Biomedical subjects

M F Alves

Publications and source records attributed to M F Alves.

5 recordsLinked to original sources

PubMed2005

A continuous fluorescent assay for the determination of plasma and tissue angiotensin I-converting enzyme activity.

A continuous assay using internally quenched fluorescent peptides with the general sequence Abz-peptidyl-(Dnp)P-OH (Abz = ortho-aminobenzoic acid; Dnp = 2,4-dinitrophenyl) was optimized for the measurement of angiotensin I-converting enzyme (ACE) in human plasma and rat tissues. Abz-FRK(Dnp)P-OH, which was cleaved at the Arg-Lys bond by ACE, was used for the enzyme evaluation in human plasma. Enzymatic activity was monitored by continuous recording of the fluorescence (lambda ex = 320 nm and lambda em = 420 nm) at 37 degrees C, in 0.1 M Tris-HCl buffer, pH 7.0, with 50 mM NaCl and 10 microM ZnCl2. The assays can be performed directly in the cuvette of the fluorimeter and the hydrolysis followed for 5 to 10 min. ACE measurements in the plasma of 80 healthy patients with Hip-His-Leu and with Abz-FRK(Dnp)P-OH correlated closely (r = 0.90, P < 0.001). The specificity of the assay was demonstrated by the complete inhibition of hydrolysis by 0.5 microM lisinopril or captopril. Abz-FRK(Dnp)P-OH cleavage by ACE was monitored in rat lung, kidney, heart, and liver homogenates in the presence of a cocktail of inhibitors containing trans-epoxy-succinyl-L-leucylamido-(4-guanido)-butene, pepstatin, phenyl-methylsulfonyl fluoride, N-tosyl-L-phenylalanyl-chloromethyl ketone, and N-tosyl-lysyl-chloromethyl ketone to prevent undesirable hydrolysis. ACE activity in lung, heart and kidney homogenates, but not in liver homogenates, was completely abolished by 0.5 microM lisinopril or captopril. The advantages of the method are the procedural simplicity and the high sensitivity providing a rapid assay for ACE determinations.

Animals↗

PubMed1999

Internally quenched fluorogenic substrates for angiotensin I-converting enzyme.

OBJECTIVE: Development of internally quenched fluorogenic substrates for sensitive and continuous assays of angiotensin I-converting enzyme (ACE). DESIGN: We synthesized internally quenched fluorogenic bradykinin-related peptides introducing Abz (ortho-aminobenzoic acid) and EDDnp (N-[2,4-dinitrophenyl]-ethylenediamine) at their N- and C-terminal groups, respectively, and these were assayed as ACE substrates. We examined two series of peptides, Abz-GFSPFRX-EDDnp and Abz-GFSPFXQ-EDDnp (X, various amino acids). METHODS: Hydrolysis of the fluorogenic substrates by ACE was followed by continuous recording of the rising fluorescence (lambda(em) = 420 nm and lambda(ex) = 320 nm). The peptides were obtained by solid-phase synthesis or by classical solution methods. RESULTS: Despite of the blocked C-terminal sequences, the internally quenched bradykinin-related peptides were hydrolysed by ACE. The best substrates for plasma guinea pig ACE were Abz-GFSPFRA-EDDnp and Abz-GFSPFFQ-EDDnp, in which the fluorescence appeared after the first cleavage that occurred at R-A and F-Q bond, respectively. This ACE activity was sensitive to NaCl concentration and the optimum pH is greater than 8.0. Measurements of ACE activity with Hip-His-Leu and Abz-GFSPFFQ-EDDnp in the serum of 20 healthy patients correlated closely (r = 0.959). Complete inhibition of the hydrolysis of Abz-GFSPFFQ-EDDnp by human serum was observed with captopril and lisinopril. CONCLUSIONS: We describe internally quenched fluorogenic substrates for ACE devoid of free C-terminal carboxyl group. They are convenient tools for ACE studies as they permit continuous fluorimetric measurements of the enzymatic activity, even in human serum.

Angiotensin-Converting Enzyme Inhibitors↗

PubMed1998

Type I and type III collagens in cutaneous mucinosis.

Cutaneous mucinoses are a heterogeneous group of diseases characterized by the focal or diffuse dermal deposition of glycosaminoglycans. The histopathologic examination of many cutaneous mucinoses reveals that the collagen fibers are fragmented. We wanted to characterize the type I (COL1) and type III (COL3) collagen distribution in skin biopsy specimens of patients with cutaneous mucinosis. The diagnosis of mucinosis was based on a modification of the classification by Rongioletti and Rebora: four patients had familial papulonodular mucinosis: four had papular mucinosis, one of which was associated with myxedema and one had scleromyxedema; and one had focal mucinosis. We performed anti-type I and type III collagens immunolabeling on frozen sections. Immunofluorescence for COL1 was increased in the superficial dermis of 2/4 familial papulonodular mucinosis, in 5/5 of papular mucinosis, and in scleromyxedema and focal mucinosis cases. The mid-dermis showed intense staining for COL1 at the periphery of collagen bundles and, in three cases of familial papulonodular mucinosis and two cases of papular mucinosis, a lacy appearance. The superficial dermis of familial papulonodular mucinosis specimens and of papular mucinosis + myxedema, scleromyxedema, and focal mucinosis specimens had decreased COL3 staining. The mid-dermis showed a more prominent fibrillar staining at the periphery of the collagen bundles, and two cases of papular mucinosis showed intense labeling for COL3. Both COL1 and COL3 distributions are altered in cutaneous mucinosis. An intense labeling with COL1 is predominantly found in the superficial layer of cutaneous mucinosis. Cases of FTP revealed decreased COL3 reactivity at the superficial layer.

Adolescent↗

PubMed1993

[Trial of labor after cesarean section. Two years' experience].

A retrospective study was undertaken to evaluate the incidence of vaginal delivery in women that previously undergone a cesarean section; in addition, incidence of complications such as uterine rupture and scar dehiscence were evaluated. During the two years reviewed, 251 out of 324 women with a previous cesarean section were allowed to labor; 64.1% had a vaginal delivery. Uterine rupture occurred in 0.8% of the patients of the trial of labor group: no other significant maternal and/or fetal complications were seen; scar dehiscence occurred in 3.3%. Statistical analysis didn't show any increase in complications when oxytocin was used to stimulate uterine activity. It is concluded that trail of labor and delivery after cesarean section is a safe option in a large number of patients.

Adult↗

PubMed1992

Cytokines and dysregulation of the immune response in human malaria.

The dysregulation of the immune response by malaria parasite has been considered as a possible constraint to the effectiveness of malaria vaccination. In spite of the important role interleukin-1 (IL-1) plays on the immunoregulation, and its ability to mimic various features of clinical malaria, reports on IL-1 in malaria are lacking. We found that only 2 out of 35 subjects with acute malaria showed increased levels of serum IL-1 alpha by enzyme immunoassay. To assess whether IL-1 could interfere with T-lymphocyte responses, blood mononuclear cells from patients infected with Plasmodium falciparum, P. vivax, or healthy subjects were cultured with phytohemagglutinin, and lymphocyte proliferation measured 72 h later by 3H-thymidine incorporation. Our data showed that T-lymphocyte responses are depressed both in P. falciparum (10,500 +/- 2,900) and P. vivax malaria (13,000 +/- 3,300), as compared to that of healthy individuals (27,000 +/- 3,000). Addition of IL-1 partially reversed depression of malaria lymphocytes, but had no effect on normal cells. On the other hand, T-lymphocytes from malaria infected-subjects presented a minimal decrease in proliferation, when cultured in the presence of exogenous PGE2. These data indicate the occurrence of two defects of immunoregulation in malaria: a deficiency of IL-1 production by monocytes/macrophages, and an increased resistance of lymphocytes to the antiproliferative effect of PGE2.

Dinoprostone↗