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Biomedical subjects

A Benito

Publications and source records attributed to A Benito.

16 recordsLinked to original sources
PubMed1995

A recombinant foot-and-mouth disease virus antigen inhibits DNA replication and triggers the SOS response in Escherichia coli.

The 3D gene of foot-and-mouth disease virus encodes the viral RNA dependent RNA polymerase, also called virus infection associated (VIA) antigen, which is the most important serological marker of virus infection. This 3D gene from a serotype C1 virus has been cloned and overexpressed in Escherichia coli under the control of the strong lambda lytic promoters. The resulting 51 kDa recombinant protein has been shown to be immunoreactive with sera from infected animals. After induction of gene expression, an immediate and dramatic arrest of cell DNA synthesis occurs, similar to that produced by genotoxic doses of the drug mitomycin C. This effect does not occur during the production of either a truncated VIA antigen or other related and non-related viral proteins. The inhibition of DNA replication results in a subsequent induction of the host SOS DNA-repair response and in an increase of the mutation frequency in the surviving cells.

Capsid↗
PubMed1995

Mitomycin C stimulates thermally induced recombinant gene expression in Escherichia coli MC strains.

The effects of mitomycin C on C1857-controlled recombinant gene expression have been explored in E. coli cultures when the drug was added simultaneously to the thermal induction. A significantly improved yield of homologous, heterologous and chimeric fusion proteins was observed in E. coli MC1061 and GE864 (a MC4100 derivative) thermoinduced cells. This feature was not detected in other E. coli strains and does not involve a gene dosage mechanism but a strain-dependent stimulation of gene expression unrelated to the RecA protease activity.

DNA, Bacterial↗
PubMed1995

In vitro susceptibility of Plasmodium falciparum to chloroquine, amodiaquine, quinine, mefloquine, and sulfadoxine/pyrimethamine in Equatorial Guinea.

Between March 1990 and June 1992, a study was carried out in Equatorial Guinea on the in vitro response of Plasmodium falciparum to different antimalarial drugs. Field work for the study was conducted both in the country's island region as well as on the mainland, and resistant isolates were found to exhibit interregional differences. On the island of Bioko, 204 tests were performed with 16% (11 of 69) resistant to chloroquine, 9% (4 of 46) resistant to quinine, 14% (6 of 43) resistant to a combination of sulfadoxine/pyrimethamine, and 6.5% (3 of 46) resistant to amodiaquine. In the mainland area of Bata, the same antimalarial drugs and mefloquine were tested with the following results: 9% (5 of 58) resistant to chloroquine; 2% (1 of 58) resistant to amodiaquine, and 3% (2 of 58) resistant to a combination of sulfadoxine/pyrimethamine. No isolates resistant to quinine or mefloquine were found. Effective concentrations (EC50, EC90, and EC99) and regression lines (log dose/response) for each antimalarial drug were calculated to establish a surveillance system for antimalarial drug chemosensitivity in Equatorial Guinea. Finally, 12 isolates from 12 patients previously treated with chloroquine were studied to compare both tests (in vivo-in vitro) and obtain a correlation between the RII and RIII types of in vivo and in vitro resistances. No correlation for the RI type was found between the two methods.

Amodiaquine↗
PubMed1995

Regulation and function of Bcl-2 during differentiation-induced cell death in HL-60 promyelocytic cells.

Polymorphonuclear leukocytes are generated by differentiation of early myeloid precursors. Once fully differentiated, blood neutrophils are programmed to die rapidly and are removed by tissue macrophages. In normal myeloid cells, the death mechanism seems to be coupled to the differentiation pathway and is accomplished by a process termed apoptosis. In the present study, we have examined the role of Bcl-2 in the differentiation pathways of the promyelocytic cell line HL-60. Treatment of HL-60 with retinoic acid or phorbol ester, which induced neutrophil or macrophage-like cell differentiation, respectively, resulted in progressive loss of cellular viability and internucleosomal DNA degradation. In HL-60, differentiation and apoptosis were coupled to down-regulation of the Bcl-2 protein. Overexpression of Bcl-2 by gene transfer inhibited apoptosis triggered by terminal differentiation of HL-60. Yet, Bcl-2 did not alter the expression of surface markers or other phenotypic changes that are induced upon myeloid differentiation. In contrast to HL-60, another immature myeloid cell line, K562, did not produce Bcl-2 but expressed a related protein, Bcl-xL, that functions as a repressor of apoptotic cell death. K562 has been shown to be relatively resistant to a variety of apoptotic stimuli. Incubation of HL-60 and K562 with inhibitors of macromolecular synthesis induced apoptosis, which appeared earlier in HL-60 than in K562. Interestingly, Bcl-2 overexpression protected K562 cells from apoptosis induced by inhibitor of macromolecular synthesis but it had little or no effect on HL-60 cells. We conclude that although differentiation and apoptosis proceed simultaneously, they can be uncoupled by expression of Bcl-2. Down-regulation of Bcl-2 appears to be part of the differentiation pathway and may serve to facilitate the apoptotic response.

Apoptosis↗
PubMed1994

Insertion of a 27 amino acid viral peptide in different zones of Escherichia coli beta-galactosidase: effects on the enzyme activity.

Seven internal, putatively exposed regions of Escherichia coli beta-galactosidase have been explored regarding their tolerance to insertions of large foreign peptides. Small sequence modifications, including amino acid substitutions and small deletions, were introduced into the lacZ gene to generate unique BamHI restriction sites. By using these mutant genes, a 27 amino acid stretch reproducing the hypervariable loop of foot-and-mouth disease virus VP1 protein (site A) was further inserted in predefined regions of the enzyme. Among the 13 resulting engineered proteins only three, carrying sequence modifications within a short region, are active, with only moderate reduction of their specific activities. The identified permissive region, which involves amino acids 275 to 279, seems to be a flexible area that could be appropriate to incorporate and study biological properties of heterologous peptides in correctly folded beta-galactosidase chimeric proteins.

Base Sequence↗
PubMed1994

[Molecular heterogeneity of beta thalassemia].

PURPOSE: To check out the incidence of beta-thalassaemia in different regions of Spain and to demonstrate its heterogeneity. PATIENTS AND METHODS: The study was performed on 60 peripheral blood samples from patients diagnosed of beta-thalassaemia by conventional methods, namely, microcytosis, HbA2 over 3.5%, increased HbF. DNA was isolated with phenol-chloroform and amplified by ARMS technique, and in one case its sequence was established according to Sanger's method. RESULTS: The mutations found in the 67 alleles studied showed similar incidence, although NT1 (IVS 1) mutation was increased and NT110 (IVS 1) mutation was decreased when compared with the findings in other Mediterranean populations. CONCLUSION: Due to historical reasons, the racial characteristics of Spanish people are the product of different ethnic groups, so hereditary disorders are here usually heterogeneous. This study appears to confirm such fact with respect to beta-thalassaemia.

Genetic Heterogeneity↗
PubMed1994

Application and evaluation of QBC malaria diagnosis in a holoendemic area.

Quantitative Buffy Coat Analysis was used in practical diagnosis situations. In the first place, sensitivity and specificity was measured in an active search survey (720 schoolchildren under 10 years of age, in nine field trips) under unfavourable laboratory conditions (onsite school diagnosis being conducted with electricity supplied by a diesel-powered generator). In this group, the QBC-based study revealed sensitivity and specificity ratings of 99.6% and 81.7% respectively. QBC proved 5.5% more sensitive than the thick-film method, but specificity was greater with the latter. QBC returned good results when used in adverse situations, viz., in the absence of air conditioning and with a diesel generator as the only power supply. QBC was used in a passive search survey (hospital group, 618 children) under good laboratory conditions (air conditioning) with patients enrolled in a trial to measure resistance to antimalarial drugs. In this survey we targeted at assessing the efficacy of chloroquine treatment at dose of 25 mg/kg/3 days. QBC proved more sensitive than the thick-film method, detecting - on day 14 of the in vivo test - low parasitaemias that had gone undetected by thick film. Lastly, this study reports on the conversion of QBC readings (parasitaemia per field) into thick-field terms (number of parasites per microliter of blood), with the aim of measuring the degree of recurring parasitaemia.

Acridine Orange↗
PubMed1993

Uses of beta-galactosidase tag in on-line monitoring production of fusion proteins and gene expression in Escherichia coli.

A simple method for monitoring and quantifying automatically the production by fermentation of beta-galactosidase fusion proteins, making use of the remaining activity of the beta-galactosidase part, is considered. A hybrid protein carrying the major antigenic domain of foot-and-mouth disease virus C1 joined at the N-terminus of beta-galactosidase has been expressed in Escherichia coli. The yield of the chimeric protein has been monitored by flow injection analysis (FIA) during batch fermentations at laboratory scale, and a high correlation between values of product concentration from FIA and from immunological quantizations has been obtained. Because of the possibility of employing FIA in large-scale experiments, and the high sampling frequency, versatility, and reproducibility offered by this method, we propose FIA as a general, simple, quick, flexible, and reliable instrument for both monitoring the yield of recombinant proteins produced industrially, and performing basic research at laboratory scale.

Aphthovirus↗
PubMed1993

Enhanced production of pL-controlled recombinant proteins and plasmid stability in Escherichia coli RecA+ strains.

Overexpression of pL-controlled foot-and-mouth disease virus recombinant proteins was studied in Escherichia coli RecA+ strains and in a recA mutant. Higher protein yield and extractable plasmid DNA amounts were found in wild type cells, in absence of detectable RecA proteolytic activity. Minor but still significant differences in pBR322 DNA amounts were also detected between RecA+ and its recA13 and lexA1 derivatives. These data should be seriously considered to select expression systems and to design production processes for recombinant proteins, specially if they are expected to be toxic for Escherichia coli cells.

Aphthovirus↗
PubMed1987

[A case of chloroquine-resistant malaria of type R3 originating in Cameroon].

World areas with chloroquine-resistant falciparum malaria are progressively spreading, in Africa from East to West. We are reporting here a new case of resistance, grade III, from Cameroon, carried out in vitro by the WHO standard macrotest. Resistance is determined by the ability of trophozoites to develop into schizonts when therapeutic doses of chloroquine have been administered.

Africa, Western↗
PubMed1984

[Normal blood pressure values in Spanish children].

Authors study 2,953 children with a range of age between 2 and 14 years, of both sexes, from schools in Madrid, Zaragoza, Lugo, Andújar, Archena and in rural areas of Guadalajara and Segovia. Blood pressure was measures three times in every child according to recommendations of the Expert Committee of WHO in respect to the method. Results are expressed in percentiles according to age and sex. Their findings indicate that both diastolic and systolic blood pressures were lower than those reported by Task Force (USA), Cassimos (Greece) and Andre (France) (p less than 0.001) with higher differences being present in the diastolic blood pressure.

Adolescent↗
PubMedDate not supplied

Ammonium-mediated reduction of plasmid copy number and recombinant gene expression in Escherichia coli.

The effect of ammonium as a medium supplement on plasmid-encoded recombinant beta-galactosidase synthesis was explored in Escherichia coli cells during aerobic growth in complex medium. After induction, only doses of ammonium chloride below 1 g/L are able to transiently enhance the yield. However, the presence of nontoxic ammonium chloride concentrations of up to 10 g/L results in lower values of beta-galactosidase in a concentration-dependent fashion. A significant reduction in plasmid DNA content explains the decrease in the yield by a gene-dosage-involving mechanism.

Ammonium Chloride↗