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Biomedical subjects

M Costa

Publications and source records attributed to M Costa.

At least 127 records · Page 7Linked to original sources
PubMed1999

Nickel enhances telomeric silencing in Saccharomyces cerevisiae.

Certain nickel compounds including crystalline nickel sulfide (NiS) and subsulfide (Ni3S2) are potent human and animal carcinogens. In Chinese hamster embryo cells, an X-linked senescence gene was inactivated following nickel-induced DNA methylation. Nickel also induced the inactivation of the gpt reporter gene by chromatin condensation and a DNA methylation process in a transgenic gpt+ Chinese hamster cell line (G12), which is located near a heterochromatic region. To determine if nickel can cause gene silencing independently of DNA methylation, based only on the induction of changes in chromatin structure, we measured its effect on gene silencing in Saccharomyces cerevisiae. Growth of yeast in the presence of nickel chloride repressed a telomeric marker gene (URA3) and resulted in a stable epigenetic switch. This phenomenon was dependent on the number of cell doubling prior to selection and also on the distance of the marker gene from the end of the chromosome. The level of TPE (telomeric position effect) increased linearly with elevations of nickel concentration. Addition of magnesium inhibited this effect, but magnesium did not silence the reporter gene by itself. The level of silencing was also assessed following treatment with other transition metals: cobalt, copper and cadmium. In the sublethal range, cobalt induced similar effects as nickel, while copper and cadmium did not change the basal level of gene expression. Silencing by copper and cadmium were evident only at concentrations of those metals where the viability was very low.

Cell Survival↗
PubMed1999

Tight binding of the 5' exon to domain I of a group II self-splicing intron requires completion of the intron active site.

Group II self-splicing requires the 5' exon to form base pairs with two stretches of intronic sequence (EBS1 and EBS2) which also bind the DNA target during retrotransposition of the intron. We have used dimethyl sulfate modification of bases to obtain footprints of the 5' exon on intron Pl.LSU/2 from the mitochondrion of the alga Pylaiella littoralis, as well as on truncated intron derivatives. Aside from the EBS sites, which are part of the same subdomain (ID) of ribozyme secondary structure, three distant adenines become either less or more sensitive to modification in the presence of the exon. Unexpectedly, one of these adenines in subdomain IC1 is footprinted only in the presence of the distal helix of domain V, which is involved in catalysis. While the loss of that footprint is accompanied by a 100-fold decrease in the affinity for the exon, both protection from modification and efficient binding can be restored by a separate domain V transcript, whose binding results in its own, concise footprint on domains I and III. Possible biological implications of the need for the group II active site to be complete in order to observe high-affinity binding of the 5' exon to domain I are discussed.

Base Sequence↗
PubMed1999

Human DNA helicase VIII: a DNA and RNA helicase corresponding to the G3BP protein, an element of the ras transduction pathway.

Human DNA helicase VIII (HDH VIII) was isolated in the course of a systematic study of the DNA unwinding enzymes present in human cells. From a HeLa cell nuclear extract a protein with an Mrof 68 kDa in SDS-PAGE was isolated, characterised and micro-sequenced. The enzyme shows ATP- and Mg2+-dependent activity is not stimulated by RPA, prefers partially unwound 3'-tailed substrates and moves along the bound strand in the 5' to 3' direction. HDH VIII can also unwind partial RNA/DNA and RNA/RNA duplexes. Microsequencing of the polypeptide showed that this enzyme corresponds to G3BP, an element of the Ras pathway which binds specifically to the GTPase-activating protein. HDH VIII/G3BP is analogous to the heterogeneous nuclear ribonucleoproteins and contains a sequence rich in RGG boxes similar to the C-terminal domain of HDH IV/nucleolin, another DNA and RNA helicase.

Adenosine Triphosphatases↗
PubMed1999

Oxidation of hypotaurine to taurine with photochemically generated singlet oxygen: the effect of azide.

Hypotaurine is oxidized to taurine by singlet oxygen (1O2) generated with methylene blue used as a photosensitizer. The oxidation rate increases in the presence of deuterium oxide as expected for the involvement of 1O2. Addition of the 1O2 quencher azide also produced an activating effect in contrast with the expected inhibition. Azidyl radicals produced by the oxidation of azide by the horseradish peroxidase/hydrogen peroxide system stimulate the oxidation of the added hypotaurine. It is concluded that azide competes with hypotaurine for 1O2 generating the azidyl radical which is a strong one-electron oxidant transfer of the radical to hypotaurine. The hypotaurine radical is then converted into taurine, possibly through the disulfone intermediate. Formation of the sulfonic hydroperoxide is the possible intermediate in the absence of azide. The finding that the azidyl radical efficiently oxidizes hypotaurine to its metabolic product taurine raises the expectation of hypotaurine being a valuable scavenger of endogenous and exogenous radicals.

Azides↗
PubMed1999

Modelling a parasystolic rhythm in a heart-transplant patient.

A parasystole from a heart-transplant patient is analysed using a beat-to-beat RR interval time series obtained from an electrocardiogram (ECG). The dysrhythmia, resulting from the co-existence of two pacemakers, the sinus node and an ectopic focus, presents distinctive regular patterns, with transitions from one pattern to another occurring abruptly. It is shown that the parasystolic rhythm can be simulated by a model involving two oscillators firing at fixed rates, under the restriction that neither is allowed to fire during the other's refractory period. It is found that the structure of the generated RR time series is essentially determined by the ratio of the periods of the two oscillators. In the case of a heart-transplant patient with a small heart-rate variability as a result of heart denervation, the model predicts the RR intervals with an error of less than 6% for an 80-beat sequence. From a physiological point of view, the results imply that the interaction between the two pacemakers in the heart is fairly weak, and hence the parasystole observed in the heart-transplant patient can be modelled as pure parasystole.

Adult↗
PubMed1999

Rapid anterograde and retrograde tracing from mesenteric nerve trunks to the guinea-pig small intestine in vitro.

A novel technique for rapid anterograde labelling of cut axons in vitro was used to visualise the peripheral branches of mesenteric nerve trunks supplying the guinea-pig small intestine. Biotinamide, dissolved in an artificial intracellular solution, was applied to the cut ends of the mesenteric nerves and the tissue was maintained in organ culture overnight. Labelled nerve fibres were visualised by fluorescein isothiocyanate (FITC)-conjugated streptavidin. Intense staining of nerve fibres and terminal varicosities in the ganglia and internodal strands of the myenteric plexus was achieved up to 15 mm from the application site. Filled fibres formed baskets around some myenteric nerve cell bodies, suggesting target-specific neurotransmission. When combined with multiple-labelling immunohistochemistry for tyrosine hydroxylase (TH), calcitonin gene-related protein (CGRP) or choline acetyltransferase (ChAT), most anterogradely labelled nerve fibres, and many pericellular baskets, were found to be TH immunoreactive, indicating their postganglionic sympathetic origin. Double-labelling immunohistochemistry revealed that the postganglionic sympathetic pericellular baskets preferentially surrounded 5-hydroxytryptamine (5-HT)-handling myenteric neurons. Some biotinamide-filled fibres were CGRP immunoreactive, and are likely to originate from spinal sensory neurons. We describe for the first time many pericellular baskets labelled from the mesenteric nerves which were ChAT immunoreactive. Retrogradely filled intestinofugal nerve cell bodies were also observed, all of which had a single axon arising from a small nerve cell body with short filamentous or lamellar dendrites. Many of these cells were ChAT immunoreactive. This in vitro technique is effective in identifying the fine arrangement of nerve terminals arising from nerve trunks in the periphery.

Animals↗
PubMed1999

Thioglycolate-elicited rat macrophages exhibit alterations in incorporation and oxidation of fatty acids.

Incorporation and oxidation of fatty acids (FA) were investigated in resident and thioglycolate-elicited (TG-elicited) rat macrophages (Mphi). Both cell types presented a time-dependent incorporation of [14C]-labeled palmitic acid (PA), oleic acid (OA), linoleic acid (LA), and arachidonic acid (AA) up to 6 h. The total amount of [14C]-FA incorporated by resident Mphi after 6 h was: AA > PA = LA > OA. TG-elicited cells presented a 50% reduction in the incorporation of LA, PA, and AA, whereas that of OA remained unchanged as compared to resident Mphi. The FA were oxidized by resident Mphi as follows: LA > OA > PA > AA. TG elicitation promoted a reduction of 42% in LA oxidation and a marked increase in AA oxidation (280%). The increased oxidation of AA in TG-elicited cells may account for the lower production of prostaglandins in Mphi under these conditions. The full significance of these findings for Mphi function, however, remains to be examined.

Animals↗
PubMed1999

HLA-DM can partially replace the invariant chain for HLA-DR transport and surface expression in transfected endocrine epithelial cells.

The function of HLA class II molecules as peptide presenters to CD4+ T cells depends on the expression of associated molecules such as the invariant chain (Ii) and DM responsible for the correct transport of and high-stability peptide binding to the class II dimers. In organs affected by autoimmune diseases, endocrine epithelial cells express class II molecules, which presumably are involved in the presentation of self-peptides to autoreactive T cells. We have transfected the rat insulinoma cell line RINm5F with different combinations of HLA-DR, Ii and HLA-DM cDNAs and have studied how Ii and DM affect the transport and stability of class II molecules expressed by the different transfectants. Immunofluorescence and biochemical analysis showed that cells transfected with DR and DM in the absence of Ii expressed mostly stable molecules in their surface, and showed a lower accumulation of DR molecules in the endoplasmic reticulum (ER) than cells expressing only DR. This suggests that, in the absence of invariant chain, DM molecules can not only exchange peptides other than class II-associated invariant chain peptide (CLIP) but may also be involved in the transport of class II molecules out of the ER towards the endosomal route. In addition, these data confirm that expression of DR alone or DR+Ii do not allow the formation of sodium dodecyl sulphate (SDS)-stable complexes, that cells expressing DR+Ii have most DR molecules occupied by CLIP and that Ii and DM molecules allow regular routing and peptide loading of class II molecules.

Animals↗
PubMed1999

Projections of nitric oxide synthase and vasoactive intestinal polypeptide-reactive submucosal neurons in the human colon.

BACKGROUND: The submucosal plexus is important in the control of secretomotor and motor function of the intestine. Our aim was to describe the projections of submucosal neurons to the mucosa within the submucosal plexus and to the circular muscle of human colon and to determine whether submucosal neurons that projected to different layers were located at different levels of the submucosa. METHODS: A retrogradely transported fluorescent dye was applied to the mucosa, submucosa or circular muscle layer of human colon which was then maintained in organotypic culture for 5 days. The submucosa was then dissected into two preparations, one containing the inner layer of the submucosal plexus and the other containing both the intermediate and outer layers. The dissected preparations were labelled with antibodies to nitric oxide synthase (NOS) or vasoactive intestinal peptide (VIP). RESULTS: Submucosal neurons projected to the mucosa, submucosa and circular muscle layers for mean distances of 3.7, 3.0 and 4.3 mm, respectively. Ninety-seven per cent of submucosal neurons labelled from the circular muscle were located in the outer or the intermediate layers, while 51% of those projecting to the mucosa were in inner layer and 49% in the intermediate/outer layers of the submucosal plexus. Eleven per cent of submucosal neurons projecting to the circular muscle were immunoreactive for NOS and 12% were immunoreactive for VIP. Forty-five per cent of those projecting within the submucosa were immunoreactive for VIP and 38% of those projecting to the mucosa were immunoreactive for VIP. CONCLUSIONS: Submucosal neurons in the human colon innervate the mucosa, circular muscle and submucosa and different functional classes of neurons are located in different layers of the submucosal plexus.

Adult↗
PubMed1999

Nickel-induced transformation shifts the balance between HIF-1 and p53 transcription factors.

Nickel (Ni) compounds are potent carcinogens and can induce malignant transformation of rodent and human cells. In an attempt to unravel the molecular mechanisms of Ni-induced transformation we investigated transcriptional activity of hypoxia-inducible factor (HIF-1) and p53 tumor suppressor protein in Ni-transformed cells. We demonstrated that the activity of HIF-1-responsive promoters was increased in Ni-transformed rodent cells resulting in the increased ratio between HIF-1- and p53-stimulated transcription. To further elucidate the roles of HIF-1 and p53 in Ni-induced transformation we used human osteosarcoma (HOS) cells and a Ni-transformed derivative, SA-8 cells. Since non-functional p53 was expressed in both HOS and SA-8 cells, acute Ni treatment induced HIF-1alpha protein and HIF-1-dependent transcription without affecting p53. In MCF-7 and A549, human cancer cells with the wild-type p53, both functional p53 and HIF-1alpha proteins accumulated following exposure to Ni. The induction of HIF-1alpha and wild-type p53 by Ni was detected after 6 h and was most pronounced by 24 h. These results suggest that acute Ni treatment causes accumulation of HIF-1alpha protein and simultaneous accumulation of wild-type, but not mutant, p53. We suggest that the induction of hypoxia-like conditions in Ni-treated cells with subsequent selection for increased HIF-1-dependent transcription is involved in Ni-induced carcinogenesis.

3T3 Cells↗
PubMed1999

Who in Brazil has a personal doctor?

BACKGROUND: Continued medical care (including having a personal doctor) is regarded as an essential aspect of a good health service. OBJECTIVES: The objectives of the present study were to investigate the reasons for not having a personal doctor, and the satisfaction with the care received by patients with and without a personal doctor. METHODS: We conducted a cross-sectional study with data collected during 20 days over 6 months in the Emergency Service of the Conceição Hospital, the busiest emergency service in Porto Alegre. The subjects were 553 patients selected through systematic random sampling. The main outcome measure was having a personal doctor. Patients who reported usually to see the same doctor and remembered their physician's name were regarded as having a personal doctor. RESULTS: Patients who usually use primary care service represented 23% of the sample, and were four times more likely to have a personal doctor (OR = 3.83, CI 95% = 2.41-6.11). Independent, statistically significant variables associated with having a personal physician were: usually receiving care from a primary health care service (OR = 3.8, CI 95% = 2.39-6.00) and from a physician in the private sector (OR = 2.16, CI 95% = 1.15-4.00). Patients who had a personal doctor reported higher satisfaction with their access to health care. The personal doctors' specialties were: internal medicine (37%), cardiologist (17%), gynaecologist-obstetrician (13%), family physician (8%) and pneumologist (6%). CONCLUSIONS: For patients who attend emergency services in Brazil, primary health care and private medical care provide better access to continuity of patient care. Patients with personal doctors report higher satisfaction with access to consultations.

Adult↗
PubMed1999

No evidence of chaos in the heart rate variability of normal and cardiac transplant human subjects.

INTRODUCTION: The variability observed in the heart rate may reflect fundamental aspects of cardiac activity. It has been under discussion whether heart rate variability (HRV) is due to noise or chaos, which is irregular behavior occurring in deterministic nonlinear systems. METHODS AND RESULTS: Using chaos analysis techniques, we analyzed HRV of five normal and five human cardiac transplant subjects at rest. HRV is studied using the beat-to-beat RR interval time series extracted from the ECG. The cardiac transplant subjects exhibited a much smaller HRV than the normal subjects because of heart denervation. We present the map and correlation dimension estimation for the RR time series. To test for nonlinear correlations in the dynamics, we built surrogate time series that have the same power spectra as the experimental time series, but also have randomized phases. The experimental and the surrogate data were compared using the correlation integral. No correlation dimension was found for the RR time series of either the normal or the cardiac transplant subjects. Nevertheless, nonlinear correlations were detected in the HRV of the normal subjects but not in HRV of the cardiac transplant subjects. For the latter, no significant changes were observed in the correlation integral as a function of time after transplantation. CONCLUSION: We found no evidence of low-dimensional chaos in the HRV of normal and cardiac transplant subjects. However, some nonlinear correlations were detected in the HRV of the normal subjects, which may be associated with autonomic nervous system influence.

Electrophysiology↗
PubMed1999

5-azacytidine induces transgene silencing by DNA methylation in Chinese hamster cells.

The cytosine analog 5-azacytidine (5-AzaC) is a demethylating agent that is also known to induce mutagenesis in mammalian cells. In this study, the mutagenic potential of this drug was tested in the G10 and G12 transgenic Chinese hamster cell lines, which have a single bacterial gpt gene integrated into the genome at different sites, with its expression driven by a simian virus 40 (SV40) promoter. We show that the mutation frequencies following a 48-h exposure to different concentrations of 5-AzaC were 10 to 20 times higher than those of any of the other numerous mutagens that have been tested in the G10-G12 system. Moreover, the mutation frequencies were much higher in the G10 cell line than in the G12 cells. Detailed molecular analysis of the 6-thioguanine (6-TG)-resistant variants demonstrated that transgene silencing by de novo DNA methylation and increased chromatin condensation in the SV40 promoter was the major factor responsible for this high level of 6-TG resistance. As would be expected, exposure to 5-AzaC lowered the overall genomic DNA methylation levels, but it unexpectedly caused hypermethylation and increased chromatin condensation of the transgene in both the G10 and G12 cell lines. These results provide the first evidence that 5-AzaC may also induce transgene-specific DNA methylation, a phenomenon that can further be used for the elucidation of the mechanism that controls silencing of foreign DNA.

Animals↗
PubMed1999

Diabetes management in a health maintenance organization. Efficacy of care management using cluster visits.

OBJECTIVE: To evaluate the effectiveness of a cluster visit model led by a diabetes nurse educator for delivering outpatient care management to adult patients with poorly controlled diabetes. RESEARCH DESIGN AND METHODS: This study involved a randomized controlled trial among patients of Kaiser Permanente's Pleasanton, CA, center who were aged 16-75 years and had either poor glycemic control (HbA1c > 8.5%) or no HbA1c test performed during the previous year. Intervention subjects received multidisciplinary outpatient diabetes care management delivered by a diabetes nurse educator, a psychologist, a nutritionist, and a pharmacist in cluster visit settings of 10-18 patients/month for 6 months. Outcomes included change (from baseline) in HbA1c levels; self-reported changes in self-care practices, self-efficacy, and satisfaction; and utilization of inpatient and outpatient health care. RESULTS: After the intervention, HbA1c levels declined by 1.3% in the intervention subjects versus 0.2% in the control subjects (P < 0.0001). Several self-care practices and several measures of self-efficacy improved significantly in the intervention group. Satisfaction with the program was high. Both hospital (P = 0.04) and outpatient (P < 0.01) utilization were significantly lower for intervention subjects after the program. CONCLUSIONS: A 6-month cluster visit group model of care for adults with diabetes improved glycemic control, self-efficacy, and patient satisfaction and resulted in a reduction in health care utilization after the program.

Adolescent↗