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M Carvalho

Publications and source records attributed to M Carvalho.

At least 73 records · Page 4Linked to original sources
PubMed1993

Protein interactions with DNA elements in variant equine infectious anemia virus enhancers and their impact on transcriptional activity.

The long terminal repeats (LTRs) from various cloned equine infectious anemia virus (EIAV) proviruses differ significantly, but all contain cis-acting DNA elements identical to MDBP-, PEA2-, AP-1-, and PU.1 (ets)-binding sites. A prototype EIAV LTR would contain one of each of these conserved elements. The LTR variations originate from the insertion of novel sequences between the PEA2 and AP-1 elements in the transcriptional enhancer unit. Viewed in this way, the LTR from provirus clone lambda 12 has an 11-bp insertion containing a PEA2 site and the LTR of the lambda 6 provirus has a 31-bp insertion/duplication containing PEA2, AP-1, and PU.1 sites. Two other LTRs were cloned by amplification of cDNAs from the persistently infected cell line, EIAV-FEA. A third LTR was generated by site-directed mutagenesis of one of the LTRs from EIAV-FEA cells. The latter three had a single base change in the element next to the TATA box that abolished PU.1 binding; however, the variable regions of these LTRs were shown by gel mobility shift assays to contain one or two PU.1 sites. One variable region was shown to have an octamer site overlapping its tandem PU.1 elements. Basal, PMA-activated, and Tat trans-activated transcriptional activities of the LTRs were compared in several different cell lines by transient transfection. The various promoters displayed different relative levels of activity depending on the cell line used and the condition of activation. This natural set of variant promoters may help define how changes in the components of the transcription complex influence transactivation by Tat. The diverse LTRs could endow their respective proviruses with a unique pattern of expression and activation in vivo.

Base Sequence↗
PubMed1993

Physical and functional characterization of transcriptional control elements in the equine infectious anemia virus promoter.

Equine infectious anemia virus (EIAV) is a lentivirus that causes a chronic disease of horses characterized by cyclic episodes of fever, anemia, and viremia. Although the genome and promoter of EIAV are much less complex than those of its relatives the primate immunodeficiency viruses, the cellular proteins that activate and regulate transcription of EIAV have not yet been identified. In this report, we show by electrophoretic mobility shift assays and DNase I footprinting that the EIAV promoter contains multiple binding sites for ubiquitous, cell type-specific, and inducible cellular proteins. Functional analysis by transient transfection of canine osteosarcoma (D17) and human epithelial carcinoma (HeLa) cells with EIAV promoters containing deletions or individually mutated DNA-binding sites demonstrated that these DNA-binding elements cooperatively regulate transcriptional activity. A methylated DNA-binding site (MDBP; also designated EF-C or EP) acts as either a positive or negative regulator of promoter activity, depending on the cell type or condition. Two PEA2 elements, an AP-1 site, and an ets/PEA3 motif confer a positive effect on promoter activity. The EIAV promoter is shown to be activated by treatment of HeLa cells with phorbol myristate acetate (PMA). DNA-binding activities were induced in PMA-treated HeLa cells and formed complexes on oligonucleotides that contain the EIAV AP-1 and ets/PEA3 elements. Functional analysis of mutated promoters indicated that the ets/PEA3 motif was the principal mediator of PMA activation.

Animals↗
PubMed1993

The PU.1/Spi-1 proto-oncogene is a transcriptional regulator of a lentivirus promoter.

The enhancer unit present in the retrovirus equine infectious anemia virus (EIAV) was previously shown to contain binding sites for proteins belonging to MDBP, PEA2, AP-1, and ets families. The EIAV ets motif matches the consensus sequence for both PEA3- and PU.1-binding sites. Here, we show by gel shift analysis that PU.1, present in nuclear extracts from monocyte and B-lymphocyte cell lines, binds to oligonucleotides containing the EIAV ets element. HeLa cells transiently transfected with a PU.1 expression plasmid expressed nuclear factors that formed complexes indistinguishable from those seen with monocyte extracts. Antibodies to PU.1 protein either supershifted or abolished formation of these complexes, depending on the PU.1 epitopes recognized. The binding of PU.1 to the EIAV ets motif in vitro correlated with transcriptional activity of the EIAV promoter in transfected monocyte cell lines. In HeLa cells, the product of PU.1 cDNA bound to the EIAV ets motif and activated transcription from the EIAV promoter. The PU.1-binding site was the primary determinant of EIAV promoter activity in cell lines that express PU.1. Nucleotide determinants of PU.1 binding and a consensus PU.1 binding sequence were defined in gel shift assays using a panel of mutated oligonucleotides. To our knowledge, this is the first report of a retroviral promoter controlled by PU.1.

Base Sequence↗
PubMed1993

[Pure autonomic failure].

We present the case of a 64-year-old woman who, in the past 5 years, complained of constipation/diarrhea, hyposudoresis, xerostomia and xerophthalmia, dysuria and orthostatic hypotension. Cardiovascular reflexes analysis revealed sympathetic and parasympathetic failure. Norepinephrine was markedly reduced, both lying and after tilt. Norepinephrine infusion determined a significant rise in blood pressure, allowing the diagnosis of denervation hypersensitivity. The diagnosis of pure autonomic failure was made. Therapy with 9 alpha fludrocortisone and metoclopramide was initiated with marked and sustained symptomatic effect.

Autonomic Nervous System Diseases↗
PubMed1993

[Neurophysiological methods for the diagnosis of genitourinary dysfunctions].

Genito-urinary function is a watershed field interesting several areas of medical concern. Neurophysiological studies allow objective assessment of central, peripheral and autonomic pathways in the nervous system contributing to topographical localisation of lesions. The authors describe different methods used, namely somatosensory evoked potentials by pudendal nerve stimulation, bulbo-cavernous reflex, cutaneous sympathetic reflex, transcranial cortical or spinal magnetic stimulation and single fiber EMG. Abnormalities in neurological diseases are presented.

Anal Canal↗
PubMed1993

[Familial amyloidotic polyneuropathy --Portuguese type-- clinical and neurophysiologic course].

Sixteen (16) patients with Familial Amyloidotic Polyneuropathy-Portuguese Type (FAP) were evaluated longitudinally, using the EMG and Clinical protocols with the aim of establishing the clinical and neurophysiological evolution rate of FAP. We conclude that spontaneous evolution of FAP patients shows a linear progression. Clinical and neurophysiological evolution times are parallel, with the EMG scores being higher than Clinical ones. We propose that Clinical and EMG scores should be used in the evaluation of future therapeutic trials, namely immunodepuration and liver transplantation.

Adult↗
PubMed1993

[Significance exercise-induced ST elevation in patients with myocardial infarction].

In 52 patients with previous myocardial infraction, 49 men and 3 women (mean age 56 +/- 7.1 years) the significance of ST-segment elevation during the stress-test, was evaluated. Of the 52 patients 15 (29%) showed St-segment elevation and 37(71%), showed no alteration of the ST-segment. Extension of coronary disease, degree of obstruction, wall motion abnormalities and the presence of residual ischemia were evaluated by coronary angiography, technetium-99M pyrophosphate imaging and exercise TL-201 scintigraphy. From the results of the study one may conclude that, in patients with previous myocardial infraction exercise, ST-segment elevation is a consequence of sub-occlusion of the left anterior descending coronary artery with severe ventricular dysfunction either in patients with one or multiple vessel disease.

Coronary Angiography↗
PubMed1992

Detecting single base substitutions as heteroduplex polymorphisms.

We have developed a sensitive technique for detecting single base substitutions in polymerase chain reaction (PCR) products from individuals heterozygous for polymorphisms or new mutations. This technique takes advantage of the formation of heteroduplexes in the PCR between different alleles from heterozygous individuals. These heteroduplexes can be detected on polyacrylamide gels because they migrate slower than their corresponding homoduplexes. Using PCR, we have generated a series of point mutations in a defined region of DNA in the equine infectious anemia virus (EIAV). Each mutation is the result of a single base substitution. By mixing the PCR products amplified from these mutations with one another, as well as with wildtype PCR products, we can generate heteroduplexes in which the identity of the mismatched bases is known. We detected eight of nine point mutations using this technique. We have also modified the electrophoretic conditions to optimize the detection of these heteroduplexes. In addition, the usefulness of this technique is demonstrated by its ability to detect a mutation in the cystic fibrosis gene that is the result of a single base substitution. This technique should prove useful for rapidly screening large numbers of individuals for new mutations or polymorphisms.

Base Sequence↗
PubMed1992

[Angina pectoris--particular aspects in the elderly].

The incidence of the coronary heart disease increases dramatically with age, representing in Portugal the second largest cause of death in patients older than 65 years. The cause of this high prevalence of the disease is not completely understood, but the pathogenic role of the cardiovascular risk factors has a singificative importance. It seems to be essential, even in elderly, to control these risk factors in order to limit the progression of the atherosclerotic disease. Regarding symptomatology, effort dyspneia appears to be one of the most frequent disease manifestations, differing from the typical chest pain presentation observed in the younger patients. Exercise stress testing can be used safely in the elderly and it is not just an important diagnostic tool, but it is also essential to the stratification of the patients to be more agressively treated. Concerning treatment, the search and correction of the precipitating factors are a key to success in the elderly patients. Pharmacologic therapy must be prescribed in agreement with the clinical pharmacologic principles, carefully adjusted to the elderly, in order to avoid adverse drug effects. In spite, coronary bypass surgery has some higher risk in the elderly. The results obtained are satisfactory, and the patients subset with ventricular disfunction, benefit the most. Percutaneous transluminal coronary angioplasty constitutes a very atractive therapeutic modality in the elderly. Cardiac reabilitation programe are very useful in the elderly, contributing to the desired psycosocial reintegration of these patients.

Aged↗
PubMed1991

Plasma catecholamines and postural hypotension in familial amyloidotic polyneuropathy of the Portuguese type.

Plasma adrenaline and noradrenaline levels were measured while supine and following head-up tilt to 45 degrees, in both normal controls and in patients with familial amyloidotic polyneuropathy of the Portuguese type. In nine patients systolic blood pressure fell by less than 15 mmHg, while in seven patients it fell by more than 15 mmHg. Plasma noradrenaline rose during tilt in the majority of patients, as in the controls. There was no correlation between levels of catecholamines and fall in blood pressure on head-up tilt. The data excludes widespread sympatho-neural failure as a cause for postural hypotension in familial amyloidotic polyneuropathy of the Portuguese type. The results are compatible with either segmental/patchy sympathetic denervation or dysfunction of the receptor/effector mechanisms in target organs such as the heart and blood vessels.

Adult↗
PubMed1991

A minimal lentivirus Tat.

Transcriptional regulatory mechanisms found in lentiviruses employ RNA enhancer elements called trans-activation responsive (TAR) elements. These nascent RNA stem-loops are cis-acting targets of virally encoded Tat effectors. Interactions between Tat and TAR increase the processivity of transcription complexes and lead to efficient copying of viral genomes. To study essential elements of this trans activation, peptide motifs from Tats of two distantly related lentiviruses, equine infectious anemia virus (EIAV) and human immunodeficiency virus type 1 (HIV-1), were fused to the coat protein of bacteriophage R17 and tested on the long terminal repeat of EIAV, where TAR was replaced by the R17 operator, the target of the coat protein. This independent RNA-tethering mechanism mapped activation domains of Tats from HIV-1 and EIAV to 47 and 15 amino acids and RNA-binding domains to 10 and 26 amino acids, respectively. Thus, a minimal lentivirus Tat consists of 25 amino acids, of which 15 modify viral transcription and 10 bind to the target RNA stem-loop.

Amino Acid Sequence↗
PubMed1991

Mutational analysis of the equine infectious anemia virus Tat-responsive element.

A hairpinlike structure is predicted to exist at the 5' end of equine infectious anemia virus (EIAV) RNA which is similar in many ways to the human immunodeficiency type 1 (HIV-1) Tat-responsive element (TAR). In EIAV, this structure has a shorter stem than in HIV-1 and lacks the uridine bulge. Primer extension analysis of EIAV RNA was used to identify the transcriptional start site in the viral long terminal repeat. Premature termination of primer elongation at the predicted double-stranded RNA region was frequently observed and suggests that the inferred hairpin structure exists under these conditions. We have functionally characterized EIAV TAR by site-directed mutagenesis and transient gene expression analysis. It is demonstrated here that the secondary structure of this element is essential for Tat action. Mutations that disrupted base pairing abolished TAR function, and compensatory mutations that restored the stem structure resulted in Tat activation. The TAR loop appears to be closed by two U.G base pairs that are likely to provide a unique structural motif recognized by the Tat protein. With one exception, substitutions of nucleotides within the EIAV loop sequence decreased TAR function. All nucleotide substitutions of the cytidine at position +14 increased EIAV Tat responsiveness; however, its deletion abolished trans activation. Our results lead us to propose that the EIAV and HIV-1 Tat systems employ closely related cis- and trans-acting components that probably act by the same mechanism.

Base Sequence↗
PubMed1991

[Myocardiopathy caused by Portuguese-type familial amyloidotic polyneuropathy. Sequential morphologic and functional study of 60 patients].

AIM OF THE STUDY: To assess the cardiac involvement of familial amyloidotic polyneuropathy--Portuguese type (FAP) in a prospective study. INTRODUCTION: FAP is a sensitive, motor and autonomic familial polyneuropathy, due to amyloid deposits on nerve and vascular structures, related to abnormal transthyretin synthesis. FAP is a progressive systemic disease. The associated cardiomyopathy is well characterized in the Japanese form, but studies on FAP--Portuguese type cardiomyopathy remains controversial, regarding its occurrence and clinical and morphofunctional aspects. POPULATION: The Authors studied 60 Portuguese caucasoid patients, belonging to 55 different families: 23 women from 27 to 67 years old (m = 40.78 +/- 18.4) and 37 men, from 31 to 60 years old (m = 40.8 +/- 12.8). The diagnosis was established by characteristic clinical aspects, electromyograms and nerve or gut biopsies, positive for amyloid. Patients were classified in phases from I to VI according to the severity of neurological involvement and disability. METHODS: All the patients carried out clinical evaluation, thorax X-Ray, conventional EKG, Holter EKG (20-24 hours), M + 2D echocardiography and phonocardiography. Morphological and functional data were obtained from 85 echocardiographic recordings; special attention was paid to the evidence of structural abnormalities, evaluation of internal chamber dimension, left ventricular mass and systolic function indexes (left ventricular fractional shortening, left ventricular ejection fraction, left ventricular end-systolic stress index and Vcf). CONCLUSIONS: Suggestive aspects of pericardial and myocardial amyloid infiltration were found even in the earliest phases of the disease; all the records of patients in advanced phases (III to VI) were abnormal. The most frequent abnormalities founded were: a bright sparkling pericardial and myocardial echo pattern and increased ventricular wall thickness, with increased left ventricular mass index in 85% of patients in phases III to VI. Vcf was abnormally increased when plotted together with left ventricular end-systolic stress index, which was normal or decreased, probably reflecting "excessive" inotropism, in 1/3 of patients in phases III to VI. Systolic function was never depressed; no cardiac chamber enlarged. So, this study points out the presence of an infiltrative and/or hypertrophic cardiomyopathy in FAP--Portuguese type; suggestive aspects of diastolic dysfunction and decrease in systolic function were recorded. However, in the clinical setting of FAP, symptoms and signs like dyspnea, fatigue, oedema, may not be diagnosis of congestive heart disease.

Adult↗
PubMed1991

[Thrombolysis in acute myocardial infarction in the aged. A greater risk of cardiac rupture?].

OBJECTIVE: To evaluate age group and i.v. thrombolytic therapy (TT) influences on cardiac rupture complicating acute myocardial infarction (AMI). CONCEPTION: Retrospective analysis of patients (pts) admitted during 1988-89 to an Intensive Care Unit of a Terciary Hospital (UCIM) with AMI. POPULATION: 430 pts were admitted with AMI. During this period 89 pts were submitted to TT (25% age greater than or equal to 65 years). Eighty four pts died during hospital stay and 42 had autopsy study (50%). Only 7 of nonautopsied pts died of undetermined cause. Dead pts had age m +/- SD 72 +/- 11 years old (yo) (37% female, 63% male and 74% age greater than or equal to 65 yo). RESULTS: Nineteen pts died of cardiac rupture (CR) (23%). CR prevalence was 1.4% (3/218) in pts less than 65 yo and 7.5% (16/212) in pts greater than or equal to 65 yo (p less than 0.01). CR prevalence in pts submitted to TT (4.5%) 4/89, was similar to pts not submitted to TT (4.4%) 15/341 (n.s.). Pts less than 65 yo had nonsignificant differences in CR wether submitted or not to TT (0% vs 2%). Elderly pts (greater than or equal to 65 yo) CR prevalence was 18% (4/22) in those submitted to TT vs 6% (12/190) in those not submitted to TT (p less than 0.05). CONCLUSIONS: Thrombolytic therapy may carry an additional risk for cardiac rupture in elderly patients (greater than or equal to 65 yo).

Age Factors↗
PubMed1990

Spinal cord compression secondary to idiopathic retroperitoneal fibrosis.

Retroperitoneal fibrosis is a rare disorder in which the abnormal fibrotic tissue compresses retroperitoneal organs. In the majority of patients no obvious cause can be found, hence the name idiopathic retroperitoneal fibrosis. The process can also arise from other anatomic areas, suggesting a multifocal origin. We report a case of dorsal epidural compression in a 63-year-old patient harboring idiopathic retroperitoneal fibrosis. A magnetic resonance imaging scan revealed the precise location of the lesion. It is suggested that retroperitoneal fibrosis should be considered in the differential diagnosis of epidural spinal cord compression.

Female↗
PubMed1990

[X syndrome: review of concepts].

Patients with syndrome X or microvascular angina present complains of effort angina associated with normal--appearing coronary arteries. Ischemia seems to be caused by myocardial perfusion abnormalities, related to microcirculatory disfunction, characterized by excessive vasoconstriction and/or inadequate vasodilation response. The intimal cause of this microcirculatory disfunction, located in the small pre-arteriolares arteries with 100-200 microns caliber, is still unknown. It seems that these patients present a generalized abnormality of the systemic smooth muscle, affecting territories other than the vascular. The diagnosis should be suspected whenever a patient presents effort angina, angiographically normal arteries and evidence of ischemia to exercise. As this syndrome is better understood some subsets of patients with different clinical characteristics and prognosis, are being identified. The calcium channel blockers seem to be the drug more effective in the control of ischemia and symptoms. The prognosis is, in general, better than in patients with coronary angiographic lesions.

Angina Pectoris↗