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A Campos-Neto

Publications and source records attributed to A Campos-Neto.

At least 37 records · Page 2Linked to original sources
PubMed1997

Molecular characterization of the heat-inducible LmSTI1 protein of Leishmania major.

We have recently isolated a cDNA encoding the Leishmania major homologue of the yeast stress-inducible protein STI1. Southern blot analyses indicate that this protein is encoded by a single copy gene in L. major and that this gene is highly conserved throughout the Leishmania genus. The STI1 gene is constitutively expressed in both L. major promastigotes and amastigotes however, STI1 transcript levels can be upregulated in promastigotes by a shift in culture temperature from 26 to 37 degrees C. Upregulation of transcript was detectable within 5' of heat shock and continued to increase for a further 8 h before returning to constitutive levels. In addition, biosynthetic incorporation of [35S]methionine followed by immunoprecipitation revealed an increase in the level of nascent STI1 protein synthesized when promastigote cultures were shifted from 26 to 37 degrees C. The L. major STI1 protein and the heat shock proteins Hsp83 and Hsp70 form a salt-sensitive complex in L. major promastigotes as evidenced by co-immunoprecipitation using an antiserum specific for L. major STI1. Furthermore, this complex can be reconstituted in vitro by adding recombinant STI1 containing an amino-terminal histidine tag to promastigote lysate and subsequent purification using metal chelate affinity chromatography.

Amino Acid Sequence↗
PubMed1996

Molecular cloning of a novel protein antigen of Leishmania major that elicits a potent immune response in experimental murine leishmaniasis.

BALB/c mice are highly susceptible to infection with the protozoan parasite Leishmania major. This susceptibility has been attributed, in part, to the expansion of parasite-specific CD4+ Th2 cells that antagonize Th1 responses and promote humoral immunity. In the present study, we have utilized sera from L. major-infected BALB/c mice to screen an L. major amastigote cDNA expression library. One of the clones detected encodes a novel Ag designated as L. major stress-inducible 1 (LmSTI1). LmSTI1 contains six copies of the tetratricopeptide consensus motif and is highly related to a family of stress-inducible proteins that is conserved from yeast to humans. Sera from L. major-infected BALB/c mice have LmSTI1-specific Ab titers in excess of 1:200,000, comprised predominantly of IgG1, IgG2A, and IgG2B isotypes. Recombinant LmSTI1 protein elicited strong proliferative responses from draining lymph node cells of L. major-infected BALB/c mice at both early (10 days) and late (28 days) stages of infection and elicited production of high levels of IFN-gamma and low levels of IL-4. In contrast, soluble leishmanial lysate elicited high levels of IL-4 and low IFN-gamma production. Thus, we have identified an Ag of Leishmania capable of eliciting a mixed cellular response that is skewed toward a Th1 phenotype in susceptible BALB/c mice with advanced infections. In addition, analyses of sera from human patients with cutaneous, visceral, and post-kala azar visceral leishmaniasis indicated that a majority of individuals from all three clinical groups mounted strong humoral responses against LmSTI1.

Amino Acid Sequence↗
PubMed1996

Chronic inflammation caused by lymphotoxin is lymphoid neogenesis.

In presenting a unifying concept for chronic inflammation and lymphoid organogenesis, we suggest that lymphotoxin's (LT, LT-alpha, TNF-beta) crucial role in these processes is pivotal and similar. Chronic inflammatory lesions that developed in the kidney and pancreas at the sites of transgene expression in rat insulin promoter-LT (RIP-LT) mice resembled lymph nodes with regard to cellular composition (T cells, B cells, plasma cells, and antigen-presenting cells), delineated T and B cell areas, primary and secondary follicles, characteristic morphologic and antigenic (ICAM-1, VCAM-1, MAdCAM-1, and PNAd) features of high endothelial venules, and ability to respond to antigen and undergo Ig class switching when obtained from mice immunized with SRBC. The vascular changes, with the exception of PNAd, appear to be the direct consequence of transgene derived LT expression, as they were also observed in RIP-LT mice lacking mature T and B cells. These data show that LT-induced chronic inflammation has the characteristics of organized lymphoid tissue.

Animals↗
PubMed1996

Terminal maturation of resting B cells induced by macrophage factors.

Mouse splenic macrophages from BALB/c nude mice (purified by plastic adherence) or cloned macrophage hybridomas stimulated with jacalin (12.5 micrograms/ml), a D-Gal binding lectin, produce one or more B-cell stimulatory factors which cause splenic B cells from BALB/c or C3H/HeJ mice to secrete immunoglobulin in a polyclonal manner as detected by reverse protein A plaque assays. Jacalin-stimulated macrophage supernatants (JacSup) activate both normal and Percoll gradient-purified small high-density (resting) B cells. Supernatants from total or resting BALB/c spleen cells cultured for 7 days in the presence of JacSup (derived from splenic BALB/c nude mice macrophages) were assayed for immunoglobulin isotypes by ELISA. Resting B cells produce only IgG3 and IgM, whereas total B cells secrete IgG3 and IgM as well as IgG1, IgG2a, IgG2b and IgA. Resting and total B cells from BALB/c nude mice are also stimulated by macrophage supernatants to secrete immunoglobulin, thus indicating that this activity is likely to be T cell independent. Moreover, jacalin-stimulated macrophage supernatants did not induce spleen cells or purified B cells to proliferate. Fractionation of factor-rich supernatants on a Sephacryl S-200 column revealed that the factor activity is located in fractions corresponding to a molecular mass of 25-27 kDa. Taken together, these results suggest that upon the action of a macrophage factor(s) resting B cells undergo terminal differentiation without proliferation in the absence of T cells.

Animals↗
PubMed1995

Cloning and expression of a Leishmania donovani gene instructed by a peptide isolated from major histocompatibility complex class II molecules of infected macrophages.

The studies reported here describe the isolation of peptides from MHC class II molecules of murine macrophages infected with Leishmania donovani, and the use of the derived peptide sequences to rescue the pathogen peptide donor protein. The isolation of the peptides was carried out by comparing the RP HPLC profile of peptides extracted from infected macrophages with the peptides extracted from noninfected cells. Several distinct HPLC peaks unique to infected macrophages were sequenced. One of the peptides that was not homologous to any known protein was used to instruct the designing of an oligonucleotide sense primer that was used in combination with an oligo dT nucleotide (anti-sense primer) to amplify by PCR a DNA fragment from L. donovani cDNA. The amplified DNA fragment was cloned and used as a probe to screen a L. donovani cDNA library. The cloned gene (Ld peptide gene) has an open reading frame of 525 bp and has no homology with any known protein/gene sequence. Northern blot analyses indicated that the Ld peptide/gene is broadly distributed and expressed among species of the Leishmania genus, in both the amastigote and promastigote life cycle forms. Using the pGEX 2T vector, the gene was expressed and the relationship of the purified recombinant protein with L. donovani was confirmed using both antibody and T cell responses from immunized or infected animals. The gene encodes a 23-kD molecule (Ldp 23) associated with the cell surface of L. donovani promastigotes. In addition, T cells purified from the lymph nodes of BALB/c mice immunized with L. donovani or infected with L. major, and from CBA/J mice infected with L. amazonensis were stimulated to proliferate by the recombinant Ldp 23 and produced high levels of IFN-gamma and no IL 4. This observation suggests that the Ldp 23 is an interesting parasite molecule for the studies concerning the host/parasite interaction because the Th1 pattern of cytokine response that it induces is correlated with resistance to Leishmania infections. These results clearly point to an alternative strategy for the purification of proteins useful for the development of both vaccines and immunological diagnostic tools not only against leishmaniasis but also for other diseases caused by intracellular pathogens.

Amino Acid Sequence↗
PubMed1995

Morphological and biochemical changes in a hematopoietic cell line induced by jacalin, a lectin derived from Artocarpus integrifolia.

Treatment of the human erythroleukemia cell line K562 with the galactose-binding lectin, jacalin, results in rapid and profound alterations in the morphology and biochemistry of the cells. Within minutes of lectin addition, the cells adhere to the plastic tissue culture surface, and within hours, the cells spread on the surface, acquiring a monocyte-like appearance. Jacalin treatment results in elevated expression of CD61 (integrin beta 3) and CD14, a monocyte-associated cell surface antigen. These results suggest that jacalin treatment of K562 cells triggers intracellular events that result in differentiation along the monocyte lineage.

Amino Acid Sequence↗
PubMed1995

Potent stimulation of murine B cells to proliferate and to secrete immunoglobulins by a lipoteichoic acid-like molecule produced by Clostridium botulinum C and D.

Bacterial products have served as important immunological tools to study lymphocyte activation. The lipopolysaccharides of the Gram-negative bacteria are well known to be potent activators of B lymphocytes. Several Gram-positive bacteria produce exotoxins that are superantigens for T cells. In the present study, we demonstrate that the Gram-positive bacteria Clostridium botulinum C and D produce a high molecular weight mitogen (Cb mitogen) that is a potent activator of murine B lymphocytes. The Cb mitogen was discovered as a consequence of our attempt to investigate a possible superantigen activity present in the botulinum exotoxins. We observed initially that mouse spleen cells were strongly stimulated to proliferate by culture supernatants of C. botulinum C and D. However, the characterization of the responding cell ruled out superantigen because only the B lymphocytes were stimulated to proliferate and to secrete immunoglobulins, and they did so independent of T cell help. In addition, the molecular characterization of the Cb mitogen demonstrated that the purified botulinum toxin was devoid of mitogenic activity. In contrast, the fractionation of the culture supernatant of C. botulinum C in an FPLC Superose 12 column indicated that the Cb mitogen was present in the void volume of the column (MW > or = 300 kDa) which had no toxigenic activity. However, the fractions containing molecules of 150 kDa were highly toxic for mice and had no mitogenic activity. The possibility that LPS was present as a contaminant in the Cb mitogen preparations was excluded because spleen cells from the LPS non-responder C3H/HeJ mice responded well to the Cb mitogen, and the antibiotic polymyxin B, which is an inhibitor of LPS, had no effect on the Cb-mitogen activity. However, an anti-lipoteichoic acid monoclonal antibody (3-1 mAb) inhibited to a great extent the proliferation of spleen cells induced by the Cb mitogen but had no effect on the LPS or concanavalin A stimulation of these cells. Moreover, the Cb mitogen was specifically adsorbed and eluted from a protein G Sepharose column to which the anti-lipoteichoic acid 3-1 mAb had been conjugated. These results support the view that lipoteichoic acid is a selective B cell mitogen.

Animals↗
PubMed1995

Immune evasion by Trypanosomatidae: normal aggregated immunoglobulin protects against lysis by the alternative complement pathway.

Immunoglobulin Fc receptors (FcRs), present in Trypanosomatidae pathogenic for mammals, may be a mechanism by which these parasites escape the host immune response. We studied the possible role of these receptors in evasion by the alternative complement pathway. Promastigotes of Leishmania amazonensis and trypsinized trypomastigotes of Trypanosoma cruzi treated with heat-aggregated normal gamma globulin and then incubated with fresh normal guinea pig serum were shown to be more resistant to lysis. When compared to log phase Leishmania promastigotes, this resistance was at least 4.5-fold greater in parasites harvested in the stationary growth phase. EDTA and EGTA plus MgCl2 inhibited the cytotoxic effect of serum, suggesting the participation of the alternative complement pathway. The distribution of FcRs among genera of Trypanosomatidae that are pathogenic, infective or noninfective for mammals and their affinity for mammalian and fowl immunoglobulin were also examined. These receptors are present only in species infective or pathogenic for mammals, a finding that suggests that this structure is essential for the establishment of infection but is not necessarily a virulence factor. Furthermore, the ligand specificity is limited to the immunoglobulin of mammalian but not of fowl origin.

Animals↗
PubMed1992

An activated murine B cell lymphoma line (A-20) produces a factor-like activity which is functionally related to human natural killer cell stimulatory factor.

In the present article we show that supernatants derived from lipopolysaccharide (LPS) or phorbol 12-myristate 13-acetate (PMA)-stimulated A-20 B cell lymphoma are able to induce polyclonal immunoglobulin (Ig) secretion by normal B cells in a T-cell-dependent manner. This activity could be blocked by neutralizing monoclonal antibodies against interferon-gamma, but not by monoclonal antibodies against interleukin (IL)-2, IL-4, IL-5, IL-6, IL-10 and granulocyte macrophage colony-stimulating factor (GM-CSF) or even a polyclonal antibody against tumor necrosis factor (TNF)-alpha. Furthermore, A-20 supernatants induced the production of measurable amounts of interferon-gamma by normal murine spleen cells and activates natural killer (NK) cells. Fractionation of factor-rich supernatants on a Sephacryl S-200 column revealed that the factor activity is located in the fractions corresponding to a molecular mass of 160-150 kDa and 80-70 kDa. The biological activities found in the A-20 supernatant are very similar to the ones described for the recently cloned human IL-12/NK cell stimulatory factor. These results suggest the existence of a murine analogous factor for the human IL-12 produced by A-20 B cell lymphoma.

Animals↗
PubMed1992

A crude extract of Artocarpus integrifolia contains two lectins with distinct biological activities.

The crude extract derived from seeds of Artocarpus integrifolia (jack fruit) contains two fractions with different biological activities for lymphocytes. One fraction is the D-galactose-binding lectin, jacalin, obtained by affinity purification on a D-galactose agarose column. The other, which is a component of the flow-through fraction (FT), is responsible for the mitogenic activity observed with human PBMC and murine spleen cells. In contrast, jacalin inhibits FT- and ConA-induced proliferative activity of human PMBC and murine spleen cells. This inhibition is not due to toxicity, because: (1) jacalin induces significant levels of IL-3/GM-CSF but not of IL-2 and/or IL-4 in murine spleen cells; (2) jacalin does not affect the capacity of these cells to secrete IL-2 or IL-4 as supernatants obtained from spleen cells sequentially stimulated with jacalin and ConA contain IL-2 and/or IL-4 as well as IL-3/GM-CSF. The ligand for the mitogen contained in the FT fraction is D-mannose as determined by sugar inhibition studies.

Animals↗
PubMed1992

Immune complex glomerulonephritis in experimental kala-azar. II: Detection and characterization of parasite antigens and antibodies eluted from kidneys of Leishmania donovani-infected hamsters.

In a previous report analysing kidney sections by immunofluorescence we showed that hamsters infected with L. donovani develop a glomerulonephritis (GN) associated with deposition of hamster immunoglobulins and parasite antigens in the glomeruli. In this study we characterize these immune components eluted from the kidneys. The eluted immunoglobulins showed specificity for L. donovani antigens and hamster immunoglobulins (rheumatoid factor-like activity). The four isotypes IgG1, IgG2, IgA and IgM were detected. Several L. donovani antigens were detected in the renal eluates by Western blot and immunoprecipitation using 125I-labelled eluates. Proteins with mol. wt of 134, 82, 52, 31, and 26 kD were detected by Western blot and proteins with 134, 110, 93, 89 and 48 kD were detected by immunoprecipitation. With the exception of the 134 kD protein which was recognized by both rabbit anti-promastigote and rabbit anti-amastigote sera all the others were recognized only by the anti-amastigote serum. The 134 kD protein was the only one isolated from the kidneys of infected hamster immunocomplexed with IgG and was the only one detected in a promastigote lysate using IgG from L. donovani-infected hamsters.

Animals↗
PubMed1992

Selective inability of spleen antigen presenting cells from Leishmania donovani infected hamsters to mediate specific T cell proliferation to parasite antigens.

Golden hamsters (Mesocricetus auratus) infected with Leishmania donovani develop a disease similar to human kala-azar. There is conspicuous hypergammaglobulinaemia and their T cells do not respond to stimulation by parasite antigens. The impairment of the cellular immune response seems to be restricted to parasite antigens since infected animals are able to develop a T cell response to the mitogen Concanavalin A (Con-A) and, after sensitization, to the antigens keyhole limpet haemocyanin (KLH) and human serum albumin (DNP-HSA). In the present investigations we studied the role played by infected macrophages in the development of the cellular unresponsiveness present in visceral leishmaniasis. Adherent spleen cells from infected hamsters were unable to present L. donovani antigens to antigen specific T cells, however they were able to present KLH. Conversely, T cells from infected animals did not respond to parasite antigens even when these antigens were presented by normal syngeneic macrophages. Interestingly, lymphocytes from inguinal lymph nodes of infected animals sensitized in their foot pad with parasite antigens proliferated well when stimulated in vitro with L. donovani antigens. These results suggest that the defect in the cellular immune response of the L. donovani infected hamsters is a consequence of a selective inability of their antigen presenting cells to process and present parasite antigens to T cells.

Animals↗
PubMed1991

Activation of T and B cells by a crude extract of Artocarpus integrifolia is mediated by a lectin distinct from jacalin.

The biological activities previously described for a crude extract derived from seeds of Artocarpus integrifolia (jack fruit) are shown in the present work to be assigned to two distinct fractions present in this extract. One fraction is the D-galactose binding lectin, jacalin, obtained by affinity purification on a D-galactose Sepharose column. The other fraction is a D-mannose-binding protein which we propose to call 'Artocarpin'. As is well documented, jacalin binds to human IgA1 and is a useful tool for the purification of this immunoglobulin. We show here that the remaining biological activities consisting of the proliferative response of mouse spleen cells and human peripheral blood mononuclear cells and polyclonal activation of human and mouse B cells for the secretion of immunoglobulin are mediated by artocarpin. Artocarpin is unique in its capacity to induce polyclonal activation of B cells in the absence of proliferation. BALB/c nu/nu spleen cells failed to proliferate which indicates that this lectin is a T cell-dependent B cell polyclonal activator.

Animals↗
PubMed1990

Synergistic signal of an anti-hamster T cell monoclonal antibody on antigen-induced T cell proliferation.

1. The use of monoclonal antibodies (mAb) has permitted the identification of T cell surface antigens and the classification of these antigens based upon phenotype and function. Some of these monoclonal antibodies can identify antigens specifically involved in T lymphocyte activation and are also able to induce, under certain conditions, T cell proliferation. 2. We describe a new mAb raised against hamster T cells which binds to a 45-kDa cell surface antigen expressed on 45% of thymic cells and 90% of mature T lymphocytes. This mAb, designated X2VA, alone does not cause T cell proliferation, but increases in a synergistic manner T cell proliferation when these cells are cultured in the presence of specific antigens, or used in mixed lymphocyte reactions. 3. When the X2VA mAb is used as a single signal for the T cells it induces the production of a T cell growth factor, suggesting that the synergist effect observed during antigen-induced T cell proliferation is mediated by one or more cytokines. 4. Our results indicate that the X2VA mAb recognizes an antigen which is expressed during T cell ontogenesis and which is involved in hamster T cell activation.

Animals↗
PubMed1988

Production of a monoclonal anti-hamster thymocyte antibody with mitogenic properties.

A monoclonal antibody (MAb), designated H3, with specificity for hamster lymphocytes, was produced by somatic cell hybridization of myeloma Sp 2/0 and spleen cells of Balb/c mice immunized with suspensions of viable hamster thymocytes. The H3 MAb (IgG 3) reacted specifically with hamster thymocyte surface membranes (immunofluorescent assay). The antibody recognized a protein of an approximate molecular weight of 44,000 Daltons in immunoblots of hamster thymocyte extracts. The soluble H3 MAb presented potent mitogenic properties as indicated by the DNA synthesis induced in in vitro hamster lymphocyte cultures.

Animals↗
PubMed1987

Immune complex glomerulonephritis in experimental kala-azar.

In the present work we demonstrate that hamsters infected with L. donovani eliminate large quantities of immunoglobulins in the urine. This alteration is clearly a consequence of a conspicuous immune complex glomerulonephritis readily detectable 7 days after the beginning of infection. L. donovani antigens and hamsters immunoglobulins (Igs) were revealed as granular deposits in the mesangial areas and contiguous loops of the glomeruli. Histopathological alterations such as focal mesangial proliferation with progression to diffuse proliferation were observed in the first 3 weeks of infection. From day 35 onwards, all diseased animals presented large deposits of amyloid material of predominantly glomerular localization. In consonance with these alterations, Igs were detected in the urine by day 21 of infection and their concentration increased substantially with the progression of disease. In contrast, serum Igs increased until day 21, when their concentration dropped steadily.

Amyloid↗
PubMed1986

Evaluation of humoral and cellular immunity in patients with urinary tract infections.

The cellular and humoral immunity of 15 patients with recurrent urinary tract infections was assessed during infection. We determined the reactivity of peripheral blood lymphocytes to microbial antigens (PPD, streptolysin O, tetanus toxoid, Salmonella typhi and Escherichia coli), to allogenic cells, and to mitogens (phytohemagglutinin, concanavalin A, pokeweed mitogen and jacalin). T and B cells were counted by the rosette technique. We also evaluated serum immunoglobulins and C3 and C4 levels by radial immunoprecipitation and specific antibodies for the etiological agent by direct agglutination, hemagglutination and indirect immunofluorescence. Urinary antibodies were assessed by the antibody-coated bacteria test (ACB). Some patients were retested for cellular immune function after cure. No significant alteration of immunoglobulin levels was detected, except for two patients who showed elevated IgM levels. C3 and C4 were elevated in one and in five cases, respectively. Specific serum antibodies for the etiological agent were present in all patients but their levels did not differ from those of normal controls. In only five patients were urinary antibodies detected by the ACB test. Patients' peripheral blood lymphocyte reactivity was depressed during infection for most of the microbial antigens and mitogens tested and returned to normal levels after cure. T and B cell numbers were within the normal range.

Adolescent↗