Biomedical subjects

V Ferreira

Publications and source records attributed to V Ferreira.

At least 37 records · Page 2Linked to original sources

PubMed1998

Discrimination between RelA and RelB transcriptional regulation by a dominant negative mutant of IkappaBalpha.

RelA and RelB belong to the nuclear factor-kappaB (NF-kappaB-Rel) transcription factor family. Both proteins are structurally and functionally related, but their intracellular and tissue distributions are different. In resting cells, RelB is found mostly in the nucleus, whereas RelA is sequestered in the cytosol by protein inhibitors, among which IkappaBalpha is the dominant form in lymphocytes. Upon cellular activation IkappaBalpha is proteolyzed, allowing RelA dimers to enter the nucleus and activate target genes. To study the selectivity of gene regulation by RelA and RelB, we generated T cell lines stably expressing a dominant negative mutant of IkappaBalpha. We show that selective inhibition of RelA-NF-kappaB decreased induction of NFKB1, interleukin-2, and interleukin-2Ralpha genes but not c-myc. Transcription driven by the IkappaBalpha promoter was blocked by the transgenic IkappaBalpha; however, wild type IkappaBalpha was expressed in the transgenic cell clones but with much slower kinetics than that in control cells. Wild type IkappaBalpha expression was concomitant with RelB up-regulation, suggesting that RelB could be involved in transcription of IkappaBalpha through binding to an alternative site. These results indicate that RelB and RelA have both distinct and overlapping effects on gene expression.

Animals↗

PubMed1997

Nuclear Rel-A and c-Rel protein complexes are differentially distributed within human thymocytes.

Nuclear factor-kappa B (NF-kappa B)/Rel proteins are inducible transcriptional regulators of numerous cellular genes. They are particularly abundant in lymphoid tissues and are thought to be critical for the transcription of genes involved in immune and inflammatory responses. We have reported previously that a nuclear NF-kappa B activity was present in freshly extracted human thymocytes in the absence of in vitro treatment of these cells. In the present report, we identified NF-kappa B proteins extracted from human thymocyte nuclei as being p50/p65 and p50/c-Rel complexes. Immunochemical and immunofluorescent staining of thymus sections using specific Abs allowed visualization of nuclear NF-kappa B proteins in both thymocytes and nonthymocyte cells. This detection suggested a preferential activation of p50/c-Rel in medullary thymocytes, whereas p50/p65 was present in both cortical and medullary regions of human thymus lobules. However, the intensity of p65 labeling was much higher in several thymocytes from the medulla. p65, p50, and c-Rel activities were found in both CD4- and CD8-positive thymocytes. These observations suggest that p65 and c-Rel complexes play distinct roles in gene expression and that both forms of NF-kappa B play critical roles during late stages of the intrathymic maturation of T cells.

Cell Nucleus↗

PubMed1996

Abnormal blood vessel development and lethality in embryos lacking a single VEGF allele.

The endothelial cell-specific vascular endothelial growth factor (VEGF) and its cellular receptors Flt-1 and Flk-1 have been implicated in the formation of the embryonic vasculature. This is suggested by their colocalized expression during embryogenesis and the impaired vessel formation in Flk-1 and Flt-1 deficient embryos. However, because Flt-1 also binds placental growth factor, a VEGF homologue, the precise role of VEGF was unknown. Here we report that formation of blood vessels was abnormal, but not abolished, in heterozygous VEGF-deficient (VEGF+/-) embryos, generated by aggregation of embryonic stem (ES) cells with tetraploid embryos (T-ES) and even more impaired in homozygous VEGF-deficient (VEGF-/-) T-ES embryos, resulting in death at mid-gestation. Similar phenotypes were observed in F1-VEGF+/- embryos, generated by germline transmission. We believe that this heterozygous lethal phenotype, which differs from the homozygous lethality in VEGF-receptor-deficient embryos, is unprecedented for a targeted autosomal gene inactivation, and is indicative of a tight dose-dependent regulation of embryonic vessel development by VEGF.

Alleles↗

PubMed1996

Leishmania amazonensis: the Asian rhesus macaques (Macaca mulatta) as an experimental model for study of cutaneous leishmaniasis.

As a means of assessing the usefulness of the Rhesus macaque (Macaca mulatta) as a nonhuman primate model for studying cutaneous leishmaniasis, monkeys were infected with Leishmania amazonensis. Variation in the level of susceptibility was found; however, animals inoculated with 10(8) promastigotes provided consistent results as indicated by an earlier onset and/or larger size of lesions. Three monkeys, which had recovered from skin lesions, were challenge-infected using the same parasite strain/dose; although these animals remained susceptible to homologous infection, lesion size was smaller and healed faster than in the initial infection. The immunologic features during infection were assessed. Levels of IgM and IgG antibodies to promastigote antigens rose during active infection and then declined; immunoblot analyses indicated that numerous leishmanial antigens (predominately >30 kDa) were recognized. Delayed type hypersensitivity (DTH) responses and proliferative responses (PBL) developed during active infection and/or rechallenge. Circulating peripheral T cell subpopulations varied throughout the course of infection. Initially (6-8 weeks p.i.), CD4+ T cells appear to predominate; subsequently (15-21 weeks p.i.), an increase in CD8+ T cells was observed. Pathologic analyses indicated that lesions contained amastigotes with a mononuclear infiltrate of macrophages, lymphocytes, and plasma cells, and formation of tuberculoid-type granulomas. As the progression and resolution of leishmanial infection in rhesus macaques are very similar to those observed in humans, this primate model could be employed for elucidating the mechanisms of protective immunity in cutaneous leishmaniasis.

Animals↗

PubMed1996

Immunomodulation of LPS ability to induce the local Shwartzman reaction.

Immunologically, the septic shock is a natural model of immunomediated vascular pathology where the interaction between cytokines and the endothelium mediates the syndrome and lethality. Tumour necrosis factor (TNF), a non-species-specific cytokine, has outstanding pleiotropic activities as an important mediator of the septic shock syndrome. In rabbits, passive immunization with anti-lipopolysaccharide (LPS) polyclonal antibodies prior to the intravenous (i.v.) injection of LPS inhibits the haemorrhagic necrotic lesion characteristic of the local Shwartzman reaction (an excellent localized in vivo correlate of the septic shock). Paradoxically, tested in an ex vivo assay (short-term whole human blood culture, stimulated with LPS), these antibodies mediated an increase in TNF production by mononuclear phagocytes and, in the rabbit model, they induced an increase in body temperature, as compared with the pre-immune reagent. Although anchoring of immune complexes containing LPS to receptors (Fc or C4b-C3b) on circulating monocytes may facilitate the access of LPS to these cells, access to localized, LPS-sensitized macrophages may be impaired. Consequently inhibition of the local Shwartzman reaction and increased TNF production in the ex vivo system were observed. Concordantly, the higher temperature in the passively immunized animals may be a consequence of a higher, immune complex-induced, systemic TNF production. These experimental results suggest that the use of anti-LPS immunoglobulins, as a potential immunotherapy for septic shock syndrome in vertebrates, may lead to increased TNF production, with adverse effects such as the pyrogenic.

Animals↗

PubMed1996

[Hepatic hydatidosis. Review of a series of 677 surgically treated patients].

A retrospective study of 677 patients who underwent surgery for hepatic hidatidosis in the Department of Surgery A and B of the Hospital Miguel Servet in Zaragoza, Spain, over the last 21 years is presented. The frequency was analyzed in regards to sex, age, symptomatology, cyst data, surgical techniques performed and postoperative morbidity. The mean age of the patients was 39 years with the incidence being practically equal in both sexes. Dyspeptic symptoms (60%), hepatomegaly and/or palpable mass in the right hypochondria (58%) and pain (46%) were the most frequent clinical signs and symptoms observed. Radical surgery was performed in 21% of all the surgical treatments, while conservative surgery was undergone in 79%. Solitary cysts were most frequent (65.7%) with localization in the right hepatic lobe (65.4%). The most frequent postoperative complication was biliary fistula (72 cases). The rate of reintervention was 18% and operative mortality 1.6% with a mean hospitalization period of 25 days.

Adolescent↗

PubMed1995

Etiology of acute respiratory tract infections among children in a combined community and hospital study in Rio de Janeiro.

We reviewed data collected between January 1987 and December 1989 on the etiology of acute respiratory infections (ARI) among 827 children in two low-income communities and a hospital in Rio de Janeiro. Respiratory syncytial virus was identified in 38% of cases of ARI, influenza A virus in 1%, parainfluenza 3 virus in 1%, and multiple viruses in 1%. Respiratory syncytial virus was most prevalent among hospitalized children, with seasonal increases in the late fall and winter. The principal bacterial pathogens were Staphylococcus aureus, gram-negative bacteria, Streptococcus pneumoniae, and alpha-hemolytic streptococci. Specimens that were most often positive were pleural fluid (46%) and specimens from other normally sterile sites (24%); normally sterile sites included the CSF, trachea, and lungs. Urine counterimmunoelectrophoresis for S. pneumoniae and Haemophilus influenzae polysaccharide antigens was positive in 3% and 2% of cases, respectively. Pharyngeal cultures yielded low numbers of S. pneumoniae and H. influenzae organisms and higher numbers of gram-negative bacteria. This study demonstrates the high incidence of ARI (4.5 cases per 100 child-weeks) among children in Rio de Janeiro and the high morbidity associated with the illness (ARI is responsible for 25%-50% of all pediatric hospitalizations) and the fact that continued attention must be paid to both viral and bacterial agents of ARI.

Acute Disease↗

PubMed1994

Optimization of the transfection of human endothelial cells by electroporation.

A method for efficient transfection of human umbilical vein endothelial cells (HUVEC) is presented here. The procedure involves cell synchronization followed by electroporation and allows the detection of the activity of low-strength promoters. Sodium butyrate, added to the culture medium after electroporation, strongly potentiates the transcriptional activity of viral promoters. The method can be successfully applied to analyze the transcriptional activity of different promoters linked to reporter genes or to express foreign genes controlled by viral promoters.

Butyrates↗

PubMed1993

Primary structure of the factor VIII binding domain of human, porcine and rabbit von Willebrand factor.

von Willebrand factor (vWF) binds to Factor VIII (FVIII) and the FVIII binding domain has been localized to the amino-terminal of the vWF subunit. The DNA sequence coding for part of the vWF precursor (provWF) including the FVIII binding domain has been compared in man, pig and rabbit. The sequenced fragment corresponds to nucleotides 2416 to 2886 of the human vWF cDNA and encodes for the last 41 amino acids of the propolypeptide, the cleavage site and the first 116 amino acids of the mature vWF subunit. The homology of the three deduced amino acids sequences is remarkable: 88% between porcine and human and 87% between rabbit and human sequences. Four contiguous amino acids are lacking in the rabbit propolypeptide (-10 to -7) when compared to the human and porcine sequences. The cleavage site of the propolypeptide is conserved in the three species as well as amino acids where mutations in the human gene lead to a binding defect of vWF to FVIII. The Asn-94 N-glycosylation site is present in the human and rabbit sequences but absent in the pig sequence.

Amino Acid Sequence↗

PubMed1993

The role of the 5'-flanking region in the cell-specific transcription of the human von Willebrand factor gene.

Transcriptional regulation of the human von Willebrand factor (vWF) gene was investigated in calf pulmonary artery endothelial (CPAE), HeLa, COS 7 and Hep G2 cells. Various lengths of flanking sequences extending up to 2123 bp 5' of the transcription initiation site and containing 19 bp of the first exon, were linked to the bacterial chloramphenicol acetyltransferase (CAT) gene and these constructs were assayed in transient transfection assays. Sequences up to 89 bp upstream of the cap site showed transcriptional activity in all cell types. Sequences between -147 and -419 bp markedly reduced CAT activity in CPAE cells and abolished it in other cell lines. A domain from -592 to -810 bp generated low levels of expression only in CPAE cells. This transcriptional activity was repressed with constructs containing 1041 to 1240 bp upstream of the cap site. Endothelial cell-specific transcription was restored by a construct that contained 1286 bp upstream of the cap site. The additional 46 bp upstream of the negative regulatory domain were within the 5' end of an inverse human Alu-family DNA repeat. RNAase-protection assays confirmed the correct transcriptional initiation. The sequence between -89 and -420 contained at least one negative regulatory element able to repress the CAT gene expression controlled by the heterologous thymidine kinase promoter in all cell types. A construct that included the sequence from -89 to -1286 bp increased the transcriptional activity directed by the thymidine kinase promoter only in CPAE cells. These results indicate that negative and positive elements in the 5'-flanking region interact to regulate vWF gene expression.

Animals↗

PubMed1992

Comparison of the primary structure of the functional domains of human and porcine von Willebrand factor that mediate platelet adhesion.

Porcine von Willebrand factor (vWF) directly aggregates human platelets in vitro indicating a conformational difference between the human and porcine molecules. We amplified and directly sequenced 1242 nucleotides of porcine vWF cDNA that encodes functional domains which mediate the binding of vWF to platelets and subendothelium. The deduced amino acid sequence corresponds to residues 473-891 of the human mature vWF subunit and is 79% homologous with the human protein. Significant differences are found in two discontinuous segments thought to be involved in the binding of vWF to platelet glycoprotein Ib. Porcine vWF lacks four contiguous residues in the first segment and has two positively charged arginine residues in the second. Three point mutations associated with human type IIB von Willebrand disease in the first segment of a botrocetin binding site are at the same position as mismatches between the pig and human. The second segment of the botrocetin site is highly conserved while the third segment shows only a 60% homology.

Amino Acid Sequence↗

PubMed1992

Determination of lead in wines by hydride generation atomic absorption spectrometry.

The optimization of lead hydride generation in aqueous ethanolic media and the influence on its generation of the wine components, both white and red, have been studied. These interferences were overcome by careful control of the parameters affecting hydride generation and the procedure was applied to the determination of Pb in wines. The method is fast, accurate and sensitive and can be used to quantify 24 ppb of Pb in wines.

Lead↗

PubMed1985

Potentialities of immunocompetent cells in high and low antibody-producing lines of mice obtained by selective breedings for responsiveness to flagellar or somatic antigens of Salmonellae.

The genetic modifications of immunocompetent cell functions were investigated in high (H) and low (L) antibody responder lines of mice obtained by selective breeding for responsiveness to flagellar and somatic antigens of Salmonellae (Selection III and Selection IV, respectively). Several lines of evidence converge to demonstrate that the differences in antibody responses between the H and L lines of the two selections are not due to the modification of antigen handling by macrophages. This contrasts with previous observations that macrophages play a major role in interline differences in Selections I and II. The choice of antibody titres after secondary challenge as the phenotypic character in Selections III and IV may explain why the regulatory role of macrophages was minimized, compared with Selections I and II which were carried out for primary responses to heterologous erythrocytes. In Selections III and IV, H mouse lymphocytes were more efficient than L mouse lymphocytes in restoring immunoresponsiveness to irradiated hosts. In contrast, allogeneic skin grafts were rejected at a similar rate in L as well as in H mice of the two Selections and in vitro lymphoproliferative responses to T cell mitogens were also equivalent in the four lines.

Animals↗

PubMed1983

Viral etiology of acute respiratory diseases in Rio de Janeiro: first two years of a longitudinal study.

A two-year study was undertaken to establish the incidence and possible viral etiology of acute respiratory diseases among the child population of a shanty town in Rio de Janeiro, Brazil. The results demonstrated that nearly half of all the illnesses seen were respiratory infections, 10% of them affecting the lower respiratory tract. Viruses were isolated from 20% of the throat swabs collected. Of the viruses identified, 47% were adenoviruses, 25% were enteroviruses, 9% were influenza A, 8% herpes simplex, 7% parainfluenza, 3% respiratory syncytial and 1% influenza B viruses.

Acute Disease↗

PubMed1979