Biomedical subjects

Armando Isibasi

Publications and source records attributed to Armando Isibasi.

10 recordsLinked to original sources

PubMed2006

Salmonella porins induce a sustained, lifelong specific bactericidal antibody memory response.

We examined the ability of porins from Salmonella enterica serovar typhi to induce a long-term antibody response in BALB/c mice. These porins triggered a strong lifelong production of immunoglobulin G (IgG) antibody in the absence of exogenous adjuvant. Analysis of the IgG subclasses produced during this antibody response revealed the presence of the subclasses IgG2b, IgG1, IgG2a and weak IgG3. Despite the high homology of porins, the long-lasting anti-S. typhi porin sera did not cross-react with S. typhimurium. Notably, the antiporin sera showed a sustained lifelong bactericidal-binding activity to the wild-type S. typhi strain, whereas porin-specific antibody titres measured by enzyme-linked immunosorbent assay (ELISA) decreased with time. Because our porin preparations contained the outer membrane proteins C and F (OmpC and OmpF), we evaluated the individual contribution of each porin to the long-lasting antibody response. OmpC and OmpF induced long-lasting antibody titres, measured by ELISA, which were sustained for 300 days. In contrast, although OmpC induced sustained high bactericidal antibody titres for 300 days, postimmunization, the bactericidal antibody titre induced by OmpF was not detected at day 180. These results indicate that OmpC is the main protein responsible for the antibody-mediated memory bactericidal response induced by porins. Taken together, our results show that porins are strong immunogens that confer lifelong specific bactericidal antibody responses in the absence of added adjuvant.

Animals↗

PubMed2005

Expression of triggering receptor on myeloid cell 1 and histocompatibility complex molecules in sepsis and major abdominal surgery.

AIM: To evaluate the surface expression of triggering receptor on myeloid cell 1 (TREM-1), class II major histocompatibility complex molecules (HLA-DR), and the expression of the splicing variant (svTREM-1) of TREM-1 in septic patients and those subjected to major abdominal surgery. METHODS: Using flow cytometry, we examined the surface expression of TREM-1 and HLA-DR in peripheral blood monocytes from 11 septic patients, 7 elective gastrointestinal surgical patients, and 10 healthy volunteers. svTREM-1 levels were analyzed by RT-PCR. RESULTS: Basal expression of TREM-1 and HLA-DR in healthy volunteers was 35.91+/-14.75 MFI and 75.8+/-18.3%, respectively. In septic patients, TREM-1 expression was 59.9+/-23.9 MFI and HLA-DR expression was 44.39+/-20.25%, with a significant difference between healthy and septic groups (P<0.05) for both molecules. In the surgical patients, TREM-1 and HLA-DR expressions were 56.8+/-20.85 MFI and 71+/-13.8% before surgery and 72.65+/-29.92 MFI and 72.82+/-22.55% after surgery. TREM-1 expression was significantly different (P = 0.0087) between the samples before and after surgery and svTREM-1 expression was 0.8590+/-0.1451 MF1, 0.8820+/-0.1460 MF1, and 2.210+/-0.7873 MF1 in the healthy, surgical (after surgery) and septic groups, respectively. There was a significant difference (P = 0.048) in svTREM-1 expression between the healthy and surgical groups and the septic group. CONCLUSION: TREM-1 expression is increased during systemic inflammatory conditions such as sepsis and the postoperative phase. Simultaneous low expression of HLA-DR molecules correlates with the severity of illness and increases susceptibility to infection. Additionally, TREM-1 expression is distinctly different in surgical patients at different stages of the inflammatory response before and after surgery. Thus, surface TREM-1 appears to be an endogenous signal during the course of the inflammatory response. svTREM-1 expression is significantly increased during sepsis, appearing to be an indicator of severity of illness. Together, these data indicate that TREM-1 may play an important role in establishing and amplifying the systemic inflammatory response. TREM-1, HLA-DR, and svTREM-1 expression analysis can provide useful diagnostic and prognostic indicators during SIRS, CARS, and sepsis.

Abdomen↗

PubMed2004

Induction of cellular immune response and anti-Salmonella enterica serovar typhi bactericidal antibodies in healthy volunteers by immunization with a vaccine candidate against typhoid fever.

Typhoid fever remains a serious public health problem. We have developed a vaccine from Salmonella enterica serovar typhi (S. typhi) outer-membrane proteins (OMPs) known as porins. A single subcutaneous dose of 10 microg of porins induced a five-fold (P = 0.05) seroconversion index consisting of IgM and IgG at 7 and 15 days after vaccination as well as the production of IgG1 and IgG2 isotypes. The porins-based vaccine induced a two-fold increase (P = 0.05) in bactericidal titres in volunteers, whom also developed a T-cell response characterized by the production of interferon-gamma (INF-gamma). Side effects after vaccination were mild and transient. The data showed that our S. typhi porins-based candidate vaccine is safe and immunogenic in healthy humans.

Adolescent↗

PubMed2004

Two Salmonella OmpC K(b)-restricted epitopes for CD8+-T-cell recognition.

We report the identification of two peptides from Salmonella OmpC porin that can bind to major histocompatibility complex class I K(b) molecules and are targets of cytotoxic T lymphocytes from Salmonella-infected mice. These peptides are conserved in gram-negative bacterial porins and are the first Salmonella porin-specific epitopes described for possible CD8(+)-T-cell elimination of infected cells.

Animals↗

PubMed2003

Leishmania lipophosphoglycan (LPG) activates NK cells through toll-like receptor-2.

Toll-like receptors (TLRs) mediate the cellular response to conserved molecular patterns shared by microorganisms. We report that TLR-2 on human NK cells is upregulated and stimulated by Leishmania major lipophosphoglycan (LPG), a phosphoglycan belonging to a family of unique Leishmania glycoconjugates. We found that purified L. major LPG upregulates both mRNA and the membrane expression of TLR-2 in NK cells. Additionally, IFN-gamma and TNF-alpha production and nuclear translocation of NF-kappaB was enhanced. The activation effect was more intense with LPG purified from infectious metacyclic parasites than from noninfectious procyclic Leishmania. Since the difference between the molecules derived from these two stages of the parasite growth cycle lies exclusively in the number of phosphosaccharide repeat domains and in the composition of glycan side chains that branch off these domains, we propose that TLR-2 possibly distinguishes between phosphorylated glycan repeats on LPG molecules. The effect of LPG on cytokine production and on membrane expression of TLR-2 could be blocked with F(ab')2 fragments of the mAb against LPG (WIC 79.3). Confocal microscopy demonstrated the co-localization of LPG and TLR-2 on the NK cell membrane. Binding of LPG to TLR-2 in NK cells was demonstrated by immunoprecipitations done with anti-TLR-2 and anti-LPG mAb followed by immunoblotting with anti-LPG and anti-TLR-2, respectively. Both antibodies recognized the immune complexes. These results suggest that NK cells are capable of recognition of, and activation by, Leishmania LPG through TLR-2, enabling them to participate autonomously in the innate immune system and thereby increasing the effective destruction of the parasite.

Animals↗

PubMed2003

Therapeutic efficacy of an E coli strain carrying an ovalbumin allergenic peptide as a fused protein to OMPC in a murine model of allergic airway inflammation.

An Escherichia coli strain expressing the ovalbumin (OVA) 323-329 allergenic peptide on the bacterial surface was evaluated for its ability to reduce the lung inflammatory response in mice allergic to OVA. BALB/c mice were rendered allergic by means of two intraperitoneal injections of OVA suspended in alum 5 days apart, and one intratracheal boost 1 week later. The mice were then treated with two intranasal, 1 week apart, doses of 4x10(9) E. coli-UH302 transformed with plasmids pST13 or pST13-OVA(323-339), which bear the OmpC porin from Salmonella enterica serovar Typhi or the OmpC with the OVA allergenic 323-339 amino acid sequence inserted in the external loop 5. The allergic inflammatory reaction was evaluated on day 31, finding that mice treated with E. coli-UH302-pST13-OVA reduced four to seven times perivascular and peribronchial infiltrates, mucus production, goblet cell hyperplasia and eosinophils when compared with mice treated with E. coli-UH302-pST13 or saline solution. These results were consistent with a significant decrease of IL-5 mRNA and induction of IFN-gamma mRNA in cells from bronchio-alveolar lavages (BAL). Specific serum IgE anti-OVA was also reduced, although the decrease did not reach statistical significance. These results demonstrate that the bacterial live vector bearing an allergenic peptide successfully moderated two important components of allergy, pulmonary inflammation and mucus overproduction.

Allergens↗

PubMed2003

Combinatorial immunoglobulin light chain variability creates sufficient B cell diversity to mount protective antibody responses against pathogen infections.

To analyze how combinatorial light (L) chain diversity influences the B cell repertoire, we studied mice with a homozygous immunoglobulin-heavy-chain null mutation (mu MT), in which the B cell developmental block was overridden by the expression of a transgenic immunoglobulin mu heavy (H) chain derived from a vesicular stomatitis virus Indiana serotype (VSV-IND)-neutralizing Ab (T11 mu MT mice). The randomly integrated transgene could not undergo secondary rearrangements and was expressed in combination with endogenous kappa or lambda chains. T11 mu MT mice had a skewed B cell repertoire as evidenced by 30-60% VSV-IND-specific peripheral B cells and spontaneous VSV-IND-neutralizing serum titers. Upon immunization, T11 mu MT mice mounted specific IgM antibody responses against VSV-IND but, interestingly, they also responded against VSV New Jersey serotype (VSV-NJ), lymphocytic choriomeningitis virus, poliovirus and Salmonella typhi porins. Variable-region sequence analysis revealed that VSV-NJ-specific antibodies expressed numerous L chains in combination with the transgenic H chain, which was devoid of hypermutations. Thus, in T11 mu MT mice combinatorial L chain variability alone is able to build up a sufficiently complex B cell repertoire to mount protective immunoglobulin responses against a variety of pathogens.

Animals↗

PubMed2002

Attenuated Salmonella enterica serovar typhi live vector with inducible chromosomal expression of the T7 RNA polymerase and its evaluation with reporter genes.

Attenuated Salmonella strains with defined gene deletions have been extensively evaluated as suitable live carriers of passenger antigens. A number of strategies for antigen delivery by these strains have been attempted, ranging from plasmid-based to chromosomal integration systems. We report here the chromosomal integration of the T7 RNA polymerase gene (T7pol) in the attenuated strain Salmonella enterica serovar Typhi (Salmonella typhi) CVD908 (aroC(-), aroD(-)). The T7pol gene was amplified by PCR from Escherichia coli BL21(DE3) and cloned in the pNir3 plasmid under the control of the anaerobically inducible nirB promoter. Then it was subcloned in a pKTN701 derivative, suicide plasmid with the R6K ori, and flanked by the aroC gene. After evaluation of its functionality in E. coli SY327, the aroC-T7pol-aroC cassette was integrated into the aroC locus of S. typhi CVD908 by homologous recombination. The resulting strain, S. typhi CVD908-T7pol, was able to transcomplement two plasmids bearing the luc or the lacZ reporter genes controlled by the T7 promoter and produce luciferase and beta-galactosidase under anaerobic culture conditions. Therefore, an inducible system for recombinant antigen production in attenuated S. typhi was achieved.

Bacterial Proteins↗

PubMedDate not supplied

Immunization with a tetramer derivative of an anti-inflammatory pentapeptide produced by Entamoeba histolytica protects gerbils (Meriones unguiculatus) against experimental amoebic abscess of the liver.

Axenically grown Entamoeba histolytica produces a pentapeptide (Met-Gln-Cys-Asn-Ser) with several anti-inflammatory properties, including the inhibition of human monocyte locomotion (Monocyte Locomotion Inhibitory Factor (MLIF)). A construct displays the same effects as the native material. It remains to be seen if MLIF is used, or even produced in vivo by the tissue-invading parasite. If MLIF were to be relevant in invasive amoebiasis, immunizing against it could diminish this parasite advantage and prevent lesions. KLH-linked MLIF mixed with Freund's adjuvant was too aggressive an immunizing material to answer this question. However, immunization with a tetramer of MLIF (but not a scrambled version of MLIF) around a lysine core (MLIF-MAPS), that displays increased antigenicity, yet lacks excessive innate immunity activation, completely protects gerbils against amoebic abscess of the liver caused by the intraportal injection of virulent E. histolytica. Liver abscesses caused by Listeria monocytogenes were not prevented. Invasive E. histolytica may produce the parent protein of MLIF in vivo, and if appropriately cleaved, it may play a role in invasive amoebiasis. MLIF may join new vaccination strategies against amoebiasis.

Animals↗

PubMedDate not supplied

Toll-like receptors: dysregulation in vivo in patients with acute respiratory distress syndrome.

OBJECTIVE: To determine the time course of the expression of five toll-like receptors (TLRs 1-5) in mixed blood mononuclear cells and their relationship to pro-inflammatory and anti-inflammatory cytokines during acute respiratory distress syndrome (ARDS). MATERIAL AND METHODS: In a prospective, a longitudinal study was done at an intensive-care unit of a university-affiliated hospital. Seven consecutive patients with ARDS were studied. We followed the onset and progression of ARDS, and subsequent patient recovery or death, and compared patient data with data from a group of healthy volunteers. We separated mixed blood mononuclear cells using Ficoll-Hypaque to detect the transcripts of human TLRs 1-5. TLR mRNAs were isolated by semiquantitative reverse-transcription PCR (RT-PCR). Each signal was expressed as the ratio of TLR mRNA to beta-actin. Cytokines, including tumor necrosis factor alpha, interferon gamma, and interleukins 4, 6, 8, 10, and 12, were assayed using commercial ELISA kits. RESULTS: Dysregulation in the transcription of TLRs gene, principally in ARDS surviving patients, was observed. Down-regulated expressions of TLR1, TLR4, and TLR5 mRNAs were observed in the first 24 h in patients who survived, and probably played a key role in the survival of patients with ARDS caused by sepsis. Serum levels of cytokines such as IL-6, 8 and 10 were significantly increased in patients with ARDS as compared with levels in the healthy volunteers. However, serum levels of IL-12 were lower in patients with ARDS than in the healthy volunteers. There was difference in serum cytokines concentration between survivors and non-survivors patients with ARDS (except for interleukin-10). CONCLUSIONS: Thus, TLRs gene dysregulation and cytokine profiles are probably important prognostic factors for patient outcome and survival after ARDS.

Aged↗