Okur-Chung neurodevelopmental syndrome-linked CK2α variants have reduced kinase activity.

The Okur-Chung neurodevelopmental syndrome, or OCNDS, is a newly discovered rare neurodevelopmental disorder. It is characterized by developmental delay, intellectual disability, behavioral problems (hyperactivity, repetitive movements and social interaction deficits), hypotonia, epilepsy and language/verbalization deficits. OCNDS is linked to de novo mutations in CSNK2A1, that lead to missense or deletion/truncating variants in the encoded protein, the protein kinase CK2α. Eighteen different missense CK2α mutations have been identified to date; however, no biochemical or cell biological studies have yet been performed to clarify the functional impact of such mutations. Here, we show that 15 different missense CK2α mutations lead to varying degrees of loss of kinase activity as recombinant purified proteins and when mutants are ectopically expressed in mammalian cells. We further detect changes in the phosphoproteome of three patient-derived fibroblast lines and show that the subcellular localization of CK2α is altered for some of the OCNDS-linked variants and in patient-derived fibroblasts. Our data argue that reduced kinase activity and abnormal localization of CK2α may underlie the OCNDS phenotype.

Stability studies in biological fluids during post-analysis custody. Opiate compounds derived from heroin consumption.

In Forensic Toxicology, the evidences have to be maintained under custody for, at least, one year. Depending on the conditions and duration of storage, drug concentrations might have changed considerably since the first analysis. The aim of this study is to evaluate in vitro stability of opiate compounds, derived from heroin consumption, 6-acetylmorphine (6-MAM), morphine (MOR) and codeine (COD), in blood and urine, during post-analysis custody. Parameters evaluated were: time of custody, temperature, addition of preservative (blood) and pH (urine). Blood and urine samples were spiked with the three analytes to give a final concentration of 1000 ng/mL. The prepared samples were divided into 2 groups and stored at two temperatures (4 °C and -20 °C). Each one of these groups was subsequently divided in other two groups: with and without preservative (1%NaF) for blood, and pH 4 and 8 in the case of urine. 6-MAM, MOR and COD were analyzed by GCMS after SPE and derivatization with BSTFA. Analyses were performed in triplicate every two weeks for a year. In blood samples 6-MAM is the only compound that degrades. The best storage conditions were at -20 °C with NaF, with 6-MAM recoveries, after one year of custody, of 47.1 ± 1.5%; while in the other conditions 6-MAM disappeared after 215 days (at 4 °C with NaF), 45 days (at -20 °C without NaF) and 15 days (at 4 °C without preservative). COD does not degrade, with recoveries higher than 90%, in all of the conditions. They ranged from 89.7 ± 3.6% in samples maintained at -20 °C without NaF to 95.9 ± 2.0% in those maintained at 4 °C with NaF. MOR recoveries were lower than those of COD. They ranged from 66.9 ± 3.6%, in frozen samples added with NaF, to 78.6 ± 0.5% in refrigerated samples without preservative. In urine samples the three compounds were stable in all the studied conditions, with the exception of 6-MAM in samples at pH 8 and stored at 4 °C. In these conditions, 6-MAM disappeared after 135 days of custody; while recoveries in the other conditions ranged from 93.7 ± 6.4%, at 4 °C and pH 4, to 85.1 ± 2.0% at -20 °C and pH 8. MOR and COD recoveries were similar in the four conditions. In the case of MOR, they ranged from 82.1 ± 1.2% at 4 °C and pH 4 to 89.5 ± 6.0% at -20 °C and pH 8. As far as COD is concerned, recoveries ranged from 111.6 ± 5.8% at 4 °C and pH 8 to 102.6 ± 1.2% at 4 °C and pH 4. In conclusion, the study showed that the most labile opiate compound is 6-MAM. Its stability mainly depends on urine pH or the addition of preservative, in blood samples. The best storage conditions for samples from heroin consumers are in the freezer, at -20 °C. In addition, blood samples must be added with 1%NaF and urine samples must be buffered at pH 4.

The tooth for molecular analysis and identification : a forensic approach.

The aim of this study is to optimize laboratory preparation of teeth for DNA identification. By sectioning the tooth topographically into two different radicular portions, it was analyzed whether these portions of mineralized tissue differ in the quantity and quality of DNA they contain. 25 teeth were subject to different experimental conditions and total DNA was quantified for each individual tooth's radicular portion: apical and remaining root, according to a 2003 study by Gaytemenn and Sweet. We verified, with statistically significant figures, that the apical portion of the tooth is that which contains the greatest quantity of DNA. Different analytical procedures were studied for various polymorphic markers to evaluate the quality of the DNA. We concluded that the tooth is topographically distinct in both DNA quantity and quality. The tooth's apical portion is the preferential choice in sample preparation of dental mineralized tissue for molecular analysis and identification.

Association of the 3050G>C polymorphism in the cyclooxygenase 2 gene with systemic sarcoidosis.

BACKGROUND: We investigated the potential association between cyclooxygenase-2 (COX-2) gene polymorphisms and clinical manifestations of sarcoidosis. METHODS: This observational cross-sectional study involved seven hospitals in Spain. We diagnosed patients with sarcoidosis according to the International Criteria. The following variables were recorded: age, gender, initial diagnostic methods, serum angiotensin-converting enzyme (ACE) levels, pulmonary function tests, radiological stage, and clinical findings at diagnosis. Manifestations of sarcoidosis were classified as systemic vs. nonsystemic. Genotyping of four COX-2 polymorphisms (COX2.5909T>G, COX2.8473T>C, COX2.926G>C, and COX2.3050G>C) was undertaken on DNA extracted from peripheral blood lymphocytes using fluorescent hybridization probes and melting curves. RESULTS: A total of 131 sarcoid patients (63 males, mean age: 47 +/- 15 years) were studied. One hundred twenty-six of these patients had one or more positive biopsies. The results demonstrated that genotype distribution for the COX2.3050G>C polymorphism was significantly different between patients with systemic sarcoidosis and those with nonsystemic forms (p = 0.046). After adjustment for age, gender, and serum ACE levels, a significant association between the carriage of at least one C allele of the COX2.3050G>C polymorphism and systemic sarcoidosis was observed (odds ratio [OR]: 2.3; 95% confidence interval [CI]: 1.03-5.12, p = 0.031). Other polymorphisms were not associated with either clinical manifestations of the disease or serum ACE levels. CONCLUSIONS: Our results indicate for the first time that the C allele of the COX2.3050G>C polymorphism is associated with systemic sarcoidosis.

[Cardiac resynchronization through a persistent left superior vena cava]. / Resincronización cardíaca a través de una vena cava izquierda persistente.

Cardiac resynchronization therapy is effective in the treatment of patients with severe heart failure and intraventricular dysynchrony. However, we are sometimes faced with the unexpected presence of a persistent left superior vena cava. We report the case of a patient with dilated cardiomyopathy and left ventricular dysynchrony in which we implanted a resynchronization pacemaker exclusively through a persistent left superior vena cava that did not communicate with the right vena cava.

[Permanent pacing of the bundle of His after radiofrequency atrioventricular node ablation in patients with suprahisian conduction disturbances]. / Estimulación permanente del haz de His tras ablación mediante radiofrecuencia del nodo auriculoventricular en pacientes con trastorno de la conducción suprahisiano.

INTRODUCTION AND OBJECTIVES: The asynchronic contraction of the left ventricle due to left bundle branch block or right ventricular pacing is inferior from a hemodynamic point of view to the synchronic contraction through the conduction system. Several authors have reported some cases of pump failure and deterioration of mitral regurgitation after AV nodal ablation. Alternative sites of pacing such as the right ventricular outflow tract pacing have been proposed in order to avoid these complications. Direct His bundle pacing might be a new alternative for permanent pacing, however, it has not been extensively evaluated in humans yet. Our aim is to prove the feasibility of permanent His pacing in terms of stability, thresholds and pump function. POPULATION: patients without structural heart disease, selected for AV nodal ablation due to uncontrolled paroxysmal atrial fibrillation, or for pacemaker implantation due to supraHis conduction disturbance, with normal conduction system. An active fixation permanent lead was placed in His position using an steering guidewire and a diagnostic catheter as an anatomical reference. We also implanted a lead in the right atrial appendage and both were connected to a DDDR generator. Pacing thresholds and ecocardiographic ventricular function parameters were evaluated (ejection fraction, cavity size, mitral regurgitation). RESULTS: 12 patients met the inclusion criteria. Successful His pacing was achieved in 8 out of 12 cases (66%) with acceptable thresholds at implantation (1.24 +/- 0.13 volts at 0.5 ms) and during follow up at 3 months (1.31 +/- 0.20 volts at 0.5 ms). Neither a significant change in the ecocardiographic parameters not a deterioration in the clinical status caused by ablation or stimulation was evidenced. CONCLUSION: The His bundle may be the site of choice for long term pacing in patients with AV block and normal infraHis conduction system.

A study on ten short tandem repeat systems: African immigrant and Spanish population data.

This work presents the results obtained from a genetic-population study for the D1S1656 system in the population of Southwest Spain (Huelva, Cádiz and Sevilla), Spaniards of Caucasian origin from North Africa (Ceuta), as well as in the black Central West African and Moroccan immigrant populations in Spain. The results of a study of the autochtonous population of the Canary Islands (n=138), and immigrant Central West African populations in Spain (n=132), obtained for nine short tandem repeat (STR) loci (D3S1358, VWA, FGA, D8S1179, D21S11, D18S51, D5S818, D13S317, D7S820), as well as the amelogenin locus, all contained in Profiler Plus (Perkin-Elmer) PCR amplification kits, are also presented. Except for the FGA and VWA data on immigrant Central West African populations in Spain, no deviations from the Hardy-Weinberg equilibrium were detected.

A study among the population of Sevilla of death due to submersion.

Death due to submersion is of great interest from the medical-legal point of view, given the increase in nautical activity among children and adults alike over the past number of years. However, the lack of reliable statistical data concerning the impact of this specific form of death in our country must be emphasized. These are the circumstances that have led us to study the incidence of this form of death in a specific area. The population analyzed lived in the city of Sevilla during the period 1967-1993.

Group-specific component (GC) polymorphism in Cádiz (southern Spain).

The genetic polymorphism of group-specific component (GC) was analysed in a sample of 443 healthy unrelated subjects of both sexes resident in the province of Cádiz (Southern Spain). Isoelectric focusing was carried out in polyacrylamide gels followed by staining with coomassie blue R 250. The estimated gene frequencies were as follows: GC*1S = 0.6185; GC*1F = 0.1162; GC*2 = 0.2652.

Polymorphism of the plasminogen (PLG) system in Cádiz Province, southern Spain.

In this work, the authors report a plasminogen (PLG) system genetic-population study in a sample of 378 healthy subjects, of both sexes and unrelated, all resident in the province of Cádiz in Southern Spain. In this study, the PLG types were determined by isoelectric focusing in polyacrylamide gels (PAGIF), followed by staining with Coomassie blue. The genic frequencies obtained were the following: PLG A = 0.833 333 3; PLG B = 0.166 666 7.

Transferrin gene frequencies in Cádiz (southern Spain).

The genetic polymorphism of transferrin (Tf) was studied in a sample of 385 healthy unrelated subjects of both sexes resident in the province of Cádiz (southern Spain). Isoelectric focusing was carried out in polyacrylamide gels, followed by staining with Coomassie Blue R250. The gene frequencies obtained were as follows: Tf C1, 0.7922; Tf C2, 0.1883; Tf C3, 0.0195.

Bradykinin-evoked modulation of cytosolic Ca2+ concentrations in cultured renal epithelial (MDCK) cells.

We have investigated the effect of bradykinin on cytosolic Ca2+ concentrations of renal MDCK cells cultured as monolayers. Bradykinin rapidly (within 5 to 20 s) increased cytosolic Ca2+ concentrations, measured by using the fluorescent indicator quin-2, from the basal value of 103 nM to a maximal value of 578 nM at about 10(-8) M bradykinin. The increase of Ca2+ was transient, returning to baseline within 1.5 to 2 min. The transient response appeared to be due to cell desensitization rather than peptide degradation. Previously desensitized cells could be resensitized after a 10 min incubation in the absence of bradykinin. The removal of extracellular Ca2+ or the addition of verapamil did not have a major effect on the maximal bradykinin-evoked changes of Ca2+, suggesting that Ca2+ released from intracellular stores plays a pivotal role in this process. Bradykinin-evoked Ca2+ metabolism may play an important role as modulator of the cellular functions of MDCK cells.