Experiencia en nefrectomía: serie de casos de cuatro años

Introducción: Las indicaciones por las cuales un paciente requiere una nefrectomía son múltiples: las neoplasias, la hidronefrosis y la exclusión funcional son las principales. En manos expertas la nefrectomía es un procedimiento seguro, especialmente porque en la actualidad el abordaje por excelencia es realizar una técnica mínimamente invasiva con conservación de nefronas. Se presenta el análisis de la experiencia en Mederi, Hospital Universitario Mayor en esta intervención. Metodología: Se realizó una serie de casos de pacientes llevados a nefrectomía entre mayo de 2008 y mayo de 2012. Se incluyeron la totalidad de los casos. Resultados: Se analizaron 72 registros, 49 mujeres y 25 hombres; 13 de ellas fueron laparoscópicas. La edad promedio fue de 58,6 años. El tiempo medio operatorio fue 169,23 minutos (118-220 minutos). El sangrado operatorio promedio fue de 680,63 ml (IC95%: 2,83-1358 ml). El tiempo de hospitalización promedio fue de 4,88 días IC95%. La mayoría de los pacientes se distribuyeron en estadios medios de la enfermedad tumoral, con poco compromiso ganglionar y metástasis; el diagnóstico histológico y estadio dominante fueron el carcinoma de células renales grado 3 de Fuhrman respectivamente. Se reportan 13 casos de compromiso de la capsula de Gerota y 11 con compromiso del hilio. Discusión: La experiencia en nefrectomía de la institución es muy positiva por el bajo número de mortalidad y complicaciones. En cuanto a la técnica, es importante promover la técnica laparoscópica

Polimorfismos del gen Butirilcolinesterasa responsables de reacciones adversas en pacientes consumidores de “cocaína”

La butirilcolinesterasa humana (BChE; EC 3.1.1.8) es una enzima polimórfica sintetizada en el hígado y en el tejido adiposo, ampliamente distribuida en el organismo y encargada de hidrolizar algunos ésteres de colina como la procaína, ésteres alifáticos como el ácido acetilsalicílico, fármacos como la metilprednisolona, el mivacurium y la succinilcolina y drogas de uso y/o abuso como la heroína y la cocaína. Es codificada por el gen BCHE (OMIM 177400), habiéndose identificado más de 100 variantes, algunas no estudiadas plenamente, además de la forma más frecuente, llamada usual o silvestre. Diferentes polimorfismos del gen BCHE se han relacionado con la síntesis de enzimas con niveles variados de actividad catalítica. Las bases moleculares de algunas de esas variantes genéticas han sido reportadas, entre las que se encuentra las variantes Atípica (A), fluoruro-resistente del tipo 1 y 2 (F-1 y F-2), silente (S), Kalow (K), James (J) y Hammersmith (H). En este estudio, en un grupo de pacientes se aplicó el instrumento validado Lifetime Severity Index for Cocaine Use Disorder (LSI-C) para evaluar la gravedad del consumo de “cocaína” a lo largo de la vida. Además, se determinaron Polimorfismos de Nucleótido Simple (SNPs) en el gen BCHE conocidos como responsables de reacciones adversas en pacientes consumidores de “cocaína” mediante secuenciación del gen y se predijo el efecto delos SNPs sobre la función y la estructura de la proteína, mediante el uso de herramientas bio-informáticas. El instrumento LSI-C ofreció resultados en cuatro dimensiones: consumo a lo largo de la vida, consumo reciente, dependencia psicológica e intento de abandono del consumo. Los estudios de análisis molecular permitieron observar dos SNPs codificantes (cSNPs) no sinónimos en el 27.3% de la muestra, c.293A>G (p.Asp98Gly) y c.1699G>A (p.Ala567Thr), localizados en los exones 2 y 4, que corresponden, desde el punto de vista funcional, a la variante Atípica (A) [dbSNP: rs1799807] y a la variante Kalow (K) [dbSNP: rs1803274] de la enzima BChE, respectivamente. Los estudios de predicción In silico establecieron para el SNP p.Asp98Gly un carácter patogénico, mientras que para el SNP p.Ala567Thr, mostraron un comportamiento neutro. El análisis de los resultados permite proponer la existencia de una relación entre polimorfismos o variantes genéticas responsables de una baja actividad catalítica y/o baja concentración plasmática de la enzima BChE y algunas de las reacciones adversas ocurridas en pacientes consumidores de cocaína.

Destabilizing Mutations Alter the Hydrogen Exchange Mechanism in Ribonuclease A

The effect of strongly destabilizing mutations, I106A and V108G of Ribonuclease A (RNase A), on its structure and stability has been determined by NMR. The solution structures of these variants are essentially equivalent to RNase A. The exchange rates of the most protected amide protons in RNase A (35ºC), the I106A variant (35ºC), and the V108G variant (10ºC) yield stability values of 9.9, 6.0, and 6.8 kcal/mol, respectively, when analyzed assuming an EX2 exchange mechanism. Thus, the destabilization induced by these mutations is propagated throughout the protein. Simulation of RNase A hydrogen exchange indicates that the most protected protons in RNase A and the V108G variant exchange via the EX2 regime, whereas those of I106A exchange through a mixed EX1 1 EX2 process. It is striking that a single point mutation can alter the overall exchange mechanism. Thus, destabilizing mutations joins high temperatures, high pH and the presence of denaturating agents as a factor that induces EX1 exchange in proteins. The calculations also indicate a shift from the EX2 to the EX1 mechanism for less protected groups within the same protein. This should be borne in mind when interpreting exchange data as a measure of local stability in less protected regions

Dynamics of QoI sensitivity in Mycosphaerella fijiensis in Costa Rica during 2000 to 2003

From 1997 onward, the strobilurin fungicide azoxystrobin was widely used in the main banana-production zone in Costa Rica against Mycosphaerella fijiensis var. difformis causing black Sigatoka of banana. By 2000, isolates of M. fijiensis with resistance to the quinolene oxidase inhibitor fungicides were common on some farms in the area. The cause was a single point mutation from glycine to alanine in the fungal target protein, cytochrome b gene. An amplification refractory mutation system Scorpion quantitative polymerase chain reaction assay was developed and used to determine the frequency of G 143A allele in samples of M. fijiensis. Two hierarchical surveys of spatial variability, in 2001 and 2002,found no significant variation in frequency on spatial scales <10 in. This allowed the frequency of G143A alleles on a farm to be estimated efficiently by averaging single samples taken at two fixed locations. The frequency of G 143A allele in bulk samples from I I farms throughout Costa Rica was determined at 2-month intervals. There was no direct relationship between the number of spray applications and the frequency of G143A on individual farms. Instead, the frequency converged toward regional averages, presumably due to the large-scale mixing of ascospores dispersed by wind. Using trap plants in an area remote from the main producing area, immigration of resistant ascospores was detected as far as 6 km away both with and against the prevailing wind.

The genome strikes back: The evolutionary importance of defence against mobile elements

Increasingly, we regard the genome as a site and source of genetic conflict. This fascinating 'bottom-up' view brings up appealing connections between genome biology and whole-organism ecology, in which populations of elements compete with one another in their genomic habitat. Unlike other habitats, though, a host genome has its own evolutionary interests and is often able to defend itself against molecular parasites. Most well-studied organisms employ strategies to protect their genomes against the harmful effects of genomic parasites, including methylation, various pathways of RNA interference, and more unusual tricks such as repeat induced point-mutation (RIP). These genome defence systems are not obscure biological curiosities, but fundamentally important to the integrity and cohesion of the genome, and exert a powerful influence on genome evolution.

The structure of interrupted human AC microsatellites

Microsatellite lengths change over evolutionary time through a process of replication slippage. A recently proposed model of this process holds that the expansionary tendencies of slippage mutation are balanced by point mutations breaking longer microsatellites into smaller units and that this process gives rise to the observed frequency distributions of uninterrupted microsatellite lengths. We refer to this as the slippage/point-mutation theory. Here we derive the theory's predictions for interrupted microsatellites comprising regions of perfect repeats, labeled segments, separated by dinucleotide interruptions containing point mutations. These predictions are tested by reference to the frequency distributions of segments of AC microsatellite in the human genome, and several predictions are shown not to be supported by the data, as follows. The estimated slippage rates are relatively low for the first four repeats, and then rise initially linearly with length, in accordance with previous work. However, contrary to expectation and the experimental evidence, the inferred slippage rates decline in segments above 10 repeats. Point mutation rates are also found to be higher within microsatellites than elsewhere. The theory provides an excellent fit to the frequency distribution of peripheral segment lengths but fails to explain why internal segments are shorter. Furthermore, there are fewer microsatellites with many segments than predicted. The frequencies of interrupted microsatellites decline geometrically with microsatellite size measured in number of segments, so that for each additional segment, the number of microsatellites is 33.6% less. Overall we conclude that the detailed structure of interrupted microsatellites cannot be reconciled with the existing slippage/point-mutation theory of microsatellite evolution, and we suggest that microsatellites are stabilized by processes acting on interior rather than on peripheral segments.

The impact of genome defense on mobile elements in Microbotryum

Repeat induced point mutation (RIP), a mechanism causing hypermutation of repetitive DNA sequences in fungi, has been described as a ‘genome defense’ which functions to inactivate mobile elements and inhibit their deleterious effects on genome stability. Here we address the interactions between RIP and transposable elements in the Microbotryum violaceum species complex. Ten strains of M. violaceum, most of which belong to different species of the fungus, were all found to contain intragenomic populations of copia-like retrotransposons. Intragenomic DNA sequence variation among the copia-like elements was analyzed for evidence of RIP. Among species with RIP, there was no significant correlation between the frequency of RIP-induced mutations and inferred transposition rate based on diversity. Two strains of M. violaceum, from two different plant species but belonging to the same fungal lineage, contained copia-like elements with very low diversity, as would result from a high transposition rate, and these were also unique in showing no evidence of the hypermutation patterns indicative of the RIP genome defense. In this species, evidence of RIP was also absent from a Class II helitron-like transposable element. However, unexpectedly the absolute repetitive element load was lower than in other strains.

Emergence of fluoroquinolone resistance in the native Campylobacter coli population of pigs exposed to enrofloxacin

Objective: The effect of a single 5 day enrofloxacin treatment on the native Campylobacter coli population in conventionally weaned 5-week-old pigs was investigated. Materials: Twelve pigs were split into two groups of six: one group was treated with a therapeutic dose (15 mg/pig/day) of enrofloxacin and the other remained untreated to act as the control. Campylobacter coli were isolated from faecal samples and tested for ciprofloxacin resistance by measuring MIC values. Mutations in the quinolone resistance-determining region (QRDR) of the gyrA gene of resistant isolates were identified by sequencing and denaturing HPLC. Levels of enrofloxacin and its primary metabolite ciprofloxacin in the pig faeces were also measured by HPLC. Results: No quinolone-resistant C. coli (n = 867) were detected in any of the pigs prior to treatment, indicating <0.1% resistance in the group. Resistant C. coli were isolated from pigs for up to 35 days after treatment with a therapeutic dose. These resistant C. coli had MIC values of 128 mg/L and 8-16 mg/L for nalidixic acid and ciprofloxacin, respectively, and the same single point mutation causing a Thr-86 to Ile substitution in the QRDR was identified in each. The concentration of enrofloxacin in the pig faeces was 2-4 mug/g faeces for the duration of the 5 day therapeutic treatment and was detected up to 10 days post-treatment. Ciprofloxacin was also measured and peaked at 0.6 mug/g faeces in the treated group. Conclusion: This study provides evidence that a single course of enrofloxacin treatment contributes directly to the emergence and persistence of fluoroquinolone resistance in C. coli.

Molecular requirements for L-type Ca2+ channel blockade by testosterone

Despite being generally perceived as detrimental to the cardiovascular system, testosterone has marked beneficial vascular effects; most notably it acutely and directly causes vasodilatation. Indeed, men with hypotestosteronaemia can present with myocardial ischemia and angina which can be rapidly alleviated by infusion of testosterone. To date, however, in vitro studies have failed to provide a convincing mechanism to account for this clinically important effect. Here, using whole-cell patch-clamp recordings to measure current flow through recombinant human L-type Ca2+ channel alpha(1C) subunits (Ca(v)1.2), we demonstrate that testosterone inhibits such currents in a concentration-dependent manner. Importantly, this occurs over the physiological range of testosterone concentrations (IC50 34 nM), and is not mimicked by the metabolite 5alpha-androstan-17beta-ol-3-one (DHT), nor by progesterone or estradiol, even at high (10 microM) concentration. L-type Ca2+ channels in the vasculature are also important clinical targets for vasodilatory dihydropyridines. A single point mutation (T1007Y) almost completely abolishes nifedipine sensitivity in our recombinant expression system. Crucially, the same mutation renders the channels insensitive to testosterone. Our data strongly suggest, for the first time, the molecular requirements for testosterone binding to L-type Ca2+ channels, thereby supporting its beneficial role as an endogenous Ca2+ channel antagonist in the treatment of cardiovascular disease.

Within-host evolution of Staphylococcus aureus during asymptomatic carriage

Background Staphylococcus aureus is a major cause of healthcare associated mortality, but like many important bacterial pathogens, it is a common constituent of the normal human body flora. Around a third of healthy adults are carriers. Recent evidence suggests that evolution of S. aureus during nasal carriage may be associated with progression to invasive disease. However, a more detailed understanding of within-host evolution under natural conditions is required to appreciate the evolutionary and mechanistic reasons why commensal bacteria such as S. aureus cause disease. Therefore we examined in detail the evolutionary dynamics of normal, asymptomatic carriage. Sequencing a total of 131 genomes across 13 singly colonized hosts using the Illumina platform, we investigated diversity, selection, population dynamics and transmission during the short-term evolution of S. aureus. Principal Findings We characterized the processes by which the raw material for evolution is generated: micro-mutation (point mutation and small insertions/deletions), macro-mutation (large insertions/deletions) and the loss or acquisition of mobile elements (plasmids and bacteriophages). Through an analysis of synonymous, non-synonymous and intergenic mutations we discovered a fitness landscape dominated by purifying selection, with rare examples of adaptive change in genes encoding surface-anchored proteins and an enterotoxin. We found evidence for dramatic, hundred-fold fluctuations in the size of the within-host population over time, which we related to the cycle of colonization and clearance. Using a newly-developed population genetics approach to detect recent transmission among hosts, we revealed evidence for recent transmission between some of our subjects, including a husband and wife both carrying populations of methicillin-resistant S. aureus (MRSA). Significance This investigation begins to paint a picture of the within-host evolution of an important bacterial pathogen during its prevailing natural state, asymptomatic carriage. These results also have wider significance as a benchmark for future systematic studies of evolution during invasive S. aureus disease.

The search of a genetic basis for noise-induced hearing loss (NIHL)

Background and aim: Knowledge about the genetic factors responsible for noise-induced hearing loss (NIHL) is still limited. This study investigated whether genetic factors are associated or not to susceptibility to NIHL. Subjects and methods: The family history and genotypes were studied for candidate genes in 107 individuals with NIHL, 44 with other causes of hearing impairment and 104 controls. Mutations frequently found among deaf individuals were investigated (35delG, 167delT in GJB2, Delta(GJB6- D13S1830), Delta(GJB6- D13S1854) in GJB6 and A1555G in MT-RNR1 genes); allelic and genotypic frequencies were also determined at the SNP rs877098 in DFNB1, of deletions of GSTM1 and GSTT1 and sequence variants in both MTRNR1 and MTTS1 genes, as well as mitochondrial haplogroups. Results: When those with NIHL were compared with the control group, a significant increase was detected in the number of relatives affected by hearing impairment, of the genotype corresponding to the presence of both GSTM1 and GSTT1 enzymes and of cases with mitochondrial haplogroup L1. Conclusion: The findings suggest effects of familial history of hearing loss, of GSTT1 and GSTM1 enzymes and of mitochondrial haplogroup L1 on the risk of NIHL. This study also described novel sequence variants of MTRNR1 and MTTS1 genes.

UBE2A Deficiency Syndrome: Mild to Severe Intellectual Disability Accompanied by Seizures, Absent Speech, Urogenital, and Skin Anomalies in Male Patients

We describe three patients with a comparable deletion encompassing SLC25A43, SLC25A5, CXorf56, UBE2A, NKRF, and two non-coding RNA genes, U1 and LOC100303728. Moderate to severe intellectual disability (ID), psychomotor retardation, severely impaired/absent speech, seizures, and urogenital anomalies were present in all three patients. Facial dysmorphisms include ocular hypertelorism, synophrys, and a depressed nasal bridge. These clinical features overlap with those described in two patients from a family with a similar deletion at Xq24 that also includes UBE2A, and in several patients of Brazilian and Polish families with point mutations in UBE2A. Notably, all five patients with an Xq24 deletion have ventricular septal defects that are not present inpatients with a point mutation, which might be attributed to the deletion of SLC25A5. Taken together, the UBE2A deficiency syndrome in male patients with a mutation in or a deletion of UBE2A is characterized by ID, absent speech, seizures, urogenital anomalies, frequently including a small penis, and skin abnormalities, which include generalized hirsutism, low posterior hairline, myxedematous appearance, widely spaced nipples, and hair whorls. Facial dysmorphisms include a wide face, a depressed nasal bridge, a large mouth with downturned corners, thin vermilion, and a short, broad neck. (C) 2010 Wiley-Liss, Inc.

Role of the Mitochondrial Mutations, m. 827A > G and the Novel m. 7462C > T, in the Origin of Hearing Loss

Samples from 30 deaf probands exhibiting features suggestive of syndromic mitochondrial deafness or from families with maternal transmission of deafness were selected for investigation of mutations in the mitochondrial genes MT-RNR1 and MT-TS1. Patients with mutation m. 1555A>G had been previously excluded from this sample. In the MT-RNR1 gene, five probands presented the m. 827A>G sequence variant, of uncertain pathogenicity. This change was also detected in 66 subjects of an unaffected control sample of 306 Brazilian individuals from various ethnic backgrounds. Given its high frequency, we consider it unlikely to have a pathogenic role on hereditary deafness. As to the MT-TS1 gene, one proband presented the previously known pathogenic m. 7472insC mutation and three probands presented a novel variant, m. 7462C>T, which was absent from the same control sample of 306 individuals. Because of its absence in control samples and association with a family history of hearing impairment, we suggest it might be a novel pathogenic mutation.

Hematologically important mutations: X-linked chronic granulomatous disease (third update)

Chronic granulomatous disease (CGD) is an immunodeficiency disorder affecting about 1 in 250,000 individuals. The disease is caused by a lack of superoxide production by the leukocyte enzyme NADPH oxidase. Superoxide is used to kill phagocytosed micro-organisms in neutrophils, eosinophils, monocytes and macrophages. The leukocyte NADPH oxidase is composed of five subunits, of which the enzymatic component is gp91-phox, also called Nox2. This protein is encoded by the CYBB gene on the X chromosome. Mutations in this gene are found in about 70% of all CGD patients. This article lists all mutations identified in CYBB in the X-linked form of CGD. Moreover, apparently benign polymorphisms in CYBB are also given, which should facilitate the recognition of future disease-causing mutations. (C) 2010 Elsevier Inc. All rights reserved.

A quasispecies approach to the evolution of sexual replication in unicellular organisms

This study develops a simplified model describing the evolutionary dynamics of a population composed of obligate sexually and asexually reproducing, unicellular organisms. The model assumes that the organisms have diploid genomes consisting of two chromosomes, and that the sexual organisms replicate by first dividing into haploid intermediates, which then combine with other haploids, followed by the normal mitotic division of the resulting diploid into two new daughter cells. We assume that the fitness landscape of the diploids is analogous to the single-fitness-peak approach often used in single-chromosome studies. That is, we assume a master chromosome that becomes defective with just one point mutation. The diploid fitness then depends on whether the genome has zero, one, or two copies of the master chromosome. We also assume that only pairs of haploids with a master chromosome are capable of combining so as to produce sexual diploid cells, and that this process is described by second-order kinetics. We find that, in a range of intermediate values of the replication fidelity, sexually reproducing cells can outcompete asexual ones, provided the initial abundance of sexual cells is above some threshold value. The range of values where sexual reproduction outcompetes asexual reproduction increases with decreasing replication rate and increasing population density. We critically evaluate a common approach, based on a group selection perspective, used to study the competition between populations and show its flaws in addressing the evolution of sex problem.