[Spectral analysis of the electroencephalogram in subjects with mild cognitive impairment]. / Análisis espectral del electroencefalogramaen sujetos con deterioro cognitivo leve.

INTRODUCTION: Mild cognitive impairment (MCI) is defined as a state of the ageing brain midway between normal cognition and dementia. Special attention has been paid to the electrophysiological substrate underlying Alzheimer's disease and MCI in order to identify as early as possible which subjects with MCI progress towards Alzheimer, which could be crucial for starting rehabilitation or pharmacological therapies. AIM: To perform a spectral characterisation of the electroencephalogram in subjects with MCI. PATIENTS AND METHODS: An electroencephalogram was carried out on 41 subjects with MCI in order to analyse the spectral measurements; apolipoprotein E genotype was also performed. RESULTS: In all, 94.8% of the sample displayed a significant increase in energy, and in 66.6% of them this was observed in the theta and delta bands, or both. Significant differences were found in the spectral measurements between carriers and non-carriers of the apolipoprotein E epsilon4 allele in the theta and alpha bands; there was also a statistically significant association between the years of schooling and being a carrier of this allele or not. An increase in the theta-alpha bands was observed in the left temporal region in subjects with a below-average number of years of schooling and carriers of the epsilon4 allele. CONCLUSIONS: In subjects with MCI and carriers of the epsilon4 allele, the alpha and theta cortical rhythms can be affected by similar pathological mechanisms and may be expressed earlier in subjects who have a lower level of schooling.

Análisis espectral del electroencefalograma en sujetos con deterioro cognitivo leve / Spectral analysis of the electroencephalogram in subjects with mild cognitive impairment

El deterioro cognitivo leve (DCL) se define como un estado del cerebro envejecido intermedio entrela cognición normal y la demencia; se ha prestado una especial atención al sustrato electrofisiológico de la enfermedad de Alzheimer y el DCL con la finalidad de identificar lo antes posible qué sujetos con DCL progresan a Alzheimer, lo cual podríaser crucial para el inicio de terapias de rehabilitación o farmacológicas. Objetivo. Caracterización espectral del electroencefalograma en sujetos con DCL. Pacientes y métodos. Se estudió a 41 sujetos con DCL a los cuales se les realizó electroencefalograma para el análisis de las medidas espectrales; además se realizó genotipo de apolipoproteína E. Resultados. El 94,8% de la muestra reveló un incremento significativo de energía; en el 66,6% de ellos se observó en las bandas theta y delta,o ambas. Se encontraron diferencias significativas en las medidas espectrales entre portadores y no portadores del alelo epsilon4 de la apolipoproteína E en las bandas theta y alfa, y además existió una asociación estadísticamente significativa entre los años de escolaridad y ser portador o no de dicho alelo. Se observó un incremento en las bandas theta-alfa en la región temporal izquierda en los sujetos por debajo de la media de años de escolaridad y portadores del alelo epsilon4. Conclusiones. En los sujetos con DCL y portadores del alelo epsilon4, los ritmos corticales alfa y theta pueden verse afectados por mecanismospatológicos similares, y puede expresarse de forma más precoz en los sujetos que presentan un nivel de escolaridad bajo

Mild cognitive impairment (MCI) is defined as a state of the ageing brain midway between normalcognition and dementia. Special attention has been paid to the electrophysiological substrate underlying Alzheimer’s disease and MCI in order to identify as early as possible which subjects with MCI progress towards Alzheimer, which could be crucial forstarting rehabilitation or pharmacological therapies. Aim. To perform a spectral characterisation of the electroencephalogram in subjects with MCI. Patients and methods. An electroencephalogram was carried out on 41 subjects with MCI in order to analyse the spectral measurements; apolipoprotein E genotype was also performed. Results. In all, 94.8% of the sampledisplayed a significant increase in energy, and in 66.6% of them this was observed in the theta and delta bands, or both. Significant differences were found in the spectral measurements between carriers and non-carriers of the apolipoprotein E epsilon4 allele in the theta and alpha bands; there was also a statistically significant association between the years of schoolingand being a carrier of this allele or not. An increase in the theta-alpha bands was observed in the left temporal region in subjects with a below-average number of years of schooling and carriers of the epsilon4 allele. Conclusions. In subjects withMCI and carriers of the epsilon4 allele, the alpha and theta cortical rhythms can be affected by similar pathological mechanisms and may be expressed earlier in subjects who have a lower level of schooling

Synthesis of a new 5'-O-triphosphate analog of 5-methyl 2'-O-deoxycytidine. Preliminary in vitro labelling for non-radioactive detection of DNA.

We report the synthesis of the triphosphate of 5-methyl 4-N-[6-(p-bromobenzamido)hex-1-yl]-2'-O-deoxycytidine 3A. We also analyzed the formation of intramolecular H-bonds of 5-methyl 4-N-[n-[6-(p-bromobenzamido) caproyl amino]alk-1-yl]-2'-deoxycytidine compounds, and confirmed their presence by 1H-NMR studies. In vitro DNA labeling with modified nucleotides is preliminarily evaluated.

Circular and linear DNA molecules in the Entamoeba histolytica complex molecular karyotype.

Entamoeba histolytica genome was analysed by pulsed field gel electrophoresis under conditions to separate linear chromosomes in the 170-1400 kb range. We identified linear DNA molecules of 227, 366, 631, 850, 1112 and 1361 kb (mean sizes obtained by three different methods) and we estimated their reorientation times and migration velocities at various experimental conditions. DNA shift mobility assays, using ethidium bromide, suggested that bands migrating at 227 and 631 kb contain linear and circular DNA, whereas a band at 436 kb has only circular DNA. We obtained a regression equation relating sizes of supercoiled DNA molecules with their migration velocities during a pulse at constant electric field and temperature. We also developed a computer program (EHPATTERNS) that predicts the migration per pulse and the resolution order of circular and linear E. histolytica DNA at different pulse times and constant driving and frictional forces. The simulation showed that linear DNA molecules frequently co-migrate with circular molecules, but circular molecules change when the pulse time varies. This molecular mixture generates broad bands and difficulties in the interpretation of the molecular karyotype of E. histolytica.

Synthesis of a new analog of thymidine for in vivo non-radioactive labeling of DNA.

The introduction of 6-(p-bromobenzoylamino)caproyl radical in the methyl group of 2'-O-deoxythymidine is described. In vivo incorporation of this nucleoside to DNA was determined using a monoclonal antibody that recognized the radical.

Comparison of DNA migrations in two clamped homogeneous electric field chambers of different sizes. Relation between sample thickness and electrophoresis time.

We present here a method to compare the mathematical descriptions of DNA migration per pulse as a function of pulse time. It is based on obtaining robust estimates and variances of DNA reorientation time, migration velocities during and after DNA reorientation; and on the statistical comparisons of these estimates. We demonstrated an equal description for the migration per pulse of each DNA molecule separated under identical conditions in clamped homogeneous electric field (CHEF) and miniCHEF chambers. However, miniCHEF resolved the patterns in shorter times, because it uses thinner samples. The relationship between sample thickness and CHEF run time is also presented.

A novel cytoplasmic structure containing DNA networks in Entamoeba histolytica trophozoites.

We report here the presence of cytoplasmic DNA arranged in networks in the trophozoites of the human parasite Entamoeba histolytica. Cytoplasmic DNA was detected in live trophozoites in a structure that we called EhkO, using the fluorescent dye acridine orange, and by in situ hybridization to trophozoites with a rDNA probe. The EhkO was found in the axenically grown clones A, L6 (strain HMI:IMSS) and MAVax (strain MAV) and in the polyxenically grown clone MAVpx (strain MAV). Bacteria present in MAVpx did not cross hybridize with the DNA probe neither in in situ hybridization or in Southern blot experiments. Autoradiography of metabolically [3H]thymidine-labeled trophozoites showed the presence of EhkO, and an EhkO-enriched fraction, purified from a nuclei-free extract and examined by light microscopy, exhibited [3H]thymidine incorporation into this structure. DNA was purified from the EhkO and enriched nuclear fractions and analyzed by transmission electron microscopy. The EhkO fraction contained DNA networks resembling those of trypanosome kDNA, whereas nuclear DNA was present mainly as linear molecules and some circles. Our findings imply that E. histolytica may be taxonomically more closely related to the Trypanosomatidae than previously suspected.

Cytoplasmic DNA in Entamoeba histolytica: its biological significance.

We report a study on the DNA organization in Entamoeba histolytica using a ribosomal DNA probe. The rDNA genes were found forming mers which were separated in a typical ladder pattern by pulse field electrophoresis. DNA rosette structures were visualized through electron microscopy in DNA eluted from bands recognized by the ribosomal probe. The in situ hybridization experiments using a DNA probe suggested that the rDNA genes are portioned between the nucleus and a cytoplasmic structure. These findings provide new data on DNA organization in E. histolytica and open the question concerning the presence of a novel organelle in this eukaryotic parasite.

Entamoeba histolytica: gene linkage groups and relevant features of its karyotype.

We identified some gene linkage groups in Entamoeba histolytica using a 4-M urea improved transversal alternating field electrophoresis (TAFE) method. Complex rosette-structured DNA molecules were found trapped along the gel lanes, explaining the fuzziness of the patterns. Using several episomal probes, including 16 S, 5.8 S, and 25 S ribosomal (r)Dna genes, an autonomous replication sequence (ARS), and EhVR1, we identified a complete ribosomal episome linkage group (CELG) at the 1.2-Mb position. Three other incomplete groups were found: IELG-1, formed by EhVR1, 16 S, 5.8 S, and 25 S genes; IELG-2 formed by EhVR1, 16 S and 25 S; and IELG-3 formed only by 5.8 S. Ehadh3, Ehpfo, and Ehredox genes migrated at the 1.8-Mb position, forming the non-ribosomal linkage group, NRLG-1.8, while the Ehenl-1 gene migrated at 1.6 Mb forming the NRLG-1.6 group. Ehhk was located at 1.2, 0.8, and 0.17 Mb in three different groups: NRLG-1.2, IELG-3-0.8, and NRLG-0.17. Putative lineal chromosomes were also identified using an heterologous telomeric probe. By in situ hybridization experiments, the rDNA and Ehhk genes were located in both nucleus and cytoplasm, while the Ehpfo and Ehredox genes were found mainly in the nucleus. We propose a model hypothezising that the 16 S and 25 S genes are in a linear molecule, duplicated in two inverted repeats, which may be looped out of the linear DNA to form an episome probably lacking or not the 5.8 S sequence, which could be added later by recombination.

Pvu II RFLPs of LDL receptor gene in normolipidemic subjects from Havana City.

Allele and genotype frequencies for Pvu II restriction fragment length polymorphism of LDL receptor gene were determined in 36 normolipidemic unrelated subjects from Havana City. The frequencies were similar to those reported for other populations.

Gene amplification in Entamoeba histolytica.

We show here data suggesting that Entamoeba histolytica, the protozoan responsible for human amebiasis, presents DNA amplification in a fashion similar to that described for transformed mammalian cells. By transmission electron microscopy (TEM), we found linear, circular and concentric circular DNA molecules exhibiting the main events of the unscheduled DNA amplification process. Loops were formed after the recombination of two nonadjacent DNA regions, and bubbles appeared from the recombinant strands without involving the looped-out sequences. Bubbles grew up and underwent further replication rounds to produce a nested set of partially replicated circles. Multicircle complexes were also formed from putative replication origin without recombination of distant DNA regions. Clones derived from the strain HM1:IMSS exhibited different DNA contents, suggesting DNA amplification. The parental clone A and its daughter clone C2 differed in rDNA gene copy numbers, but this was observed only when total DNA was separated by pulse field gel electrophoresis, and no significant differences were detected in nuclear DNA. The dissection of the events observed by TEM led us to propose an onion skin model for gene amplification in E. histolytica.

Amplificación del gen de la ADN polimerasa I por multiplicación del Lambda polA mediante ciclo lítico en la E. coli WK6: obtención y purificación de la enzima / Amplification of the gene for DNA polimerasa I by the multiplication of Lambda polA in E. coli WK6: obtention and purification of the enzyme

El fago Lambda polA (Qam 73, Sam 7) fue aislado y purificado de un lisógeno de la E. coli 594, y posteriormente se multiplicó en una cepa SupE, SupF. Se obtuvo una cantidad considerable de DNA polimerasa I multiplicándolo por vía ciclo lítico, tras la infección a alto MOI (Multiplicity of Infection) de una cepa libre de supresores amber (E. coli WK6). En este trabajo se muestran los resultados de la estandarización de las condiciones para obtener un nivel adecuado de expresión del gen polA, el esquema de purificación utilizado y se discuten las modificaciones realizadas para la obtención de la DNA polimerasa I a gran escala. La DNA pol I purificada incorporó P32-dATP por Nick translation en fragmentos de DNA entre 800-50 000 bp con un alto marcaje específico

Un prototipo para la electroforesis de campo pulsante. Estandarización de las condiciones para la resolución de grandes moléculas de ADN / A prototype for the pulse field electrophoresis: condition for the resolution of large DNA molecules

Se desarrolló un prototipo para la electroforesis de campo pulsante en gel de agarosa, que permite separar moléculas de ADN de hasta 2 000 kb. El equipo se fundamenta en la propiedad que tienen las moléculas grandes de ADN, de fraccionarse en geles de agarosa si son sometidas alternadamente a dos campos eléctricos no homogéneos de orientación perpendicular. En los experimentos, los tiempos de pulso fueron 60s, y se estudió la influencia de la fuerza iónica y el tiempo de corrida sobre la resolución de las bandas. El método fue utilizado para obtener el cariotipo electroforético de dos cepas de Saccharomyces cerevisiae. El trabajo describe las distintas partes del equipo, el protocolo electroforético, y la caracterización parcial de algunas bandas mediante hibridización ADN-ADN con sondas específicas para diferentes cromosomas