Biomedical subjects

A M Goes

Publications and source records attributed to A M Goes.

At least 37 records · Page 2Linked to original sources

PubMed1999

Nitric oxide-mediated immune complex-induced prostaglandin E(2) production by peripheral blood mononuclear cells of humans infected with Schistosoma mansoni.

Granuloma reaction around Schistosoma mansoni eggs is the prominent lesion in human schistosomiasis. Studies have suggested the involvement of a series of suppressive mechanisms in the control of this reaction. Using an in vitro model of granuloma formation, we have shown that immune complexes (IC) isolated from sera of chronic intestinal schistosomiasis patients were able to reduce granulomatous reaction developed against soluble egg antigen-conjugated polyacrylamide beads. In this system, the role of the l-arginine-nitric oxide (NO) pathway in the formation of prostaglandin E(2) (PGE(2)) by human peripheral blood mononuclear cells (PBMC) of patients infected with schistosomiasis was investigated using IC. Preincubation of PBMC with IC produced a significant increase of both nitrite and PGE(2) levels in the cell supernatant. This effect was inhibited by coincubation of cells with Nomega-nitro-l-arginine methyl ester (L-NAME), a NO synthase inhibitor, showing that the release of PGE(2) subsequent to IC stimulation was driven by NO. The inhibitory effect of L-NAME on PGE(2) release by IC-treated PBMC was reversed by sodium nitroprusside, a known NO donor. Our results indicate that NO could be an important second signal for the stimulation of PGE(2) production induced by IC in PBMC from human schistosomiasis patients.

Adolescent↗

PubMed1999

Phenotype and cytokine profile of Schistosoma mansoni specific T cell lines and clones derived from schistosomiasis patients with distinct clinical forms.

It is essential to distinguish the role of T lymphocytes on the physiopathology associated to more severe forms of schistosomiasis and on the immunomodulation that evolves in the majority of infected people. In this study, we generated Schistosoma mansoni-specific T cell lines and clones from patients with the acute and chronic (intestinal and hepatosplenic forms) phases of disease, from former ones, and from uninfected individuals sensitized to parasite soluble antigens. T cell lines derived from nontreated acute infected donors were capable of producing IL-4 and IL-5, while cells from treated patients secreted IFN-gamma. Lines from intestinal chronic and antigen-sensitized donors preferentially produced IFN-gamma, while those from hepatosplenic patients secreted all three cytokines. The cytokine analysis of CD4+ T cell clones revealed a Th2/Th0 pattern (clones producing IL-4 and IL-5 and clones producing all three cytokines) for those derived from infected patients, while cells from antigen-sensitized donors exhibited an opposite Th1/Th0 pattern (clones producing IFN-gamma and clones producing all three cytokines). The possible role of these T cell populations on human schistosomiasis mansoni is discussed.

Acute Disease↗

PubMed1999

Evidence for nitric oxide action on in vitro granuloma formation through pivotal changes in MIP-1alpha and IL-10 release in human schistosomiasis.

Nitric oxide (NO) has been implicated, both and paradoxically, as a pro- and anti-inflammatory agent in a wide range of circumstances. It is of common concern that NO can be either up- or downregulated by different inflammatory cytokines. Attempting to assess the contribution of NO to the granulomatous response, we used the in vitro granuloma (IVG) model which consists on a reaction of mononuclear cells around polyacrylamide beads conjugated to antigens. Our assays employed Schistosoma mansoni antigens and human peripheral blood mononuclear cells (PBMC) from schistosomiasis patients. Recently, we have described evidence for a regulatory role of NO, with the aid of an inhibitor of NO synthesis, L-NAME. The addition of L-NAME to IVG cultures elicited an increase on the granuloma formation index. Based on these data we decided to investigate the mechanisms involved in the effects of L-NAME-enhanced granuloma formation. Cytokines and chemokines are involved in inflammatory responses by, particularly the latter, inducing migration and adhesion of leukocytes, which led us on this search for their interactions with NO on granulomatous reaction. We evaluated the cytokine/chemokine-secreting profile of PBMC (treated and not treated with L-NAME) on the IVG reaction in order to investigate how NO could interfere on the release of these soluble mediators. Comparison of cell culture releasing amounts of IL-2, IL-10, TNFalpha, IFNgamma, MIP-1alpha, MCP-1, and RANTES demonstrated that MIP-1alpha had increased levels when NO production was blocked with L-NAME, whereas IL-10 secretion decreased in presence of L-NAME. The other tested cytokines (IL-2, TNFalpha, and IFNgamma) and chemokines (MCP-1 and RANTES) showed no significant differences between the presence or absence of L-NAME. Results obtained in this work suggest that inhibition of NO production could upregulate the IVG reaction on human schistosomiasis through changes in the cytokine/chemokine profile released by PBMC. The mechanisms involved may lead to a MIP-1alpha-increased and IL-10-decreased secretion under our experimental conditions, which could partly account for the previously ascribed IVG-exacerbating action of NO inhibition.

Chemokine CCL3↗

PubMed1999

Nitric oxide and IL-10 production induced by PIII--a fraction of Schistosoma mansoni adult worm antigenic preparation--associated with downregulation of in vitro granuloma formation.

Identification and characterization of Schistosoma mansoni antigens that can provide protective immunity, as well as an investigation of their role in host-parasite interaction, is crucial for understanding the complex immunoregulatory events that modulate granuloma formation in schistosomiasis. Previous work by our laboratory identified a fraction of S. mansoni soluble adult worm antigenic preparation (SWAP), named PIII, obtained by anionic chromatography on fast protein liquid chromatography (FPLC). This fraction was able to elicit significant in vitro cell proliferation and at the same time lower in vitro and in vivo granuloma formation when compared either to SWAP or to soluble egg antigens (SEA). In the present work, we investigate some biological activities of PIII, such as the stimulation of nitric oxide (NO) and cytokine production. Our data demonstrated that SEA, SWAP and specially PIII were able to induce a time-dependent increased NO production during in vitro granuloma reaction. Besides that, PIII evoked increased IL-10 production, but not IL-2 or IFNgamma. Collectively, our results indicate the possibility that the modulation role of PIII on in vitro granuloma might be mediated in part by its ability to induce the higher production, initially of IL-10, and lately of NO.

Animals↗

PubMed1999

In vitro human immune reactivity of fast protein liquid chromatography fractionated Paracoccidioides brasiliensis soluble antigens.

Soluble antigens of Paracoccidioides brasiliensis yeast cells (PbAg) were fractionated in a fast protein liquid chromatography (FPLC) system, using Q-Sepharose anion-exchange resin, in order to characterize antigenic fractions that could elicit cell reactivity and antibody recognition in human paracoccidioidomycosis (PCM). PbAg fractions were eluted by 20 mM Tris-HCl solution (pH 9.6) with an increasing gradient up to 1 M NaCl. The FPLC system was able to resolve 7 fractions, enumerated from 0 to VI, according to the elution on the NaCl gradient. The analysis of each fraction on SDS-PAGE showed that fractions 0 to V were constituted by multiple protein bands with molecular mass ranging from 18 to 114 kDa. Large amounts of nucleic acids were evidenced in fraction VI, as revealed by agarose gel stained with ethidium bromide. Sera from PCM patients presenting different clinical forms contained antibodies that recognized antigens in all fractions with the exception of fraction VI as detected by ELISA. Further studies were designed to investigate the capacity of these fractions to induce cell proliferation. It was demonstrated that fractions III and V (200 and 450 mM NaCl, respectively) stimulated a significant proliferative response of peripheral blood mononuclear cells, while fraction 0 induced the lowest proliferative response among patients with PCM, in either acute, acute treated, or chronic forms.

Acute Disease↗

PubMed1999

Main features of DNA-based immunization vectors.

DNA-based immunization has initiated a new era of vaccine research. One of the main goals of gene vaccine development is the control of the levels of expression in vivo for efficient immunization. Modifying the vector to modulate expression or immunogenicity is of critical importance for the improvement of DNA vaccines. The most frequently used vectors for genetic immunization are plasmids. In this article, we review some of the main elements relevant to their design such as strong promoter/enhancer region, introns, genes encoding antigens of interest from the pathogen (how to choose and modify them), polyadenylation termination sequence, origin of replication for plasmid production in Escherichia coli, antibiotic resistance gene as selectable marker, convenient cloning sites, and the presence of immunostimulatory sequences (ISS) that can be added to the plasmid to enhance adjuvanticity and to activate the immune system. In this review, the specific modifications that can increase overall expression as well as the potential of DNA-based vaccination are also discussed.

Enhancer Elements, Genetic↗

PubMed1999

Immunological properties of gene vaccines delivered by different routes.

Gene vaccines represent a new and promising approach to control infectious diseases, inducing a protective immune response in the appropriate host. Several routes and methods of genetic immunization have been shown to induce antibody production as well as T helper (Th) cell and cytotoxic T lymphocyte activation. However, few studies have compared the nature of the immune responses generated by different gene vaccination delivery systems. In the present study we reviewed some aspects of immunity induced by gene immunization and compared the immune responses produced by intramuscular (i.m.) DNA injection to gene gun-mediated DNA transfer into the skin of BALB/c mice. Using a reporter gene coding for beta-galactosidase, we have demonstrated that i.m. injection raised a predominantly Th1 response with mostly IgG2a anti-beta gal produced, while gene gun immunization induced a mixed Th1/Th2 profile with a balanced production of IgG2a and IgG1 subclasses. Distinct types of immune responses were generated by different methods of gene delivery. These findings have important implications for genetic vaccine design. Firstly, a combination between these two systems may create optimal conditions for the induction of a broad-based immune response. Alternatively, a particular gene vaccine delivery method might be used according to the immune response required for host protection. Here, we describe the characteristics of the immune response induced by gene vaccination and the properties of DNA involved in this process.

Animals↗

PubMed1999

Antigenic stimulation is more efficient than LPS in inducing nitric oxide production by human mononuclear cells on the in vitro granuloma reaction in schistosomiasis.

Nitric oxide (NO) is an extremely important and versatile messenger in biological systems. It has been identified as a cytotoxic factor in the immune system, presenting anti- or pro-inflammatory properties under different circumstances. In murine monocytes and macrophages, stimuli by cytokines or lipopolysaccharide (LPS) are necessary for inducing the immunologic isoform of the enzyme responsible for the high-output production of NO, nitric oxide synthase (iNOS). With respect to human cells, however, LPS seems not to stimulate NO production in the same way. Addressing this issue, we demonstrate here that peripheral blood mononuclear cells (PBMC) obtained from schistosomiasis-infected patients and cultivated with parasite antigens in the in vitro granuloma (IVG) reaction produced more nitrite in the absence of LPS. Thus, LPS-induced nitrite levels are easily detectable, although lower than those detected only with antigenic stimulation. Concomitant addition of LPS and L-N-arginine methyl ester (L-NAME) restored the ability to produce detectable levels of nitrite, which had been lost with L-NAME treatment. In addition, LPS caused a mild decrease of the IVG reaction and its association with L-NAME was responsible for reversal of the L-NAME-exacerbating effect on the IVG reaction. These results show that LPS alone is not as good an NO inducer in human cells as it is in rodent cells or cell lines. Moreover, they provide evidence for interactions between LPS and NO inhibitors that require further investigation.

Animals↗

PubMed1998

Role of nitric oxide on human schistosomiasis mansoni: upregulation of in vitro granuloma formation by N omega-nitro-L-arginine methyl ester.

Over the past decade, nitric oxide has been intensely studied due to its relevance as a widespread intra- and intercellular messenger and as a cytotoxin released during several physiopathological events, including immunological reactions and inflammation. In the present paper, we investigate the effect of inhibition of NO synthesis, using an analogue of L-arginine, N omega-nitro-L-arginine methyl ester (L-NAME), on in vitro granulomatous formation of human peripheral blood mononuclear cells (PBMC) from Schistosoma mansoni-infected individuals. The results demonstrated that human PBMC are capable of in vitro NO production and that inhibition of its production through the addition of L-NAME is responsible for exacerbating granulomatous reaction. This L-NAME-induced granuloma enhancement (ranging from 30 to 65%) was measured using the granuloma index. Furthermore, we observed a general time-dependent increase in NO production during the period of cell culture (21 days) and an inverse relationship between nitrite detection and granuloma reactivity. Collectively, our results point to a possible regulatory role of NO on the development of granulomatous inflammation.

Antigens, Helminth↗

PubMed1998

Down modulation of MHC surface molecules on B cells by suppressive immune complexes obtained from chronic intestinal schistosomiasis patients.

Granulomatous inflammation around parasite eggs is the prominent lesion in human schistosomiasis. Studies have suggested the involvement of a series of suppressive mechanisms in the control of this reaction, such as macrophages, cytokines, idiotypic interactions and immune complexes (IC). The studies examine the role of IC obtained from chronic intestinal schistosomiasis patients (ISP) in the reactivity of peripheral blood mononuclear cells (PBMC). The results have shown that these immune complexes are able to suppress cell reactivity by inducing an increase in the production of soluble mediators such as prostaglandins and IL-10. To gain a better understanding of how this suppression occurs the present study examines the phenotypic pattern of PBMC after immune complex treatment in cell proliferation assays. These data show that cultures including immune complex present a higher percentage of B lymphocytes in which a lower expression of a MHC-class II gene product, HLA-DR was detected. This altered expression of the HLA-DR molecule on B lymphocytes after IC treatment suggests a novel mechanism for the suppression observed, that is, IC might decrease the antigen-presenting function of B lymphocytes.

Antigen-Antibody Complex↗

PubMed1998

Inhibition of protein kinases prevents lymphocyte activation by Schistosoma mansoni antigens and reduces in vitro [correction of in vivo] granuloma reaction.

T-cell activation is regulated by signal transduction events initiated by protein kinases. The role of different protein kinases during stimulation of peripheral blood mononuclear cells (PBMC) from chronic intestinal schistosomiasis patients was evaluated using specific inhibitors of protein kinases. We have assayed their ability to interfere with cell proliferation, in vitro granuloma reaction and calcium mobilization. Taken together, our results suggest that Schistosoma mansoni antigen activation of PBMC involves protein kinases. The results observed could be important in the understanding of the mechanism involved in the immunomodulation of schistosomiasis.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1998

Cytokine profile associated to effector functions of human T cell clones specific for Schistosoma mansoni antigens.

T cell lines and clones specific for Schistosoma mansoni antigens were established to study cellular immunity in human schistosomiasis. Flow cytometric analysis of the clones demonstrated that all of them were of the helper inducer T cell subset (CD3+, CD4+, CD8-), and expressed the alphabeta T cell receptor, besides the IL-2 low affinity receptor CD25. Lymphokine analysis revealed that clones presented Th1, Th2, or either Th0 patterns of secretion. More interestingly, the capability of clones to induce in vitro granuloma reactions seems to be related to the presence of TNF-alpha and the absence of IL-10. In counterpart, IL-10 producer clones did not help in vitro granuloma formation, even in the presence of TNF-alpha.

Animals↗

PubMed1998

Characterization of Schistosoma mansoni specific human T-T hybrids.

In this study, we generated antigen-specific T-T hybrids by the fusion of an established T lymphoma, CEM, with T-cell lines derived from Schistosoma mansoni antigen-stimulated peripheral blood mononuclear cells (PBMC) of schistosomiasis patients. These PBMC were capable of in vitro proliferation and granuloma formation upon proper stimulation with antigens derived from different phases of S. mansoni development. Culture supernatants collected from T-T hybrids were responsible for the stimulation or inhibition of antigen-specific responses of other PBMC, on in vitro proliferation and granuloma reaction assays. Clones derived from a T-T hybrid were characterized as CD4+ CD8- HLA-DR- and expressed the cellular markers CD3 and CD25, especially after antigenic stimulation. Their supernatants, as that from the T-T hybrid line, were capable of modulation of in vitro cellular responses, and IL-2, IL-10, and TNF-alpha were detected upon cellular stimulation with antigen or phytohemagglutinin-A (PHA).

Animals↗

PubMed1998

Schistosomiasis and associated infections.

In hospital-based series viral hepatitis B has been frequently described in association with schistosomiasis whilst in field-based studies the association has not been confirmed. The association between schistosomiasis and Salmonella bacteraemia has been well documented. More recently, acute schistosomiasis has been shown to be a facilitating factor in the genesis of pyogenic liver abscesses caused by Staphylococcus aureus. New evidences indicate an interaction between the acquired immunodeficiency syndrome (AIDS) and schistosomiasis. In this paper, data on the association of schistosomiasis with other infections are updated.

Acquired Immunodeficiency Syndrome↗

PubMed1998

Immunization with PIII, a fraction of Schistosoma mansoni soluble adult worm antigenic preparation, affects nitric oxide production by murine spleen cells.

Nitric oxide (NO) is an important effector molecule involved in immune regulation and defense. NO produced by cytokine-activated macrophages was reported to be cytotoxic against the helminth Schistosoma mansoni. Identification and characterization of S. mansoni antigens that can provide protective immunity is crucial for understanding the complex immunoregulatory events that modulate the immune response in schistosomiasis. It is, then, essential to have available defined, purified parasite antigens. Previous work by our laboratory identified a fraction of S. mansoni soluble adult worm antigenic preparation (SWAP), named PIII, able to elicit significant in vitro cell proliferation and at the same time lower in vitro and in vivo granuloma formation when compared either to SEA (soluble egg antigen) or to SWAP. In the present work we report the effect of different in vivo trials with mice on their spleen cells ability to produce NO. We demonstrate that PIII-immunization is able to significantly increase NO production by spleen cells after in vitro stimulation with LPS. These data suggest a possible role for NO on the protective immunity induced by PIII.

Animals↗

PubMed1998

Protective immunity induced in mice by F8.1 and F8.2 antigens purified from Schistosoma mansoni eggs.

Schistosoma mansoni soluble egg antigens (SEA) were fractionated by isoelectric focusing, resulting in 20 components, characterized by pH, absorbance and protein concentration. The higher absorbance fractions were submitted to electrophoresis, and fraction 8 (F8) presented a specific pattern of bands on its isoelectric point. Protein 3 was observed only on F8, and so, it was utilized to rabbit immunization, in order to evaluate its capacity of inducing protective immunity. IgG antibodies from rabbit anti-F8 serum were coupled to Sepharose, and used to obtain the specific antigen by affinity chromatography. This antigen, submitted to electrophoresis, presented two proteic bands (F8.1 and F8.2), which were transferred to nitrocellulose membrane (PVDF) and sequenciated. The homology of F8.2 to known proteins was determined using the Basic Local Alignment Search Tool program (BLASTp). Significant homologies were obtained for the rabbit cytosolic Ca2+ uptake inhibitor, and for the bird a1-proteinase inhibitor. Immunization of mice with F8.1 and F8.2, in the presence of Corynebacterium parvum and Al(OH)3 as adjuvant, induced a significant protection degree against challenge infection, as observed by the decrease on worm burden recovered from portal system.

Animals↗

PubMed1998

Induction of protective immunity against Schistosoma mansoni infection by antigens purified from PIII, a fraction of adult worm, associated to the downregulation of granuloma formation.

This study was performed in order to define Schistosoma mansoni antigens able to function as modulator agents in BALB/c mice granulomatous hypersensitivity to parasite egg. The antigens P-24, P-35 and P-97 were purified by affinity chromatography from a fraction of S. mansoni adult worm antigenic preparation, denominated PIII, involved in the inhibition of granulomatous response to eggs. Immunization of mice with these antigens, in the presence of Corynebacterium parvum and Al(OH)3 as adjuvant, induced a significant protection degree against challenge infection, as observed by the decrease on worm burden recovered from portal system. In vitro blastogenesis assays revealed that purified antigens were able to induce significant proliferation of spleen cells from S. mansoni-infected mice. This protection was correlated to significant decrease in granuloma size induced by PIII. From these results, we concluded that PIII preparation contains antigens capable of mediating protective anti-parasite immunity and down-regulating granulomatous hypersensitivity to S. mansoni eggs.

Animals↗

PubMed1998

The role of T cell subsets and cytokines in the regulation of intracellular bacterial infection.

Cellular immune responses are a critical part of the host's defense against intracellular bacterial infections. Immunity to Brucella abortus crucially depends on antigen-specific T cell-mediated activation of macrophages, which are the major effectors of cell-mediated killing of this organism. T lymphocytes that proliferate in response to B. abortus were characterized for phenotype and cytokine activity. Human, murine, and bovine T lymphocytes exhibited a type 1 cytokine profile, suggesting an analogous immune response in these different hosts. In vivo protection afforded by a particular cell type is dependent on the antigen presented and the mechanism of antigen presentation. Studies using MHC class I and class II knockout mice infected with B. abortus have demonstrated that protective immunity to brucellosis is especially dependent on CD8+ T cells. To target MHC class I presentation we transfected ex vivo a murine macrophage cell line with B. abortus genes and adoptively transferred them to BALB/c mice. These transgenic macrophage clones induced partial protection in mice against experimental brucellosis. Knowing the cells required for protection, vaccines can be designed to activate the protective T cell subset. Lastly, as a new strategy for priming a specific class I-restricted T cell response in vivo, we used genetic immunization by particle bombardment-mediated gene transfer.

Animals↗