Biomedical subjects

A R Melo

Publications and source records attributed to A R Melo.

5 recordsLinked to original sources

PubMed2005

Expression of the virulence factor, BfpA, by enteropathogenic Escherichia coli is essential for apoptosis signalling but not for NF-kappaB activation in host cells.

Localized adherence (LA) of enteropathogenic Escherichia coli (EPEC) to epithelial cells results in attaching and effacing of the surface of these cells. LA depends on the gene bfpA, which codes for the BfpA protein. We found that EPEC-E. coli adherence factor (EAF)((+)), expressing BfpA, significantly reduced HeLa cell viability in comparison with EPEC-EAF((-)), as evaluated by the mitochondrial-dependent succinate dehydrogenase conversion of 3'-[4,5,-dimethylthiazol-2yl]2,5-diphenyltetrazolium bromide (MTT) to its formazan. Apoptosis accounts for a substantial loss of the cell viability, because the cells incubated with EPEC-EAF((+)) or with cloned BfpA (data not shown), but not with EPEC-EAF((-)), were positive for annexin-V binding, demonstrated chromatin condensation and nuclei fragmentation and exhibited a high level of caspase-3 activity. Because the blockade of bacterial cell-surface-associated BfpA by anti-BfpA immunoglobulin (Ig)Y antibody suppressed apoptotic death induced by EPEC-EAF((+)), BfpA may be the trigger for apoptosis. Both EPEC-EAF((+)) and EPEC-EAF((-)), as well as recombinant BfpA (data not shown), activated nuclear factor (NF)-kappaB in a similar manner as analysed by the electrophoretic mobility shift assay (EMSA). EMSA supershift analysis demonstrated the presence of p65/RelA in a DNA-binding complex. In contrast to DNA binding, NF-kappaB-dependent reporter gene transactivation was stimulated more strongly by EPEC B171/EAF((+)), suggesting a role for this virulence factor in the regulation of transcriptional activity of NF-kappaB. Because suppression of NF-kappaB activation by BAY11-7085, a NF-kappaB inhibitor, neither induced apoptosis by itself nor blocked apoptosis induction by EPEC-EAF((+)), it may be suggested that apoptosis is not regulated by the NF-kappaB pathway in HeLa cells.

Apoptosis↗

PubMed2001

A single strain of Mycobacterium bovis bacillus Calmette-Guérin (BCG) grown in two different media evokes distinct humoral immune responses in mice.

Two attenuated bacillus Calmette-Guérin (BCG) preparations derived from the same Moreau strain, Copenhagen but grown in Sauton medium containing starch and bacto-peptone (onco BCG, O-BCG), or asparagine (intradermal BCG, ID-BCG), exhibited indistinguishable DNA sequences and bacterial morphology. The number of viable bacilli recovered from spleen, liver and lungs was approximately the same in mice inoculated with the vaccines and was similarly reduced (over 90%) in mice previously immunized with either BCG vaccine. The humoral immune response evoked by the vaccines was, however, distinct. Spleen cell proliferation accompanying the growth of bacilli in tissue was significantly higher in mice inoculated with O-BCG. These cells proliferated in vitro upon challenge with the corresponding BCG extract. Previous cell treatment with mAb anti-CD4 T cells abolished this effect. Anti-BCG antibodies, as assayed either in serum by ELISA or by determining the number of antibody-producing spleen cells by the spot-ELISA method, were significantly higher in mice inoculated with ID-BCG. Anti-BCG antibodies were detected in all immunoglobulin classes, but they were more prevalent in IgG with the following distribution among its isotypes: IgG1>(IgG2a = IgG2b)>IgG3. When some well-characterized Mycobacterium tuberculosis antigens were used as substitutes for BCG extracts in ELISA, although antibodies against the 65-kDa and 96-kDa proteins were detected significantly, antibodies against the 71-kDa, 38-kDa proteins and lipoarabinomannan were only barely detected or even absent. These results indicate that BCG bacilli cultured in Sauton-asparagine medium permitted the multiplication of bacilli, tending to induce a stronger humoral immune response as compared with bacilli grown in Sauton-starch/bacto-peptone-enriched medium.

Animals↗

PubMed1997

Major stress protein Hsp70 interacts with NF-kB regulatory complex in human T-lymphoma cells.

Polypeptides belonging to the Hsp70 major stress protein family and to the NF-kB/Rel multi-functional regulatory complex are known to be involved in cellular defense mechanisms. It was suggested that both systems may interact in cells that respond to injuring stimuli. To check this, Molt4 human lymphoma cells were heated at 43 degrees C for 15 min and, after a 6 h post-shock recovery period, the cells were activated with phorbol ester or bacterial lipopolysaccharide. It was found that mild heat shock caused a substantial increase of the intracellular Hsp70 content with the concomitant suppression of NF-kB complexes, though the latter was properly activated in non-stressed cells. After a 24 h period of being inactive the complex fully recovered its activity and p65 and c-Rel subunits migrated to the nucleus. This new active period lasted even longer than that in non-heated control cells. As this suggested the existence of a Hsp70-related mechanism of NF-kB/Rel complex retention in cytoplasm, we carried out immunoprecipitation with the use of anti-Hsp70 and anti-Rel antibodies. All three Rel family members p65, c-Rel, p50, but not their precursors and IkB alpha inhibitory protein were shown to co-precipitate with the stress protein and anti-Hsp70 antibodies from both heated and non-heated cells. We conclude that the Hsp70 stress protein may confer a new mechanism of NF-kB regulation in cells affected by elevated temperature or other factors related to the cellular response to stress.

Blotting, Western↗

PubMed1990

Controversies in posttraumatic epilepsy.

In civilian accidents the factors involved in the origin of posttraumatic epilepsy are controversial. In this study of 506 consecutive head trauma patients and 101 epileptic patients developing seizures after head trauma, we have examined the importance of the duration of coma, type of seizure and drug therapy, time to first seizure, age and focal lesions and compared our results with the literature. The importance is stressed of focal lesions and of neurological deficits for the origin of posttraumatic seizures.

Adolescent↗

PubMedDate not supplied

[Suppurative thyroiditis with fatal course in diabetics].

The authors report two case studies of suppurative thyroiditis in type II diabetics patients, giving emphasis to the rapid evolution for sepsis, which took one of the patients to death. An earlier diagnosis, antibiotictherapy and a large surgical drainage are essential to avoid a fatal course in these patients. A brief review of suppurative thyroiditis is also presented.

Adult↗