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Biomedical subjects

G R Castro

Publications and source records attributed to G R Castro.

At least 19 recordsLinked to original sources

Lindane removal induction by Streptomyces sp. M7.

Gamma-hexachlorocyclohexane (gamma-HCH or lindane), one of the most commonly used pesticides, has been mainly used in agriculture; this pesticide is known to be highly toxic and persistent, causing serious water and soil contamination. The objective of the present work is to study the effect of low glucose concentration and the addition of lindane at different growing time on the pesticide detoxification ability of Streptomyces M7. After 96 h of incubation in synthetic medium containing glucose 0.6 g l(-1) with the addition of lindane 100 microg l(-1) at 20 h of incubation, a typical diauxic curve was obtained: glucose was the preferred substrate until 24 h, at 48 h, when the carbohydrate was depleted, the microorganism consumed the pesticide like carbon source. On the other hand, lindane removal induction was observed, which was greater when the pesticide was added to the medium at 20 h than 6 h of incubation. Between 72 and 96 h, a maximum of approximately 86% of the Cl(-) was released when lindane was added to the medium at 20 h, whereas approximately 70% and 67% Cl(-) was released in the medium when the pesticide was added at 0 and 6 h of incubation respectively. This is the first report of chloride release from inoculated medium supplemented with lindane, suggesting that the pesticide was degraded by Streptomyces sp. under aerobic conditions.

Biodegradation, Environmental↗

Tangential-flow ultrafiltration: a versatile methodology for determination of complexation parameters in refractory organic matter from Brazilian water and soil samples.

In this work the copper(II) complexation parameters of aquatic organic matter, aquatic and soil humic substances from Brazilian were determined using a new versatile approach based on a single-stage tangential-flow ultrafiltration (TF-UF) technique (cut-off 1 kDa) and sensitive atomic spectrometry methods. The results regarding the copper(II) complexation capacity and conditional stability constants obtained for humic materials were compared with those obtained using direct potentiometry with a copper-ion-selective electrode. The analytical procedure based on ultrafiltration is a good alternative to determine the complexation parameters in natural organic material from aquatic and soil systems. This approach presents additional advantages such as better sensibility, applicability for multi-element capability, and its possible to be used under natural conditions when compared with the traditional ion-selective electrode.

Brazil↗

Determination of Cu, Ni, and Zn in fuel ethanol by FAAS after enrichment in column packed with 2-aminothiazole-modified silica gel.

This work describes the synthesis and characterization of 2-aminothiazole-modified silica gel (SiAT), as well as its application for preconcentration (in batch and column technique) of Cu(II), Ni(II) and Zn(II) in ethanol medium. The adsorption capacities of SiAT determined for each metal ion were (mmol g(-1)): Cu(II)=1.20, Ni(II)=1.10 and Zn(II)=0.90. In addition, results obtained in flow experiments, showed a recovery of ca. 100% of the metal ions adsorbed in a column packed with 500 mg of SiAT. The eluent was 2.0 mol L(-1) HCl. The sorption-desorption of the studied metal ions made possible the development of a preconcentration method for metal ions at trace level in fuel ethanol using flame AAS for their quantification.

Automobiles↗

Isolation of four aquatic streptomycetes strains capable of growth on organochlorine pesticides.

Ninety-three wild-type isolates identified as actinomycetes were tested against 11 organochlorine pesticides (OPs): aldrin, chlordane, DDD, DDE, DDT, dieldrin, heptachlor, and heptachlor epoxides, lindane, and methoxychlor. Qualitative screening agar assays displayed 62-78% tolerance of strains to OPs. Four strains designed M4, M7, M9 and M15 were selected based on multi-OP-tolerance, and identified as members of the streptomycetes group. Different growth profiles were observed in cultures of the four selected streptomycetes cultured in synthetic medium containing 5-50 microg x l(-1) aldrin or chlordane or lindane. Increase of aldrin removal by the selected microorganisms was concomitant with the 4.8-36.0 microg x l(-1) pesticide concentration range. After 72 h of streptomycete M7 growth in synthetic medium containing 48.0 microg x l(-1) aldrin, the remaining OP concentration in the supernatant was approximately 10% of the initial concentration. Also, in stationary growth phase less than 2.5 microg x l(-1) aldrin residual concentration was detected in the medium.

Biodegradation, Environmental↗

Homogeneous biocatalysis in organic solvents and water-organic mixtures.

Biocatalysis in non-aqueous media has undergone tremendous development during the last decade, and numerous reactions have been introduced and optimized for synthetic applications. In contrast to aqueous enzymology, biotransformations in organic solvents offer unique industrially attractive advantages, such as: drastic changes in the enantioselectivity of the reaction, the reversal of the thermodynamic equilibrium of hydrolysis reactions, suppression of water-dependent side reactions, and resistance to bacterial contamination. Currently, the field is dominated by heterogeneous biocatalysis based primarily on lyophilized enzyme powders, cross-linked crystals, and enzymes immobilized on inert supports that are mainly applied in enantioselective synthesis. However, low reaction rates are an inherent problem of the heterogeneous biocatalysis, while the homogeneous systems have the advantage that the elimination of diffusional barriers of substrates and products between organic and water phases results in an increase in the reaction rate. Here the discussion is focused on the correlation between activity and structure of the intact enzymes dissolved in neat organic solvents, as well as modifications of natural enzymes, which make them soluble and catalytically active in non-aqueous environment. Factors that influence conformation and stability of the enzymes are also discussed. Current developments in non-aqueous biocatalysts that combine advantages of protein modification and immobilization, i.e., HIP plastics, enzyme chips, ionic liquids, are introduced. Finally, engineering enzymes for biotransformations in non-conventional media by directed evolution is summarized.

Catalysis↗

Phosphatidylinositol-specific phospholipase C activity in Lactobacillus rhamnosus with capacity to translocate.

Phosphatidylinositol-specific phospholipase C (PI-PLC) activity was investigated in 25 different lactic acid bacteria (LAB) strains belonging to the genera Lactobacillus, Weisella, and Enterococcus. PI-PLC activity was detected in 44% of the strains studied in culture medium without carbon source. From the PI-PLC positive strains, Lactobacillus rhamnosus ATCC 7469 was selected for translocation studies. Healthy mice were orally administered with a daily dose of 2.0 x 10(9) of viable L. rhamnosus suspension. Viable bacteria were detected in liver and spleen of mice fed with LAB for 7 days. Bacterial colonies isolated from liver were biochemically characterized, and further subjected to randomly amplified polymorphic DNA. Amplification patterns of five strains displayed identical profiles to L. rhamnosus. PI-PLC activity was determined in the strains recovered from liver.

Animals↗

Chromium accumulation by two Streptomyces spp. isolated from riverine sediments.

Strains designated R22 and R25, isolated from Salí River sediments, Argentina, were highly resistant to chromium. These strains were shown by 16S rRNA sequencing studies to be Streptomyces spp.; this affiliation was consistent with morphological and chemical characteristics. Growth of strains R22 and R25 in medium containing 100 mg l(-1) chromate was reduced by only 23% and 34%, respectively, compared with growth in medium without added chromium. Streptomyces sp. strains R22 and R25 both accumulated chromium with yields of 10.0 and 5.6 mg Cr g(-1) of dry weight, respectively, and a chromate concentration of 50 mg ml(-1). Cell fractionation studies with strain R22 showed that the great majority of the chromium were associated with the cell wall fraction. Streptomyces strains R22 and R25 may have applications in bioremediation of chromium contamination.

Biodegradation, Environmental↗

Alterations in the main steps of reverse cholesterol transport in male patients with primary hypertriglyceridemia and low HDL-cholesterol levels.

Hypertriglyceridemia is a complex pathological entity strongly connected to low HDL-C levels but controversially related to the risk of coronary artery disease. In this study, we evaluated the main steps of the antiatherogenic pathway called reverse cholesterol transport in a group of patients with primary hypertriglyceridemia and low HDL-C levels in comparison to normotriglyceridemic subjects with or without hypoalphalipoproteinemia. In patients with primary hypertriglyceridemia, low HDL-C levels were accompanied by decreased apo A-I and apo A-II concentrations. These reductions were manifested by a selective reduction in LpA-I:A-II particles. In addition, apo C-III Lp non B was found to be elevated and HDL lipid percentage composition showed a triglyceride enrichment and cholesterol depletion. The capacity of serum samples from hypertriglyceridemic patients to promote cellular cholesterol efflux was reduced, as evidenced by using two different cellular models, Fu5AH and J774 cells. This impaired cholesterol efflux promotion was also corroborated by incubations of isolated HDL fractions with Fu5AH cells. Lecithin:cholesterol acyltransferase (LCAT) activity, the driving force of reverse cholesterol transport, showed a tendency towards lower values in hypertriglyceridemic patients, but this difference was not statistically significant. Additionally, cholesteryl ester transfer protein (CETP) activity was increased in this group of patients. Therefore, hypertriglyceridemia was found to induce quantitative and qualitative alterations in HDL and its subclasses and, consequently, in some steps of reverse cholesterol transport. The abnormalities found in this antiatherogenic pathway and its promoters could constitute a possible connection between hypertriglyceridemia and atherosclerosis.

Adult↗

Abnormal capacity to induce cholesterol efflux and a new LpA-I pre-beta particle in type 2 diabetic patients.

In this study, we first characterized the lipoprotein components of serum samples obtained from a group of well-controlled diabetic patients and from healthy subjects in fasting and postprandial states. We then explored some aspects of reverse cholesterol transport in the same population. Patients showed high levels of fasting triglycerides, postprandial triglyceride responses and LpC-III levels (3.18+/-0.86 vs 2.17+/-0.54 mg/dl, P < 0.001). There were also positive correlations between LpC-III and fasting triglycerides (r = 0.82, P < 0.001), total triglyceride area (r = 0.75, P < 0.001) and incremental triglyceride area (r = 0.54, P < 0.001). HDL-C and apo A-I were significantly decreased in diabetic patients due to a selective reduction in LpA-I subfraction, whose antiatherogenic role is generally accepted (37.4+/-8.0 vs 49.2+/-12.5 mg/dl, P < 0.001). In addition, HDL from patients proved to be triglyceride enriched and cholesteryl ester depleted, alterations which were further amplified in the postprandial state. The molar ratio HDL-C/apo A-I + apo A-II, already defined as a predictor of apo A-I fractional catabolic rate, was significantly diminished in the patient group (15.1+/-2.2 vs 20.8+/-3.3, P < 0.001), thus suggesting an accelerated catabolism of apo A-I. For the first time, we describe here the presence of a small apo A-I-containing particle, isolated by two-dimensional electrophoresis and characterized by immunoblotting, only in samples from diabetic patients. This particle that we named pre-beta0, has an apparent molecular weight of 40 kDa. As regards the capacity of serum samples to promote cholesterol efflux from [3H]cholesterol-labeled Fu5AH rat hepatoma cells, patient samples were found to induce significantly lower cholesterol efflux than controls only in the postprandial state (21.2+/-3.3 vs 23.8+/-1.8%, P = 0.012). The presence of pre-beta0 in samples from diabetic patients might therefore be associated to an altered capacity of these serum samples to promote cellular cholesterol efflux. Overall, these abnormalities may contribute to a delay in the reverse cholesterol transport pathway in type 2 diabetic patients.

Adult↗

[Partial lecithin-cholesterol acyltransferase (LCAT) deficiency syndrome].

This syndrome is a pathological entity of low incidence which mainly affects high density lipoprotein (HDL) metabolism. We here show the first case reported in our country, observed in a 63-year-old woman who showed bilateral corneal opacity and eruptive xanthomas in both arms. The lipoprotein profile disclosed severe hypertriglyceridemia and normocholesterolemia, although the percentage of cholesteryl esters was low. Plasma levels of HDL-cholesterol and HDL major apolipoproteins, A-I and A-II, were markedly decreased. The patient also showed glucose intolerance and hematological alterations related to abnormal lipid composition of erythrocyte membranes. As evaluated by the exogen substrate method, LCAT activity proved to be 82% lower in the patient than in a control subject. It is noteworthy that the patient had experienced cardiac events and presented hypertension, neither of which has been commonly documented in partial LCAT deficiency syndromes.

Cholesterol, HDL↗

Optical design of the general-purpose Spanish X-ray beamline for absorption and diffraction.

This paper describes the optical design of the general-purpose Spanish beamline for absorption and diffraction (SpLine), which will be installed on a bending-magnet port of the ESRF. This beamline is planned to cover the needs of the Spanish synchroton radiation community with broad scientific fields of interest, covering physics, chemistry, materials science and biology. By using the advantages of a wide front end (9 mrad), the Spanish beamline will be split into two branches. Both branches will be equipped with focusing optics and experimental stations. Thus, each branch can be operated simultaneously and independently from each other. One branch (A) will have two experimental stations, one for high-resolution powder diffraction (HRPD) and the other for X-ray absorption spectroscopy (XAS) and X-ray standing waves (XSW). The other branch (B) will have facilities for macromolecular crystallography and for single-crystal diffraction analysis, including that of surfaces and interfaces, as well as an X-ray diffraction/scattering camera for non-crystalline specimens.

Journal Article↗

VLDL compositional changes and plasma levels of triglycerides and high density lipoprotein.

VLDL chemical composition is related to plasma levels of triglycerides and HDL-cholesterol. We evaluated patients with primary hypertriglyceridemia with or without hypoalphalipoproteinemia and subjects with normotriglyceridemia with hypoalphalipoproteinemia. The pattern observed in all the groups was an enrichment in the triglyceride content of VLDL and in apo B-VLDL. Compared to controls, LpC-III:B levels were higher in hypertriglyceridemic patients with low or normal HDL-cholesterol levels (7.3 +/- 0.6 vs. 14.9 +/- 1.8 and 12.3 +/- 2.8 mg/dl; P < 0.005 and P < 0.01, respectively) and LpE:B concentration was only increased in patients with hypertriglyceridemia and normal HDL-cholesterol levels (3.1 +/- 0.5 vs. 6.3 +/- 1.0 mg/dl; P < 0.01). The activity of the cholesteryl ester transfer protein was higher in hypertriglyceridemic patients with low HDL-cholesterol levels than in controls (380 +/- 25 vs. 262 +/- 14% cholesteryl esters/ml.h; P < 0.001). The most atypical VLDL particle was found in patients who combined an accumulation of VLDL particles and a reduction in HDL-cholesterol concentration. These two parameters represent both ends of the cholesteryl ester-triglyceride transfer, a crucial factor for VLDL chemical composition and HDL levels.

Adult↗

Rapid electrophoretic separation of pre-beta-migrating high density lipoproteins using automated PhastSystem: application to analysis of lecithin:cholesterol acyltransferase-deficient plasma.

To identify pre-beta-high density lipoproteins, a rapid two-dimensional separation by electrophoresis (1 hour 30 minutes) was performed on an automated Phast System. This procedure used commercially available polyacrylamide gradient gels (4-15%) and allows sensitive and reproducible results. Pre-beta-1- and pre-beta-2-high density lipoproteins were clearly identified by this method. In addition, our procedure was successfully applied to diagnosis of a patient with familial lecithin:cholesterol acyltransferase deficiency, characterized by the absence of alpha-high density lipoproteins.

Automation↗

Antimicrobial activity of flavonoids from leaves of Tagetes minuta.

The total extract and fractions with different solvents, obtained from leaves of Tagetes minuta, showed several degrees of antimicrobial activity against Gram positive and Gram negative microorganisms. The same fractions were inactive against Lactobacillus, Zymomonas and Saccharomices species. The major component of the extract: quercetagetin-7-arabinosyl-galactoside, showed significant antimicrobial activity on pathogen microorganisms tested. Correlation results were carried out using chloramphenicol as standard antibiotic.

Anti-Bacterial Agents↗

Chymase in exocytosed rat mast cell granules effectively proteolyzes apolipoprotein AI-containing lipoproteins, so reducing the cholesterol efflux-inducing ability of serum and aortic intimal fluid.

Degranulated mast cells are present in human fatty streaks. Chymase in granules released from degranulated rat serosal mast cells, i.e., in granule remnants, proteolyzes human high density lipoprotein3 (HDL3), and so reduces its ability to induce cholesterol efflux from macrophage foam cells in vitro. In this study we found that remnant chymase, by proteolyzing human serum and human aortic intimal fluid, prevents these two physiologic fluids from effectively inducing cholesterol efflux from cultured macrophage foam cells. Inhibition was strongest when remnants were added to apolipoprotein AI (apoAI)-containing lipoproteins; the remnants had no effect on the weaker efflux produced by apoAI-deficient serum. Western blot analysis showed that granule remnants degrade apoAI in serum and in internal fluid. When released from remnants, chymase lost its ability to proteolyze HDL3 in the presence of serum. Thus, remnant chymase (but not isolated chymase) was able to resist the natural protease inhibitors present in serum and in intimal fluid. The results imply participation of exocytosed mast cell granules in foam cell formation in atherogenesis.

Animals↗

Structural domain of apolipoprotein A-I involved in its interaction with cells.

Apolipoprotein A-I (apo A-I) is the major protein constituent of high-density lipoprotein (HDL), the lipoprotein fraction which mediates the reverse cholesterol transport. This apolipoprotein plays an important role in the binding of HDL to cells and participates in the efflux of cellular cholesterol. We have recently compared six different genetic variants of apo A-I and found that the apo A-I (Pro 165-->Arg) mutant is defective in promoting cellular cholesterol efflux from murine adipocytes and peritoneal macrophages and we have proposed that this region of apo A-I may be involved in their interaction with cells. To confirm this hypothesis, four monoclonal antibodies (mAbs) specific for apo A-I were used to study the inhibition of the interaction of palmitoyloleoylphosphatidylcholine (POPC): apoA-I complexes with HeLa cells and adipocytes. Among these antibodies, the apo A-I epitope recognized by the A44 mAb lies in the COOH terminal region (amino acid residues 149-186) including the proposed region. The antibodies A05, and A03 react with residues 25-82, 135-140, respectively and the A11 mAb corresponds to a discontinuous epitope at residues 99-105 and 126-132. Our results show clearly that the A44 and A05 mAbs reduce both the binding to HeLa cells and the cholesterol efflux from adipocytes. The inhibition of POPC: apoA-I complexes binding to both cell types is more strictly observed with the Fab fragments of monoclonal antibodies A44 and A05. Partial cotitration curves of these mAbs in a solid phase assay (RIA), indicated partial competition between these two antibodies. We propose a structural model for the POPC: apoA-I complexes where the N-terminal domain of one apo A-I molecule is in close spatial relationship with the C-terminal domain of the adjacent apo A-I molecule. We therefore suggest that the domain around amino acid 165 of apo A-I and which is recognized by mAb A44 (149-186) forms or contains some specific regions which mediate selectively the interaction with the binding site of cells and is involved in the efflux of cellular cholesterol.

Adipocytes↗

Apolipoprotein A-I-containing particles and reverse cholesterol transport: evidence for connection between cholesterol efflux and atherosclerosis risk.

It is now clearly established that apo A-I-containing lipoproteins exist as two major families, those containing apo A-I and apo A-II (LpA-I:A-II) and those containing apo A-I but free of apo A-II (LpA-I). Metabolic studies utilizing radiolabeled lipoprotein particles suggested that there is a kinetic difference between LpA-I and LpA-I:A-II family and support the concept that there may be important functional differences between the lipoprotein particles present within HDL. Of considerable significance was the finding that proteins stimulating reverse cholesterol transport (lecithin:cholesterol acyltransferase (LCAT), cholesteryl ester transfer protein (CETP)) are mainly present in LpA-I and not in LpA-I:A-II family. Cholesterol efflux mediated by A-I-containing particles has been studied in different cells. Long term exposure to LpA-I family promoted cholesterol efflux whereas less efflux was observed in the presence of LpA-I:A-II family. The fact that LpA-I:A-II family can inhibit the LpA-I promoted cholesterol efflux strongly supports the role of apo A-II as an antagonist in the production of cholesterol efflux. These results which emphasize that LpA-I and LpA-I:A-II families behave as distinct entities have been confirmed in other studies showing that they have different clinical significance. The results in mice transgenic for apo A-I indicate that overexpression of apo A-I induces more cholesterol efflux and protects C57BL/6 mice from atherosclerosis. Increased expression of apo A-II in mice appears to decrease cholesterol efflux and to promote rather than retard aortic fatty streak development.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Tangier disease: isolation and characterization of LpA-I, LpA-II, LpA-I: A-II and LpA-IV particles from plasma.

Tangier disease (TD) is characterized by extremely low plasma levels of HDL, apoA-I and apoA-II due to very rapid catabolism. However, the risk of premature coronary heart disease (CHD) is not markedly increased in TD. In order to gain insight into reverse cholesterol transport in TD, we isolated LpA-I, LpA-I:A-II, LpA-II and LpA-IV particles from fasting plasma of 5 TD patients. LpA-I composition was similar to control LpA-I, but TD LpA-I had more LCAT and CETP activity (respectively, 0.35 +/- 0.14 and 0.14 +/- 0.04 mumol of cholesterol esterified/h/micrograms of protein, and 7 +/- 2.5 and 1.4 +/- 0.3 mumol of cholesteryl ester transferred/h/micrograms of protein). In contrast, TD LpA-I:A-II had abnormal composition, with a low molar ratio of apoA-I to apoA-II (0.2-1.33). In addition, LpA-I:A-II in TD contained a substantial amount of apoA-IV compared with control, making this particle an LpA-I:A-II:A-IV complex. LpA-I:A-II from normal plasma do not promote cholesterol efflux from adipocytes cells, whereas TD LpA-I:A-II:A-IV complexes promoted cholesterol efflux from these cells. Moreover LpA-I:A-II:A-IV complexes have more LCAT and CETP activity than control (respectively 1.2 +/- 0.16 and 0.05 +/- 0.01 mumol of cholesterol esterified/h/micrograms of protein and, 41 +/- 3.7 and 1 +/- 0.4 mumol of cholesteryl ester transferred/h/micrograms of protein). The LpA-II particle in TD represented in fact an LpA-II:A-IV complex (75% mol apoA-II and 22% mol apoA-IV).(ABSTRACT TRUNCATED AT 250 WORDS)

Adipose Tissue↗