Exploring the genetic basis of stroke. Spanish stroke genetics consortium.

This article provides an overview of stroke genetics studies ranging from the candidate gene approach to more recent studies by the genome wide association. It highlights the complexity of stroke owing to its different aetiopathogenic mechanisms, the difficulties in studying its genetic component, and the solutions provided to date. The study emphasises the importance of cooperation between the different centres, whether this takes places occasionally or through the creation of lasting consortiums. This strategy is currently essential to the completion of high-quality scientific studies that allow researchers to gain a better knowledge of the genetic component of stroke as it relates to aetiology, treatment, and prevention.

A study on superovulation using FSH and eCG in Awassi ewes.

The present study was conducted to evaluate superovulatory treatments in Awassi ewes by eCG and FSH. High number of unovulated follicles (P < 0.05) was observed in ewes treated with eCG in non-breeding season. It could be concluded that using FSH to induce superovulation in Awassi ewes is better than eCG.

Comprehensive survey of radionuclides in contemporary smokeless tobacco products.

There is considerable interest in the chemical composition of smokeless tobacco products (STPs), owing to health concerns associated with their use. Previous studies have documented levels of 210Po, 210Pb and uranium in STP samples. Here, the levels of 13 α-particle and 15 ß-radiation emitting radionuclides have been measured in a broad and representative range of contemporary STPs commercially available in the United States and Sweden. For each radionuclide, the level of radioactivity and calculated mass per gram of STP are reported. The results indicate that, among 34 Swedish snus and 44 US STPs, a more complex radionuclide content exists than previously reported for these products. Of the 28 radionuclides examined, 13 were detected and quantified in one or more STPs. The most frequently identified radionuclides in these STPs were 40K, 14C, 210Po and 226Ra. Over half the STPs also contained 228Th, and an additional 8 radionuclides were identified in a small number of STPs. The presence of 14C, 3H and 230Th are reported in tobacco for the first time. The activity of ß-emitters was much greater than those of α-emitters, and the ß-emitter 40K was present in the STPs with both the greatest radioactivity and mass concentrations. Since the three radionuclides included in the FDA's HPHC list were either not detected (235U), identified in only three of 78 samples (238U), and/or had activity levels over fifty times lower than that of 40K (210Po, 238U), there may be a rationale for reconsidering the radionuclides currently included in the FDA HPHC list, particularly with respect to 40K. Using a model of the physical and biological compartments which must be considered to estimate the exposure of STP users to radionuclides, we conclude that exposure from α-emitters may be minimal to STP users, but 40K in particular may expose the oral cavities of STP users to ß-radiation. Although a more comprehensive picture of the radioisotope content of STPs has emerged from this study, epidemiological evidence suggests that the levels of radionuclides measured in this study appear unlikely to present significant risks to STP users.

Epigallocatechin-3-gallate treatment reduces thermal hyperalgesia after spinal cord injury by down-regulating RhoA expression in mice.

BACKGROUND: (-)-Epigallocatechin-3-gallate (EGCG) is the major polyphenolic constituent found in green tea. It has been reported that may be a natural agent for reducing thermal and mechanical pain after nervous system injuries. However, the molecular pathways implicated in these beneficial effects have not been completely elucidated. This study aimed to assess the EGCG treatment effects on thermal hyperalgesia, spinal cord gliosis and modulation of Ras homologue gene family member A (RhoA), fatty acid synthase (FASN) and tumour necrosis factor alpha (TNF-α) expression after spinal cord contusion in mice. METHODS: Animals were subjected to a spinal cord contusion. Thirty minutes after contusion and daily during the first week post-surgery, animals were treated with EGCG or dimethyl sulfoxide-saline (DMSO-saline). At 7 and 14 days post-operation, motor recovery was evaluated using the Basso Mouse Scale, and nociceptive response was evaluated using the Hargreaves test. Furthermore, at 14 days, the expression of RhoA, FASN and TNF-α proteins was quantified in the lesion site of spinal cord by Western blot technique. Finally, spinal cord samples were processed by immunohistochemical techniques for observing astrocytes, microglia and afferent nerve fibres. RESULTS: At short time, EGCG treatment reduced significantly thermal hyperalgesia but had no effect on locomotor recovery in spinal cord injured mice. Furthermore, EGCG treatment down-regulated the RhoA, FASN and TNF-α proteins expression, and decreased astro- and microglia reactivity in spinal cord. CONCLUSION: These findings suggest that at short time EGCG treatment reduces thermal hyperalgesia and gliosis via FASN and RhoA pathway, causing a decrease in cytokines in spinal cord.

Presence in frog urinary bladder of proteins immunologically related to the aquaporin-CHIP.

Aquaporin-CHIP, a 28 kDa channel forming protein already referred to as CHIP28, has been identified as the water channel in red blood cells as well as in mammalian renal tubule cells. Another member of the aquaporin family, WCH-CD, has been found in the apical membrane of collecting duct principal cells and may represent the ADH-sensitive water channel. The present study investigates the possible presence of CHIP28-like proteins in amphibian urinary bladder, where the presence of water channels has been postulated. For this purpose, we raised polyclonal antibodies against human erythrocyte CHIP28. Immune serum precipitated a protein of about 30 kDa from the whole homogenate of urinary epithelial cells. By Western blotting, in addition to the reaction with the 30 kDa component, the immune serum reacted with higher molecular weight components from the bladder homogenate. The 30 kDa band was detected by Western blot only in bladders having a high water permeability. Moreover, a 30 kDa protein was also recognized in frog red blood cell membranes by the anti-CHIP28 antibodies. In line with the immunoblotting studies, in immunohistofluorescence anti-CHIP28 antibodies stained frog red blood cells and urinary bladder epithelial cells. However, in whole tissue water permeability studies apical treatment with the anti-CHIP28 antibodies had no effect on either the hydrosmotic response to ADH or on the basal net water flow of the bladder. All together, these results indicate the presence in the frog red blood cells and urinary epithelium of proteins sharing immunological analogies with aquaporin-CHIP.

The role of aquaporin-4 in the blood-brain barrier development and integrity: studies in animal and cell culture models.

Aquaporin-4 (AQP4) is the major water channel expressed in brain perivascular astrocyte processes. Although the role of AQP4 in brain edema has been extensively investigated, little information exists regarding its functional role at the blood-brain barrier (BBB). The purpose of this work is to integrate previous and recent data regarding AQP4 expression during BBB formation and depending on BBB integrity, using several experimental models. Results from studies on the chick optic tectum, a well-established model of BBB development, and the effect of lipopolysaccharide on the BBB integrity and on perivascular AQP4 expression have been analyzed and discussed. Moreover, data on the BBB structure and AQP4 expression in murine models of Duchenne muscular dystrophy are reviewed. In particular, published results obtained from mdx(3cv) mice have been analyzed together with new data obtained from mdx mice in which all the dystrophin isoforms including DP71 are strongly reduced. Finally, the role of the endothelial component on AQP4 cellular expression and distribution has been investigated using rat primary astrocytes and brain capillary endothelial cell co-cultures as an in vitro model of BBB.

Role of EEG monitoring and cross-clamping duration in carotid endarterectomy.

The usefulness of electroencephalographic (EEG) monitoring as well as the significance of the period of cross clamping in carotid endarterectomy have not been completely defined. In particular, the clinical importance of major EEG changes has not been fully investigated and some recent studies seem to indicate that the method has little value. As to the duration of cross clamping, there is strong evidence that occlusion times of about 15 minutes are tolerated under general anesthesia, but no information is available regarding longer periods of occlusion. The authors describe a consecutive series of 141 carotid endarterectomies in which the patients with EEG changes were shunted only when occlusion was anticipated to last longer than 30 minutes. Early major EEG changes (during the first 4 minutes) occurred in 14% of the cases. In the absence of EEG changes, long occlusion periods of 40 to 50 minutes were well tolerated. In contrast, the 20 patients with major persistent EEG changes did not tolerate protracted occlusion and three of them had immediate postoperative neurological complications. It seems that, in these circumstances, the incidence of neurological deficit is a function of the duration of cross clamping: these three patients had undergone occlusion for 15 to 30 minutes. Their deficits partially resolved. On the basis of these results it is concluded that: EEG recording is a reliable monitoring system in carotid artery cross clamping. No major strokes due to temporary carotid artery occurred in the series. The clinical significance of major persistent EEG changes is not negligible. Cross clamping for longer than 15 minutes in the presence of significant EEG alterations is potentially dangerous.

[Standard parenteral nutrition preparations in complex clinical situations]. / Preparados estándar de nutrición parenteral en situaciones clínicas complejas.

GOAL: Binary and ternary parenteral nutrition preparations may be of limited use in certain cases. The goal of this study is to establish difficult nutritional situations to handle and analyze the type of formula used in these situations. MATERIAL AND METHODS: The study included patients treated with parenteral nutrition over 9 months. Three clinically complex situations were defined: long duration, lasting more than 25 days; kidney failure, uraemia > 20 mmol/L or serum creatinine > 200 micromol/L; and liver failure, total bilirubin > 30 mmol/L or ALT > 2 microkat/L and alkaline phosphatase > 3 microkat/L or GGT > 3 microkat/L. Mortality and hypoalbuminaemia (< 35 g/L) were studied and compared by means of a chi squared test (p < 0.05) against the rest of the patients. The use of individualized formulas was studied using a multiple logarithmic regression model, the dependent variable being the administration or not of an individualized formula and the independent variables being the 3 groups of patients in clinical situations defined as complex. The Odds Ratio (OR) was studied as the measure of risk. RESULTS: A total of 511 patients receiving 8,015 feeds with parenteral nutrition were studied. Of these, 283 were included in one or more of the 3 complex clinical situations. All three groups presented higher levels of mortality and hypoalbuminaemia with statistically significant differences when compared to the group in a non-complex clinical situation. The use of individualized formulas was greater in the three groups defined, with statistically significant differences resulting: OR=6.7 (CI 95%; 3.78-11.91) with long duration; OR=3.66 (CI 95%; 2.68-5.68) in kidney failure; and OR=1.5 (CI 95%; 1.01-2.35) in liver failure. CONCLUSIONS: Patients in complex clinical situations present greater visceral malnutrition, a worse clinical evolution and, at our hospital, their nutritional treatment by parenteral means is based on a greater use of individualized formulas.

A model to control intermittent aeration phases.

Dealing with intermittent aeration as a useful practice to improve nitrogen removal efficiency of activated sludge plants, the possibility to plan optimal temporisation during daytime was investigated. A mathematical model (NIDEN) that allows us to manage different situations, with respect to influent load, environmental conditions and operating strategy, was then developed. The model represents a useful tool especially to plan the aeration cycles in small and medium sized plants, where high costs of automatic control through on-line instrumentation might not be justified. Once the input variables have been defined and the set-point values for tank nutrient concentration have been fixed, NIDEN gives an optimal phase temporisation, to obtain either the maximum energy saving or the best total nitrogen removal.

Surgery of thyroid cancer: twelve years' personal experience.

Diagnosis and treatment of thyroid carcinoma require a multidisciplinary approach. The close and long-standing collaboration between the Otorhinolaryngology, Pathological Anatomy and Nuclear Medicine Departments of Legnano Hospital has led to a precise diagnostic and therapeutic protocol in thyroid patients. In the 1990-2002 period, 131 patients underwent total thyroidectomy after diagnosis of thyroid cancer at the Otorhinolaryngology--Head and Neck Surgery Department. Patients submitted to lobectomy for differentiated thyroid cancer were excluded from the present study. The patient population is composed of 96 females (73%) and 36 males (27%) aged between 22 and 85 years. Of the 131 patients, 115 (87%) presented papillary carcinoma, 13 (10%) follicular carcinoma, 2 (2%) medullary carcinoma and one (1%) undifferentiated carcinoma. Two patients (2%) suffered from a preoperative monolateral recurrent nerve palsy. Total thyroidectomy was performed in all 131 patients. Selective neck dissection was performed only in patients with positive lymph nodes for papillary (37/115, 32%) and follicular carcinoma histotype (2/13, 15%) and, in both patients with medullary carcinoma (100%). Of the 131 patients, 15 (11%) did not undergo routine follow-up and were, therefore, excluded from the study, the remainder completed a mean follow-up of 47 months. During follow-up, the incidence of the two most frequent complications of thyroid surgery were evaluated: recurrent nerve paralysis and permanent hypoparathyroidism (exceeding the postoperative 6 months). Results of treatment have been evaluated considering the incidence of local and/or distant recurrences and patient survival rate. As far as concerns papillary and follicular histotype, we have considered as healed (absence of signs suggesting loco regional and distant recurrence) only those patients presenting both negligible levels of plasma thyroglobulin and a negative total-body 131I scintigraphy. Briefly, in 3 cases (3%), all papillary carcinomas, local recurrence occurred; 9 (8%), all with papillary carcinoma, developed lateral neck recurrence; 6 (5%), 5 with papillary carcinoma and one with follicular carcinoma, developed distant metastases, of which 3 pulmonary, 2 bone and 1 hepatic. Serum thyroglobulin values were considered during the last control visit in 95/113 patients (84%). Of these, 86 (91%) with negligible thyroglobulin levels and negative 131I scintigraphy, were considered healed. All 113 patients with differentiated thyroid carcinoma were alive at the last control visit. Both patients with medullary carcinoma are alive with no sign of illness at the last follow-up control. The patient presenting undifferentiated carcinoma died 2 months after surgery. In conclusion, at the last follow-up control, 1 (1%) patient has died, 5 patients (4%) are alive with disease (2 of whom suffered from multiple recurrences) and the remaining 110 (95%) patients are alive without evidence of disease. As far as concerns complications of surgery, iatrogenic recurrent palsy and permanent hypoparathyroidism are present in 2 (2%) and 10 patients (8%), respectively.

Uranium and thorium sequential separation from norm samples by using a SIA system.

This study presents a sequential radiochemical separation method for uranium and thorium isotopes using a novel Sequential Injection Analysis (SIA) system with an extraction chromatographic resin (UTEVA). After the separation, uranium and thorium isotopes have been quantified by using alpha-particle spectrometry. The developed method has been tested by analyzing an intercomparison sample (phosphogypsum sample) from International Atomic Energy Agency (IAEA) with better recoveries for uranium and thorium than the obtained by using a classical method (93% for uranium using the new methodology and 82% with the classical method, and in the case of thorium the recoveries were 70% for the semi-automated method and 60% for the classical strategy). Afterwards, the method was successfully applied to different Naturally Occurring Radioactive Material (NORM) samples, in particular sludge samples taken from a drinking water treatment plant (DWTP) and also sediment samples taken from an area of influence of the dicalcium phosphate (DCP) factory located close to the Ebro river reservoir in Flix (Catalonia). The obtained results have also been compared with the obtained by the classical method and from that comparison it has been demonstrated that the presented strategy is a good alternative to existing methods offering some advantages as minimization of sample handling, reduction of solvents volume and also an important reduction of the time per analysis.

The histone acetyltransferase component TRRAP is targeted for destruction during the cell cycle.

Chromosomes are dynamic structures that must be reversibly condensed and unfolded to accommodate mitotic division and chromosome segregation. Histone modifications are involved in the striking chromatin reconfiguration taking place during mitosis. However, the mechanisms that regulate activity and function of histone-modifying factors as cells enter and exit mitosis are poorly understood. Here, we show that the anaphase-promoting complex or cyclosome (APC/C) is involved in the mitotic turnover of TRRAP (TRansformation/tRanscription domain-Associated Protein), a common component of histone acetyltransferase (HAT) complexes, and that the pre-mitotic degradation of TRRAP is mediated by the APC/C ubiquitin ligase activators Cdc20 and Cdh1. Ectopic expression of both Cdh1 and Cdc20 reduced the levels of coexpressed TRRAP protein and induced its ubiquitination. TRRAP overexpression or stabilization induces multiple mitotic defects, including lagging chromosomes, chromosome bridges and multipolar spindles. In addition, lack of sister chromatid cohesion and impaired chromosome condensation were found after TRRAP overexpression or stabilization. By using a truncated form of TRRAP, we show that mitotic delay is associated with a global histone H4 hyperacetylation induced by TRRAP overexpression. These results demonstrate that the chromatin modifier TRRAP is targeted for destruction in a cell cycle-dependent fashion. They also suggest that degradation of TRRAP by the APC/C is necessary for a proper condensation of chromatin and proper chromosome segregation. Chromatin compaction mediated by histone modifiers may represent a fundamental arm for APC/C orchestration of the mitotic machinery.

Determination of 9°Sr and ²¹°Pb in sludge samples using a LOV-MSFIA system and liquid scintillation counting.

There has been increasing interest recently in the capacity of water treatment plants to concentrate and eliminate radionuclides from raw water. As the normal operation in these plants generates high quantities of sludge, which can be considered a naturally occurring radioactive material, it is important to gather information about its radiological content. Therefore, in order to determine the activity values of two radioactive beta emitters with minimal sample manipulation, an automated lab-on-valve and multisyringe flow injection system has been developed to achieve the sequential preconcentration and separation of (90)Sr and (210)Pb using an extraction chromatographic resin (Sr-spec). Activities of both isotopes were measured by liquid scintillation counting (LSC). The developed method was validated by analyzing three samples from three different intercomparison exercises and good Z-score values (between 0.1 and 1) and trueness values (between 10% and 17%) were obtained. The method was then also satisfactorily applied to sludge samples taken from a Spanish drinking water treatment plant which treats water from the Ebro River. The proposed method offers advantages over existing methods as it allows the sequential separation of both isotopes, simply by changing the elution conditions and using a semi-automated approach. Therefore, the method is less time consuming and environmentally friendly.

Comparative study of different analytical methods for the determination of 238U, 234U, 235U, 230Th and 232Th in NORM samples (Southern Catalonia).

Uranium and thorium isotopes from sludge samples taken from an area of influence of a dicalcium phosphate factory, located close to the Ebro River reservoir in Flix, and from mineral raw materials (coal and phosphate) which were deposited in Tarragona Harbour (Catalonia) have been measured. To do that, several procedures to determine these isotopes have been applied and compared in terms of accuracy, precision and total analysis time. In particular, digestion procedures, such as one based on conventional acidic digestion or another based on the use of microwave digestion, have been tested. Moreover, different radiochemical separation procedures have been applied. In this regard, one method based on liquid-liquid extraction using TBP/Xylene and another based on the use of extraction chromatography using UTEVA resins have been compared. Firstly, the different proposed methods were tested by analyzing a phosphogypsum sediment and a moss-soil sample from two different intercomparison exercises. The microwave digestion - UTEVA method provided the best results (recoveries higher than 82% for uranium and thorium isotopes) and was then applied to the NORM samples. The obtained results showed that the higher uranium and thorium activities values were found in phosphate raw material and in the more superficial samples located in sludge sediments next to the dicalcium phosphate factory.

Distribution of naturally occurring radioactive materials in sediments from the Ebro river reservoir in Flix (Southern Catalonia, Spain).

Industrial waste containing radioactive isotopes (from U-decay series) was released into Ebro river basin due to the activity of a dicalcium phosphate (DCP) plant for a period of more than two decades. Gross alpha, gross beta, (40)K, (226)Ra and (210)Pb activities were determined in several sludge samples taken at different depths from different points in the area of influence of the DCP plant located in Flix. Samples were collected from two different zones: one in front of the DCP plant and the second in front of a wastewater treatment plant installed several years after the DCP plant. The data obtained verify the influence of industrial DCP production on radioactivity levels present in the area.

Higher order structure of aquaporin-4.

Unlike other mammalian AQPs, multiple tetramers of AQP4 associate in the plasma membrane to form peculiar structures called Orthogonal Arrays of Particles (OAPs), that are observable by freeze-fracture electron microscopy (FFEM). However, FFEM cannot give information about the composition of OAPs of different sizes, and due to its technical complexity is not easily applicable as a routine technique. Recently, we employed the 2D gel electrophoresis BN-SDS/PAGE that for the first time enabled the biochemical isolation of AQP4-OAPs from several tissues. We found that AQP4 protein is present in several higher-order complexes (membrane pools of supra-structures) which contain different ratios of M1/M23 isoforms corresponding to AQP4-OAPs of different size. In this paper, we illustrate in detail the potentiality of 2D BN/SDS-PAGE for analyzing AQP4 supra-structures, their relationship with the dystrophin glycoprotein complex and other membrane proteins, and their role as a specific target of Neuromyelitis Optica autoantibodies.

[Venous thromboembolism and hyperhomocysteinemia as first manifestation of pernicious anemia]. / Tromboembolia venosa e hiperhomocisteinemia como primera manifestación de una anemia perniciosa.

INTRODUCTION AND OBJECTIVE: Hyperhomocysteinemia is associated to thrombosis and atherosclerosis. Vitamin B12 is among its main causes and may be due to a pernicious anemia. This study aimed to know the prevalence of this disease in patients who have venous thromboembolism and hyperhomocysteinemia. PATIENTS AND METHODS: A total of 80 consecutive patients (55 men and 25 women; age: mean [standard deviation] 63 [15] years) with pulmonary embolism and/or venous thrombosis and elevated values of homocysteine (> 12 micromol/l) were studied. RESULTS: Pernicious anemia was diagnosed (positive Schilling test, presence of anti-intrinsic factor antibodies and/or anti-parietal cells and fundal atrophic gastritis) in 5 patients (6.25% with range of age: 42-73 years. Only one of them had macrocytic anemia and there were no alterations in any of them in the thrombophilia study. The patients were treated with vitamin B12, administering it orally (1 mg/day) in 4 of them. The homocysteine and vitamin B12 values were normalized in every case at 6 months. CONCLUSIONS: Although the prevalence of pernicious anemia is not elevated in patients with venous thromboembolism and hyperhomocysteinemia, its existence must be ruled out to avoid other thrombotic and neurological complications.

Continuous intraoperative electroencephalographic monitoring in carotid endarterectomy.

Continuous electroencephalographic (EEG) monitoring was done during 143 consecutive carotid endarterectomies in 130 patients to detect intolerance to cross-clamping. An intraluminal shunt was inserted when EEG changes appeared (in the first 100 procedures) or when EEG changes appeared and a long clamping time was supposed (in the last 43 cases). 121 patients had no EEG changes at clamping time and only 1 had a postoperative deficit due to embolization, which occurred before clamping and was revealed by a decrease of voltage on tracing. 15 patients showed early (within 4 min after clamping) changes and in 11 patients shunting led to the reversal of EEG anomalies in all cases but 2. The state of these 2 patients was worse after surgery, in one because of embolic problems, in the other (shunted 30 min after clamping) probably for hemodynamic reasons. 4 patients with early EEG changes, not shunted because of the short clamping time, had no neurological deficit. 7 patients, 2 of which were shunted, showed late (after 4 min) EEG changes. Only 1 nonshunted patient in this group awoke with a minor, transitory neurological complication. Moreover, EEG changes have been correlated with the preoperative clinical condition and with the presence, on angiography, of contralateral carotid lesions. Also, EEG findings have been correlated with the computed tomography data (in 71 cases) and with the values of interior carotid artery (ICA) back pressure (in 58 cases). No possibility to predict the tolerance to clamping appeared from these correlations. However, a relatively higher risk of intolerance to clamping in patients with contralateral ICA occlusion was noted.(ABSTRACT TRUNCATED AT 250 WORDS)