicans. At 1 and 4 days postinfection, natural killer (NK) cell-enriched fractions from the spleens of antibody-treated mice displayed a dramatic reduction in 5E6+ lymphocytes and negligible anti-YAC-1 cytotoxic activity in vitro. Nevertheless, the frequency of IFN-gamma-producing cells in those fractions was reduced by less than half, on average, by anti-NK-1.1 treatment in vivo. In addition, the antibody-treated and infected mice demonstrated unchanged T helper cell responses, as measured by yeast-specific footpad reactions, resistance to reinfection, occurrence of antibodies of different isotypes, and production in vitro of interleukin-2 (IL-2), IFN-gamma, IL-4, and IL-10 by CD4+ cells. Therefore, although NK cells may contribute to early IFN-gamma production in Candida-vaccinated mice, these cells apparently do not play a dominant role in the qualitative development of yeast-specific T helper responses.

Animals↗
PubMed1992

Immunogenic tumor variants induced by drug treatment of the L5178Y lymphoma: search for serologically defined antigens at the clonal level.

Highly immunogenic tumor variants are generated by in vitro or in vivo treatment of the murine L5178Y lymphoma line with triazene derivatives. Most of these variants express new transplantation- and antibody-defined antigens that previous studies have shown to be closely related. One such 80-kDa protein on the surface of clone-D cells was found to be related to xenotropic MuLV gp70 molecules. To investigate the possible relevance of clone-D data to general properties of immunogenic variants in this tumor model system, polyclonal syngeneic antisera raised to a panel of immunogenic clones (including clone D) of the drug-treated L5178Y lymphoma line were employed in the immunoprecipitation of cell-surface and intrinsically labeled variant cells. In all clones, 1- and 2-dimensional electrophoretic analysis of the immunoprecipitates detected an antigen of approximately 80 kDa, and 35S-labeled 80-kDa molecules could be cross-precipitated from all clones by the panel of clone-specific antisera. In addition, 45- and 30-kDa components were also found in metabolically labeled variant cells. While the surface 80-kDa component was reactive with anti-xenotropic gp70 antibodies, the 30-kDa molecule was removed by anti-gag p30 antibody in sequential immunoprecipitation experiments. These data suggest that expression of aberrant, retrovirus-related proteins is a common finding in immunogenic cells of the drug-treated L5178Y lymphoma line.

Animals↗
PubMed1992

Neutralizing antibody to interleukin 4 induces systemic protection and T helper type 1-associated immunity in murine candidiasis.

An interleukin 4 (IL-4)-specific monoclonal antibody (mAb) was administered to mice infected systemically with the yeast Candida albicans, and the animals were monitored for mortality, development of delayed-type hypersensitivity, production of antibodies of different isotypes, release of IL-2, IL-4, IL-6, and interferon gamma (IFN-gamma) in vitro by splenic CD4+ lymphocytes, and levels of IL-4 and IFN-gamma mRNA in these cells. Neutralization of IL-4 by three weekly injections of mAb in several independent experiments resulted in an overall cure rate of 81% versus 0% of controls. Cure was associated with efficient clearance of the yeast from infected organs and histologic evidence of disease resolution, detection of strong T helper type 1 (Th1) responses, and establishment of long-lasting protective immunity. Soon after infection, and as a result of the first or second injection of mAb, there was a decrease in IL-4 mRNA in CD4+ cells, which was accompanied by an increase in the levels of IFN-gamma-specific transcripts. Our data thus indicate that the production of IL-4 by Th2 cells may limit Th1-associated protective immunity in murine candidiasis.

Animals↗
PubMed1992

Antigen-specific cytolysis of infected cells in murine candidiasis.

Immune L3T4+ and Lyt-2+ lymphocytes play an important role in the acquired resistance of mice to challenge with virulent Candida albicans, and release macrophage-activating cytokines in response to yeast cells in vitro. To determine whether antigen (Ag)-specific cytotoxic T lymphocytes are generated during fungal infection, purified L3T4+ and Lyt-2+ lymphocytes from immunized mice were cultured in the presence of syngeneic accessory cells, Candida Ag, and IL-2. Yeast-infected bone marrow macrophages and peritoneal exudate neutrophils were used as target cells in a standard 51Cr release assay. Ag-specific, MHC-unrestricted lysis of infected macrophages was evident with immune Lyt-2+ cells after 5-10 days in culture. Under the same experimental conditions, the cytotoxic activity of L3T4+ cells was negligible, but its expression could be induced by the addition of anti-CD3 antibody. Culturing immune Lyt-2+ cells for shorter periods of time (1-2 days) resulted in preferential lysis of infected neutrophils. In addition, at limiting effector cell numbers, Ag-specific MHC-restricted lymphocytes with cytotoxic activity to infected macrophages could be identified. We suggest that C. albicans infection stimulates multiple cytotoxic T-cell precursors with varying recognition stringency, which may have an important role in antifungal resistance in vivo.

Animals↗
PubMed1992

O6-methylguanine-DNA methyltransferase activity and induction of novel immunogenicity in murine tumor cells treated with methylating agents.

To investigate the mechanism of the generation of immunogenic tumor variants by mutagenic drugs, murine leukemia cells exhibiting different sensitivity to killing by the alkylator 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) and different ability to repair O6-methyl-guanine in their DNA were treated in vitro with a series of methylating agents, including triazene derivatives, temozolomide, and streptozotocin. At the population level, we found that BCNU-resistant cells (L1210/BCNU) that appeared to be cross-resistant to killing by a dimethyltriazene and expressed high levels of O6-methylguanine-DNA methyltransferase activity (mer+ phenotype) failed to generate highly immunogenic variant sublines on repeated exposure to the methylating agents. In contrast, all cells (L1210) that were susceptible to DNA alkylation damage and deficient in O6-methylguanine repair (mer-) developed immunogenic variant sublines. A noticeable exception was represented by streptozotocin treatment, which was equally effective in mer+ and mer- cells. At the clonal level, a single exposure to streptozotocin or a triazene derivative resulted in a high incidence (33% and 50%, respectively) of immunogenic cell generation in mer- cells only. In mer+ cells, streptozotocin treatment led to a 33% incidence of immunogenic clones only when the cells were concurrently exposed to O6-methylguanine as a free base. The activity of O6-methylguanine-DNA methyltransferase in mer+ cells was greatly reduced by treatment with O6-methylguanine or streptozotocin, and the combination of the two drugs led to enzyme levels similar to those observed in mer- cells. Taken together, these data suggest that the mechanism of O6-alkylation may be operative in the induction of novel tumor-cell antigenicity by methylating agents.

Alkylating Agents↗
PubMed1992

Involvement of the Th1 subset of CD4+ T cells in acquired immunity to mouse infection with Trypanosoma equiperdum.

Heat- or merthiolate-inactivated Trypanosoma equiperdum was administered to recipient mice that were subsequently challenged with viable inocula of the same stabilate. Only mice inoculated with merthiolate-killed parasites were completely protected from a challenge inoculum of 10(3) trypanosomes, an effect that was abolished by prior immunosuppression of mice. Immune sera from protected animals contained high levels of interferon (IFN)-gamma and specific IgG2a antibodies. Spleen cells from these mice produced high amounts of interleukin (IL)-2 and IFN-gamma in vitro in response to specific antigen or concanavalin A, whereas splenocytes from mice receiving heat-killed parasites produced high amounts of IL-6. In contrast, the production of tumor necrosis factor (TNF)-alpha and colony-stimulating activity (CSA) was not significantly different in mice receiving either killed parasite preparation. The protection in immunized mice was associated with the detection of strong delayed-type hypersensitivity (DTH) to T. equiperdum antigens, an effect that could be adoptively transferred onto naive recipients by specifically immune CD4+ lymphocytes. These results suggest that the development of protective immunity in mice to T. equiperdum by our immunization protocol may involve the activity of helper/DTH T cells, particularly those of the Th1 subset.

Animals↗
PubMed1992

Tumor-specific L3T4+ and Lyt-2+ lymphocytes in mice primed to mutagenized cell variants.

We have investigated the tumor-specific reactivity of different T-cell subsets from mice primed with clonal variants of L5178Y and P815 cells treated with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). In both tumor systems, anti-parental tumor immunity and protection against non-immunogenic clones were only induced by vaccinating the hosts with highly immunogenic cell variants, and the effect correlated with the detection of TATA-specific delayed-type hypersensitivity (DTH) reactions. The footpad reaction was transferable with spleen cell populations from immunized mice, and enrichment of splenic lymphocytes in L3T4+ but not Lyt-2+ lymphocytes increased the footpad swelling. Unfractionated spleen cell populations from immunized mice released high amounts of IL-2 and IFN-gamma in vitro in response to parental antigens. Purified L3T4+ and Lyt-2+ lymphocytes also produced IFN-gamma when incubated in vitro with the parental tumors and accessory cells. It is suggested that the mechanisms of anti-parental tumor immunity induced by MNNG-treated variants may be similar to those described previously for triazene-xenogenized L5178Y/DTIC cells, and may involve induction of a tumor-specific DTH reaction and IFN-gamma-mediated stimulation of non-specific tumoricidal effects.

Animals↗
PubMed1992

Modulation of colony-stimulating activity by interleukin 1 in mice: opposing effects of combined treatment with indomethacin or prostaglandin E2.

Following previous observations that interleukin 1 (IL-1) may have both positive and negative effects on the levels of circulating colony-stimulating factors (CSF) in mice, we have investigated the impact of human rIL-1 beta administration on serum concentrations of colony-stimulating activity (CSA, as defined by biossay) and macrophage-specific colony-stimulating factor (CSF-1, measured by RIA). In addition, we have studied the effects of IL-1 administered in conjunction with indomethacin or prostaglandin (PG) E2. Besides confirming the finding that exogenous IL-1 leads to a rapid increase in CSF detection, we obtained evidence that IL-1 may also result in the production of cyclo-oxygenase pathway products that down-regulate the IL-1-induced burst in CSA and CSF-1 levels. While co-treatment of mice with indomethacin led to a further increase in CSF detection, the combined exposure to IL-1 and PGE2 resulted in a significant impairment of the stimulatory activity of IL-1.

Animals↗
PubMed1992

Experimental studies of immunotoxicity of a photosensitizing agent (Photofrin II) in mice.

Immunotoxicity studies have been performed on the photosensitizing agent Photofrin II (PHFR), a porphyrin derivative used in photodynamic therapy. Hybrid CD2F1 (H-2d/H-2d) or inbred C57Bl/6 (H-2b) male mice were injected with graded doses of the agent (from 1.2 to 12 mg/Kg ip) on day -5, -3 and -1 before assays. The animals, or spleen cells collected from them on day 0 with respect to PHFR treatment, were tested for: a) competence of producing GVHD upon cell transfer into allogeneic, immunosuppressed recipients; b) graft response against challenge with allogeneic lymphoma cells; c) delayed-type hypersensitivity (DTH) against sheep red blood cells; d) in vitro response to mitogens; e) NK cell activity; f) in vitro generation of alloreactive cytotoxic T lymphocytes (CTL); g) resistance against the challenge of a sublethal dose of Pseudomonas aeruginosa. Moreover the LD50 of the drug given ip has been determined in male CD2F1 mice. The results show that PHFR, even at the highest doses used, does not affect most of the immunological parameters studied, except for a marginal inhibition of CTL generation and increment in proliferative responses to Con A or LPS. These data along with parallel studies performed by our group on human models in vitro, showing increased susceptibility of PHFR-treated tumors to NK or LAK effector cells, point out that PHFR, in the absence of systemic photoactivation, is essentially non-immunotoxic in vivo and could render tumor cells more susceptible to natural immunity.

Animals↗
PubMed1992

Course of primary candidiasis in T cell-depleted mice infected with attenuated variant cells.

Anti-CD4, anti-CD8, or anti-interferon-gamma (IFN-gamma) antibodies or combinations of them were administered in the early stages of chronic infection of mice with a Candida albicans live vaccine strain, and the animals were monitored for course of primary infection, development of delayed-type hypersensitivity, resistance to reinfection, production of interleukin 2 (IL-2) and IFN-gamma in vitro by splenic lymphocytes, and levels of IL-2 and IFN-gamma transcripts in these cells. CD4+ cell and IFN-gamma depletion resulted in the development of fatal candidiasis by the attenuated yeast vaccine. In contrast, either treatment alone modified the course but not the outcome of primary infection, though each prevented the development of resistance to reinfection. Our data thus indicate that both IFN-gamma and CD4+ cells participate in resistance to primary infection with attenuated yeast cells and are critical in the induction of persistent systemic anticandidal immunity.

Animals↗
PubMed1992

Gamma interferon modifies CD4+ subset expression in murine candidiasis.

A single injection of monoclonal antibody to gamma interferon administered in conjunction with a live Candida albicans yeast cell vaccine resulted in the detection of nonprotective Th2 rather than protective Th1 responses and altered the early expression of interleukin 4 and gamma interferon mRNA in CD4+ cells.

Animals↗
PubMed1991

Intrasplenic immunization for the induction of humoral and cell-mediated immunity to nitrocellulose-bound antigen.

Intrasplenic immunization of mice has been shown to induce both specific humoral and cell-mediated immunity to minute amounts of nitrocellulose-bound antigen, electroblotted from sodium dodecyl sulfate-polyacrylamide gels. The test antigens used were aberrantly expressed molecules immunoprecipitated from the lysate of highly immunogenic ('xenogenized') murine lymphoma cells, derived by mutagenesis from a parental, nonimmunogenic cell line. The stained bands of nitrocellulose blots corresponding to the appropriate molecular weights were cut out and the resulting strips deposited in the spleens of recipient mice on three occasions at 15 day intervals. 2 weeks later, the antibody response in the serum was analyzed using a standard ELISA procedure. Cell-mediated immunity was investigated in vitro in terms of cytotoxic T lymphocyte (CTL) activity to radiolabeled xenogenized tumor target cells. In vivo, the immunized mice were assayed for their ability to mount a delayed-type hypersensitivity (DTH) response following footpad challenge with the xenogenized tumor. Our results confirm previous data that the intrasplenic deposition of minute amounts of protein immobilized on a solid matrix effectively stimulates production of specific antibodies. In addition, our results demonstrate that this procedure may also result in the development of T cell-dependent responses detectable in in vitro and in vivo assays of cell-mediated immunity.

Animals↗
PubMed1991

Candida albicans-specific Ly-2+ lymphocytes with cytolytic activity.

To determine whether antigen (Ag)-specific cytotoxic T lymphocytes are generated during experimental Candida albicans infection, purified L3T4+ and Ly-2+ lymphocytes from immunized mice were cultured in the presence of syngeneic accessory cells, C. albicans Ag, and interleukin 2. Yeast-infected bone marrow macrophages were used as target cells in a standard 51Cr-release assay. Freshly isolated L3T4+ and Ly-2+ lymphocytes failed to lyse either target cell type. However, Ag-specific, major histocompatibility complex (MHC)-unrestricted lysis of infected macrophages was evident with immune Ly-2+ cells after 7-10 days in culture. The cultured cells were greater than 98% Thy-1+, CD3+, L3T4-, Ly-2+, T cell receptor alpha/beta + T cells, and their lytic activity was potentiated by the addition of anti-CD3 monoclonal antibodies. At limiting effector cell numbers, Ag-specific MHC-restricted lymphocytes with cytotoxic activity against infected macrophages could be identified. We suggest that C. albicans infection stimulates multiple cytotoxic cell precursors with varying recognition stringency, which include MHC class I-restricted, Ag-specific cytotoxic T lymphocytes.

Animals↗
PubMed1991

Modulation of circulating colony-stimulating activity in mice: combined effects of IL-1 and bacterial or indomethacin treatment.

We have investigated the effects of interleukin 1 (IL-1) administration on circulating levels of colony-stimulating activity (CSA) in intact or neutropenic mice. Intact or cyclophosphamide-treated mice received human rIL-1 beta according to different regimens, and their sera were assayed for CSA at 4, 24 or 48 h. The results indicated that (1) cyclophosphamide alone significantly increased the level of circulating CSA, (2) administration of IL-1 to intact or neutropenic mice resulted in a biphasic pattern of CSA response, an early burst at 4 h being followed at 24-48 h by a significant decrease. In nongranulocytopenic mice, the combined treatment with IL-1 and bacterial cells also resulted in a biphasic pattern of CSA response. However, when IL-1 was administered in concurrence with the cyclo-oxygenase inhibitor indomethacin, sustained CSA levels could be observed for a prolonged period of time. These data expand upon our previous observations on modulation of CSA by IL-1 in granulocytopenic mice, and further support the concept that IL-1 may have both positive and negative effects on the expression of circulating CSA.

Animals↗
PubMed1991

Th1 and Th2 cytokine secretion patterns in murine candidiasis: association of Th1 responses with acquired resistance.

Two chemically mutagenized agerminative variants of Candida albicans were used to immunize mice against challenge with highly virulent cells of the parent strain. Although both mutants (Vir- 3 and Vir- 13) resulted in nonlethal infection and could be recovered from mouse organs for many days after the intravenous inoculation of 10(7) to 10(6) cells, significant protection to systemic challenge with virulent C. albicans was induced by only one (Vir- 3) of the two variants. Anticandidal resistance in Vir- 3-infected mice was associated with the occurrence in vivo of strong delayed-type hypersensitivity to Candida antigen, detection in vitro of highly fungicidal effector macrophages, and presence in the serum of a large proportion of Candida-reactive antibodies of the immunoglobulin G2a isotype. Bulk cultures of purified CD4+ lymphocytes from mice infected with either mutant were compared for their ability to produce gamma interferon (IFN-gamma), interleukin-2 (IL-2), IL-4, and IL-6 in vitro. After stimulation with specific antigen, CD4+ cells from Vir- 3-immunized mice released large amounts of the Th1-specific cytokines, IFN-gamma and IL-2, at a time when CD4+ cells from Vir- 13-infected mice predominantly secreted the characteristic Th2 cytokines, IL-4 and IL-6. These results were confirmed by quantitative analysis of cytokine-producing Th1 and Th2 cells. In addition, only mice infected with Vir- 3 displayed a high frequency of CD8+ cells with the potential for in vitro lysis of yeast-primed bone marrow macrophages. Purified CD4+ cells from Vir- 3-infected mice, but not a mixture of these cells with CD4+ lymphocytes from mice infected with Vir- 13, could adoptively transfer delayed-type hypersensitivity reactivity onto naive mice. Taken together, these data suggest that both Th1 and Th2 CD4+ lymphocytes may be activated during experimental C. albicans infection in mice.

Animals↗