Programa de formación de ayudantes de investigación: conocimiento y satisfacción de los estudiantes / Research assistant formation program: students' knowledge and satisfaction / Programa de formação de assistentes de pesquisa: conhecimento e satisfação dos estudantes

Introducción: La inserción de estudiantes de pregrado en actividades de investigación es relevante para fomentar la crítica y aplicación de resultados de investigación. La formación de ayudantes de investigación se plantea como una alternativa válida para que los alumnos se expongan, experimenten y participen activamente en el proceso de investigación. Objetivo: Identificar el nivel de conocimiento y grado de satisfacción de los estudiantes después de participar en un programa de formación de ayudantes de investigación. Método: Estudio de pre y post test en 20 estudiantes de pregrado de Enfermería. Se evaluó el conocimiento alcanzado y la satisfacción de los estudiantes con el programa de formación, el cual considera tres módulos de aprendizaje distribuidos en 180 horas cronológicas. Resultados: El conocimiento de los estudiantes incrementó significativamente con el programa, no se relacionó con el conocimiento que el alumno trae al inicio del mismo, así como tampoco con el semestre que cursa. El grado de satisfacción fue en el 100% de los alumnos muy satisfecho. Discusión: Este artículo reporta un programa de formación de ayudantes de investigación que es posible ser replicado en otras realidades, que incrementa el conocimiento, con un alto grado de satisfacción en los estudiantes. La duración del programa podría considerarse una debilidad. Conclusión: La participación de estudiantes de pregrado en un programa de formación de ayudantes de investigación, puede considerarse una estrategia para involucrar a alumnos en investigación, y que valoren su aporte en la disciplina de enfermería.

Introduction: An early integration of undergraduate students into activities of research can help them develop skills for a critical application of the corresponding findings. The formation of research assistants program is proposed as an experience for students to expose, experiment, and actively participate in diverse researching related activities. Objective: To identify the level of knowledge and degree of satisfaction of students after their participation in a research assistant formation program. Method: This is pre-post-test study involving 20 undergraduate nursing students. Their achieved knowledge and satisfaction in the three-module-180-hours program were assessed. Results: Unrelated to parallel experiences, the students' knowledge increased significantly after the program. All students stated having felt very satisfied. Discussion: This article informs on a program of formation of research assistants which is prone to be extended to other contexts, however its duration could be adjusted. Conclusion: The participation of undergraduate students in a research assistant formation program is a novel strategy to engage students into nursing research.

Introdução: A inserção de estudantes de graduação em atividades de pesquisa é relevante para promover a crítica e aplicação de resultados de pesquisa. A formação de assistentes de pesquisa contempla-se como una alternativa válida para os alunos se exporem, experimentarem e participarem ativamente no processo de pesquisa. Objetivo: Identificar o nível de conhecimento e grau de satisfação dos estudantes depois de participarem em um programa de formação de assistentes de pesquisa. Método: Estudo de pre e post teste de 20 estudantes de graduação de Enfermagem. Avaliou-se o conhecimento atingido e a satisfação dos estudantes com o programa de formação, o qual considera três módulos de aprendizagem distribuídos em 180 horas cronológicas. Resultados: O conhecimento dos estudantes aumentou significativamente com o programa, não se relaciona com o conhecimento que o aluno traz ao início do mesmo, assim como também não, com o semestre que cursa. O grau de satisfação foi do 100% dos alunos: muito satisfeito. Discussão: Este artigo informa um programa de formação de assistentes de pesquisa que é possível ser reproduzido em outras realidades, que aumenta o conhecimento, com um alto grau de satisfação nos estudantes. A duração do programa podia considerar-se uma debilidade. Conclusão: A participação de estudantes de graduação em um programa de formação de assistentes de pesquisa, pode considerar-se uma estratégia para envolver os alunos em pesquisa, e que valorizem seu aporte na disciplina de enfermagem.

Impact of cognitive impairment, depression, disease activity, and disease damage on quality of life in women with systemic lupus erythematosus.

OBJECTIVES: To define the relative role of cognitive impairment, depression, disease activity, and disease damage in the decreased health-related quality of life (HRQoL) frequently observed in systemic lupus erythematosus (SLE) patients. METHOD: We studied 101 Chilean female SLE patients and applied the 12-item Medical Outcomes Study (MOS) Short Form Health Survey version 2 (SF-12v2) to assess HRQoL and the Cambridge Neuropsychological Test Automated Battery (CANTAB) to assess cognitive function. Analysis of covariance (ANCOVA) models included demographic and disease-related factors and cognitive function tests of sustained attention, memory, and executive function. RESULTS: All measures of HRQoL were lower in the 101 female SLE patients compared to the women from the Chilean general population. HRQoL was associated with the following factors: (i) depression symptoms, which were detrimental to all components of the physical and mental HRQoL scores; (ii) executive dysfunction (spatial planning), which was associated with lower scores on role limitations due to physical health problems and emotional problems, and general health perceptions; (iii) higher activity and organ damage were deleterious to role physical, bodily pain, and physical summary scores; and (iv) higher damage also impacted physical function. Impairments in sustained attention and memory did not decrease the HRQoL. CONCLUSIONS: Our results highlight the relevance of executive dysfunction to poor physical and mental health components of HRQoL in SLE together with depression, while disease activity and disease damage are associated with lower HRQoL physical components. The need for cognitive function evaluation and rehabilitation in SLE is indicated.

Effect of foods and Mediterranean diet during pregnancy and first years of life on wheezing, rhinitis and dermatitis in preschoolers

BACKGROUND: There is a conflictive position if some foods and Mediterranean diet (MedDiet) consumed by the mother during pregnancy and by the child during the first years of life can be protective for current wheezing, rhinitis and dermatitis at preschool age. METHODS: Questionnaires of epidemiological factors and food intake by the mother during pregnancy and later by the child were filled in by parents in two surveys at two different time points (1.5 yrs and 4 yrs of life) in 1000 preschoolers. RESULTS: The prevalences of current wheezing, rhinitis and dermatitis were 18.8%, 10.4%, and 17.2%, respectively. After multiple logistic analysis children who were low fruit consumers (never/occasionally) and high fast-food consumers (≥3 times/week) had a higher risk for current wheezing; while intermediate consumption of meat (1 or 2 times/week) and low of pasta by mothers in pregnancy were protected. For current rhinitis, low fruit consumer children were at higher risk; while those consuming meat <3 times/week were protected. For current dermatitis, high fast food consumption by mothers in pregnancy; and low or high consumption of fruit, and high of potatoes in children were associated to higher prevalence. Children consuming fast food >1 times/week were protected for dermatitis. MedDiet adherence by mother and child did not remain a protective factor for any outcome. CONCLUSION: Low consumption of fruits and high of meat by the child, and high consumption of potatoes and pasta by the mother had a negative effect on wheezing, rhinitis or dermatitis; while fast food consumption was inconsistent

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Increased prevalence of Mycoplasma pneumoniae serological positivity in Chilean young children

BACKGROUND: Mycoplasma pneumoniae is a frequent cause of respiratory infections in school children and adolescents. Epidemiological suspicion is important, since there are no specific symptoms or signs to help in diagnosing infection caused by this agent. OBJECTIVE: To determine the variation in prevalence over the last 10 years of M. pneumoniae IgM seropositivity according to age, particularly in pre-schoolers. Method: The results of M. pneumoniae IgM serological testing between January 2004 and December 2013 were analysed. Variables such as gender and month and year of sample processing were studied according to age groups (<5, 5-18, 19-50, 51-70 and >70 years of age). RESULTS: Of a total of 20,020 serological samples, 31.9% proved positive for M. pneumoniae. All age groups showed increases in percentage seropositivity over the last 10 years, although the most significant increase corresponded to the 5-18 years group (from 15.8% to 54%), followed by children <5 years of age (from 8.6% to 30%). Seropositivity was significantly higher in women in all age groups, except in those over 50 years of age. CONCLUSIONS: Children under five years of age were the group with the second highest increase in seropositivity

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Increased prevalence of Mycoplasma pneumoniae serological positivity in Chilean young children.

BACKGROUND: Mycoplasma pneumoniae is a frequent cause of respiratory infections in school children and adolescents. Epidemiological suspicion is important, since there are no specific symptoms or signs to help in diagnosing infection caused by this agent. OBJECTIVE: To determine the variation in prevalence over the last 10 years of M. pneumoniae IgM seropositivity according to age, particularly in pre-schoolers. METHOD: The results of M. pneumoniae IgM serological testing between January 2004 and December 2013 were analysed. Variables such as gender and month and year of sample processing were studied according to age groups (<5, 5-18, 19-50, 51-70 and >70 years of age). RESULTS: Of a total of 20,020 serological samples, 31.9% proved positive for M. pneumoniae. All age groups showed increases in percentage seropositivity over the last 10 years, although the most significant increase corresponded to the 5-18 years group (from 15.8% to 54%), followed by children <5 years of age (from 8.6% to 30%). Seropositivity was significantly higher in women in all age groups, except in those over 50 years of age. CONCLUSION: Children under five years of age were the group with the second highest increase in seropositivity.

Effect of foods and Mediterranean diet during pregnancy and first years of life on wheezing, rhinitis and dermatitis in preschoolers.

BACKGROUND: There is a conflictive position if some foods and Mediterranean diet (MedDiet) consumed by the mother during pregnancy and by the child during the first years of life can be protective for current wheezing, rhinitis and dermatitis at preschool age. METHODS: Questionnaires of epidemiological factors and food intake by the mother during pregnancy and later by the child were filled in by parents in two surveys at two different time points (1.5 yrs and 4 yrs of life) in 1000 preschoolers. RESULTS: The prevalences of current wheezing, rhinitis and dermatitis were 18.8%, 10.4%, and 17.2%, respectively. After multiple logistic analysis children who were low fruit consumers (never/occasionally) and high fast-food consumers (≥3 times/week) had a higher risk for current wheezing; while intermediate consumption of meat (1 or 2 times/week) and low of pasta by mothers in pregnancy were protected. For current rhinitis, low fruit consumer children were at higher risk; while those consuming meat <3 times/week were protected. For current dermatitis, high fast food consumption by mothers in pregnancy; and low or high consumption of fruit, and high of potatoes in children were associated to higher prevalence. Children consuming fast food >1 times/week were protected for dermatitis. MedDiet adherence by mother and child did not remain a protective factor for any outcome. CONCLUSION: Low consumption of fruits and high of meat by the child, and high consumption of potatoes and pasta by the mother had a negative effect on wheezing, rhinitis or dermatitis; while fast food consumption was inconsistent.

Many respiratory viruses have temporal association with meningococcal disease

BACKGROUND: Previous ecological studies have shown a temporal and spatial association between influenza epidemics and meningococcal disease (MNG); however, none have examined more than two respiratory viruses. METHODS: Data were obtained in Chile between 2000 and 2005 on confirmed cases of MNG and all confirmed cases of respiratory viruses (influenza A and B; parainfluenza; adenovirus; and respiratory syncytial virus [RSV]). Both variables were divided by epidemiological weeks, age range, and regions. Models of transference functions were run for rates of MNG. RESULTS: In this period, 1022 reported cases of MNG and 34,737 cases of respiratory virus were identified (25,137 RSV; 4300 parainfluenza; 2527 influenza-A; 356 influenza-B; and 2417 adenovirus). RSV was the major independent virus temporally associated to MNG (it appears one week before MNG), followed by parainfluenza, influenza-B, adenovirus, and influenza-A. CONCLUSIONS: The rate of MNG in Chile is temporally associated to all of the respiratory viruses studied, but with variability according age range, and regions

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Methacholine challenge test by wheezing and oxygen saturation in preschool children with asthma

Methacholine challenge test (MCT) performed with spirometry is a commonly used test to evaluate bronchial hyperreactivity (BHR) in children. However, preschoolers do not usually collaborate. OBJECTIVES: To assess the usefulness of MCT through clinical evaluation (wheezing auscultation and decreased pulse arterial oxygen saturation [SpO2]) in recurrent wheezing preschoolers with asthma, in comparison to healthy controls. METHODS: We performed the MCT (modified Cockroft method) on healthy and on asthmatic preschoolers. The end point was determined by the presence of wheezing in the chest and/or tracheal auscultation (PCw) and/or a decrease in SpO2 of ≥5 from the baseline value (PCSpO2). Maximal methacholine concentration was 8 mg/ml. RESULTS: The study population comprised 65 children: 32 healthy and 33 asthmatic children. There were no differences in demographic characteristics between the groups. The median methacholine doses for PCw and for PCSpO2 were significantly lower among asthmatic than healthy children: 0.5 mg/ml (0.25-0.5 mg/ml) vs. 2 mg/ml (1-4 mg/ml), respectively, p < 0.001; and 0.25 mg/ml (0.25-0.5 mg/ml) and 2 mg/ml (0.5-4 mg/ml), respectively, p < 0.001. The best cut-off point of PCw was observed at a methacholine concentration of 0.5 mg/ml (AUC = 0.72 [95% CI = 0.66-0.77]), its sensitivity was 91%, specificity 43%, PPV 16% and NPV 98%. For PCSpO2 the best cut-off point was a methacholine concentration of 1 mg/ml (AUC = 0.85 [95% CI 0.81-0.89]), with sensitivity of 80%, specificity 74%, PPV 49%, and NPV 92%. There were no adverse reactions. CONCLUSION: MCT using clinical parameters such as wheezing auscultation and SpO2 measurement could be a useful and safe test to confirm BHR among preschoolers

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Methacholine challenge test by wheezing and oxygen saturation in preschool children with asthma.

UNLABELLED: Methacholine challenge test (MCT) performed with spirometry is a commonly used test to evaluate bronchial hyperreactivity (BHR) in children. However, preschoolers do not usually collaborate. OBJECTIVES: To assess the usefulness of MCT through clinical evaluation (wheezing auscultation and decreased pulse arterial oxygen saturation [SpO2]) in recurrent wheezing preschoolers with asthma, in comparison to healthy controls. METHODS: We performed the MCT (modified Cockroft method) on healthy and on asthmatic preschoolers. The end point was determined by the presence of wheezing in the chest and/or tracheal auscultation (PCw) and/or a decrease in SpO2 of ≥5 from the baseline value (PCSpO2). Maximal methacholine concentration was 8 mg/ml. RESULTS: The study population comprised 65 children: 32 healthy and 33 asthmatic children. There were no differences in demographic characteristics between the groups. The median methacholine doses for PCw and for PCSpO2 were significantly lower among asthmatic than healthy children: 0.5 mg/ml (0.25-0.5 mg/ml) vs. 2 mg/ml (1-4 mg/ml), respectively, p<0.001; and 0.25 mg/ml (0.25-0.5 mg/ml) and 2 mg/ml (0.5-4 mg/ml), respectively, p<0.001. The best cut-off point of PCw was observed at a methacholine concentration of 0.5 mg/ml (AUC=0.72 [95% CI=0.66-0.77]), its sensitivity was 91%, specificity 43%, PPV 16% and NPV 98%. For PCSpO2 the best cut-off point was a methacholine concentration of 1 mg/ml (AUC=0.85 [95% CI 0.81-0.89]), with sensitivity of 80%, specificity 74%, PPV 49%, and NPV 92%. There were no adverse reactions. CONCLUSION: MCT using clinical parameters such as wheezing auscultation and SpO2 measurement could be a useful and safe test to confirm BHR among preschoolers.

Anti-N-methyl-D-aspartate receptor and anti-ribosomal-P autoantibodies contribute to cognitive dysfunction in systemic lupus erythematosus.

OBJECTIVE: Autoantibodies against N-methyl-D-aspartate receptor (anti-NMDAR) and ribosomal-P (anti-P) antigens are potential pathogenic factors in the frequently observed diffuse brain dysfunctions in patients with systemic lupus erythematosus (SLE). Although studies have been conducted in this area, the role of anti-NMDAR antibodies in SLE cognitive dysfunction remains elusive. Moreover, the specific contribution of anti-P antibodies has not been reported yet. The present study attempts to clarify the contribution of anti-NMDAR and anti-P antibodies to cognitive dysfunction in SLE. METHODS: The Cambridge Neuropsychological Test Automated Battery (CANTAB) was used to assess a wide range of cognitive function areas in 133 Chilean women with SLE. ANCOVA models included autoantibodies, patient and disease features. RESULTS: Cognitive deficit was found in 20%. Higher SLEDAI-2K scores were associated with impairment in spatial memory and learning abilities, whereas both anti-NMDAR and anti-P antibodies contributed to deficits in attention and spatial planning abilities, which reflect fronto-parietal cortex dysfunctions. CONCLUSIONS: These results reveal an association of active disease together with specific circulating autoantibodies, such as anti-NMDAR and anti-P, with cognitive dysfunction in SLE patients.

Many respiratory viruses have temporal association with meningococcal disease.

BACKGROUND: Previous ecological studies have shown a temporal and spatial association between influenza epidemics and meningococcal disease (MNG); however, none have examined more than two respiratory viruses. METHODS: Data were obtained in Chile between 2000 and 2005 on confirmed cases of MNG and all confirmed cases of respiratory viruses (influenza A and B; parainfluenza; adenovirus; and respiratory syncytial virus [RSV]). Both variables were divided by epidemiological weeks, age range, and regions. Models of transference functions were run for rates of MNG. RESULTS: In this period, 1022 reported cases of MNG and 34,737 cases of respiratory virus were identified (25,137 RSV; 4300 parainfluenza; 2527 influenza-A; 356 influenza-B; and 2417 adenovirus). RSV was the major independent virus temporally associated to MNG (it appears one week before MNG), followed by parainfluenza, influenza-B, adenovirus, and influenza-A. CONCLUSIONS: The rate of MNG in Chile is temporally associated to all of the respiratory viruses studied, but with variability according age range, and regions.

Systemic lupus erythematosus impairs memory cognitive tests not affected by depression.

OBJECTIVE: Our aim was to assess the contribution of depression to cognitive impairment in patients with systemic lupus erythematosus (SLE). METHODS: Clinical features, education, age, and Hospital Anxiety and Depression Scale (HADS) were evaluated in 82 patients with SLE and 22 healthy controls, all Chilean women. The Cambridge Neuropsychological Test Automated Battery (CANTAB eclipseTM) assessing attention, spatial memory, and learning and executive function domains was applied. Cognitive deficit definition: a cut-off for definite impairment was defined as a score below -2 standard deviations in at least one outcome measure in two or more domains. ANCOVA with stepwise selection evaluated influences of health status (SLE or control), age, education, and HADS depression and anxiety scores on cognitive outcomes. To avoid overfitting, a shrinkage method was performed. Also, adjusted p-values for multiple comparisons were obtained. RESULTS: Cognitive deficit affected 16 (20%) patients, and no controls (p=0.039). Median HADS depression score in SLE patients was 6 (range 0-19) and in controls was 0 (0-19), p<0.001). ANCOVA and shrinkage models showed that worse cognitive performance in sustained attention and spatial working memory tests was explained by the presence of SLE but not depression, whereas depression only affected a measure of executive function (I/ED Stages completed). CONCLUSION: Depression has a limited role in cognitive impairment in SLE. Impairments in sustained attention and spatial working memory are distinctly influenced by yet-unknown disease-intrinsic factors.

[A prolonged capillary refill is predictor of low central venous oxygen saturation]. / El tiempo de llene capilar prolongado es predictor de una saturación venosa central de oxígeno disminuida.

INTRODUCTION: Educational programs in pediatric life support endorse a capillary refill time>2 s as an indicator of shock. In the emergency room, a barrier to the implementation of an early goal directed therapy, aiming at central venous oxygen saturation (ScvO2)≥70% is the insertion of central venous catheter (CVC). OBJECTIVE: To establish the predictive value of capillary refill time>2 s to detect ScvO2<70% in children admitted to Intensive Care Units. PATIENTS AND METHOD: Prospective study. We included 48 children admitted in the first 24 hours in ICU with superior vena cava CVC. Simultaneously, we measured ScvO2 and capillary refill time in the heel of upper extremity or toe. RESULTS: There were 75 paired measurements ScvO2 (75,9±8,4%) and capillary refill capillary (1,9±1,0 s). We found an inverse correlation between capillary refill time and ScvO2 (r-0,58). The ROC curve analysis revealed an excellent ability for the capillary fill time>2 s to predict ScvO2<70% (AUC 0,94) (95% CI 0,87-0,98). CONCLUSIONS: A prolonged capillary refill time>2 s, is a predictor of ScvO2<70% in children admitted to ICU, which supports the current recommendations. This finding may be relevant in emergency units where the use of CVC is limited and ScvO2 is not available.

Reproducibility and performance of one or two samples of salivary cortisol in the diagnosis of Cushing's syndrome using an automated immunoassay system.

The purpose of this article is to evaluate the variability and reproducibility of late night salivary cortisol (LNSC) using electrochemiluminescence immunoassay (ECLIA) and compare the accuracy of one or two samples in diagnosis of Cushing's syndrome (CS). We prospectively included 64 healthy volunteers (HV), 35 patients with clinically suspected CS (S), and 26 patients with confirmed CS. Nine patients in the CS group had 24-h urinary free cortisol (UFC) less than two times the upper limit of normal (mild CS). UFC and two consecutive LNSC (LNSC1, LNSC2) were collected at home. All patients in the S group had normal UFC and low-dose dexamethasone suppression test. No differences were found between the HV and S groups in UFC, LNSC1, and LNSC2. Intra-individual variability between the two samples of LNSC was 22% in HV (1.6-91%), 32% in the S group (1.6-144%), and 51% (1.6-156%) in the CS group. Variability was higher in CS patients than those in the HV (P < 0.001) and S groups (P = 0.05). The AUC of LNSC1 was 0.945 (IC 95% 0.880-1.004); when considering the highest LNSC, the AUC was 0.980 (IC 95% 0.954-1.007) (P < 0.01). We found 23% of discordant LNSC in the S group and 11% in the CS group. Three patients with CS had only one elevated LNSC, all of them with mild CS. Our results suggest that LNSC is variable, and reproducibility is affected in both CS and S patients. We found significant improvements in the diagnostic accuracy of the LNSC measurement by obtaining two samples.

Survivin is a key factor in the differential susceptibility of gastric endothelial and epithelial cells to alcohol-induced injury.

We previously demonstrated that the anti-apoptosis protein, survivin, plays a protective role against alcohol-induced gastric injury. Since the endothelium is a primary target of alcohol-induced gastric damage, we investigated whether survivin expression is a key factor in the greater susceptibility of gastric endothelial vs. epithelial cells to alcohol-induced injury. Here, we demonstrate that rat gastric epithelial cells (RGM1 cells, an epithelial cell line derived from normal rat gastric mucosa) expressed 7.5-fold greater survivin protein levels vs. rat gastric endothelial cells. Survivin expression correlated with resistance of gastric epithelial vs. endothelial cells to both alcohol-induced cell damage and alcohol-induced apoptosis. Suppression of survivin protein expression levels using siRNA rendered the gastric epithelial cells as susceptible to both alcohol-induced cell damage and apoptosis as the gastric endothelial cells. Conversely, forced overexpression of survivin by transient transfection rendered gastric endothelial cells as resistant to both alcohol-induced cell damage and apoptosis as mock-transfected gastric epithelial cells. Moreover, overexpression of a threonine-34 to glutamate phosphorylation mimic mutant survivin construct rendered gastric endothelial cells significantly more resistant to alcohol-induced damage and apoptosis vs. mock-transfected gastric epithelial cells. These findings indicate that disparate survivin expression levels can explain the discrepancy between gastric epithelial and endothelial cell susceptibility to alcohol-induced injury; and, that a negative charge at amino acid residue 34 on survivin, such as that which naturally occurs by phosphorylation of threonine-34, enhances its property in conferring gastric mucosal protection.

Novel properties of a mouse gamma-aminobutyric acid transporter (GAT4).

We expressed the mouse gamma-aminobutyric acid (GABA) transporter GAT4 (homologous to rat/ human GAT-3) in Xenopus laevis oocytes and examined its functional and pharmacological properties by using electrophysiological and tracer uptake methods. In the coupled mode of transport (Na+/ Cl-/GABA cotransport), there was tight coupling between charge flux and GABA flux across the plasma membrane (2 charges/GABA). Transport was highly temperature-dependent with a temperature coefficient (Q10) of 4.3. The GAT4 turnover rate (1.5 s(-l); -50 mV, 21 degrees C) and temperature dependence suggest physiological turnover rates of 15-20 s(-1). No uncoupled current was observed in the presence of Na+. In the absence of external Na+, GAT4 exhibited two distinct uncoupled currents. (i) A Cl- leak current (ICl(leak)) was observed when Na+ was replaced with choline or tetraethylammonium. The reversal potential of (ICl(leak)) followed the Cl- Nernst potential. (ii) A Li+ leak current (ILi(leak)) was observed when Na+ was replaced with Li+. Both leak currents were inhibited by Na+, and both were temperature-independent (Q10 approximately 1). The two leak modes appeared not to coexist, as Li+ inhibited (ICl(leak)). The results suggest the existence of cation- and anion-selective channel-like pathways in GAT4. Flufenamic acid inhibited GAT4 Na+/Cl-/GABA cotransport, ILi(leak), and ICl(leak), (Ki approximately 30 microM), and the voltage-induced presteady-state charge movements (Ki approximately 440 microM). Flufenamic acid exhibited little or no selectivity for GAT1, GAT2, or GAT3. Sodium and GABA concentration jicroumps revealed that slow Na+ binding to the transporter is followed by rapid GABA-induced translocation of the ligands across the plasma membrane. Thus, Na+ binding and associated conformational changes constitute the rate-limiting steps in the transport cycle.

Biochemical characterization of recombinant and circulating human Spalpha.

Human Spalpha is a soluble protein expressed by macrophages present in lymphoid tissues (spleen, lymph node, thymus, and bone marrow), for which little functional and structural information is available. It belongs to the group B of the scavenger receptor cysteine-rich superfamily (SRCR-SF) that includes the lymphocyte surface receptors CD5 and CD6 among others. Spalpha is able to bind to different cells of the immune system (monocytes and lymphocytes), which suggests that it may play an important role in the regulation of this system. To study Spalpha, an episomal mammalian expression system (pCEP-Pu/HEK 293-EBNA) was used to produce a recombinant form (rSpalpha) that was utilized for biochemical studies and for the generation of specific hybridomas. Four monoclonal antibodies were selected for their reactivity against rSpalpha by Western blot, immunoprecipitation, and enzyme-linked immunosorbent assays. The monoclonal antibodies recognized three different epitopes on Spalpha. The monoclonal antibodies revealed the existence of two Spalpha isoforms of 38 and 40 kDa, resulting from different sialic acid content. They also showed that Spalpha is a relatively abundant serum protein (60 micro g/ml) that mostly circulates in association with other serum proteins. Accordingly, rSpalpha allowed affinity chromatography isolation of polyclonal and monoclonal immunoglobulin M (IgM). These data indicate that Spalpha is a circulating protein that may play a role in the homeostasis of IgM antibodies.

Calyculaglycosides D and E, novel cembrane glycosides from the Caribbean gorgonian octocoral Eunicea species and structural revision of the aglycon of calyculaglycosides A-C.

A recent investigation of the constituents of Eunicea sp. from Colombia and Puerto Rico has provided two novel cembrane glycosides, calyculaglycosides D (6) and E (7), and a new cembranoid diterpene in free form, (+)-nephthenol (4). Metabolites 6 and 7 possess a glycosyl-fused cembrane skeleton with a 1S configuration. Biogenetic considerations have led to a revision of the previously assigned dilophol skeleton of calyculaglycosides A-C, which was confirmed by extensive 2D NMR investigation and a chemical degradation study. Consequently, the true structures for calyculaglycosides A-C are 8-10, respectively, not 1-3. This is the first report of the occurrence of marine diterpene glycosides having a cembrane aglycon.

High circulating levels of soluble scavenger receptors (sCD5 and sCD6) in patients with primary Sjögren's syndrome.

OBJECTIVE: To determine the existence of circulating levels of soluble scavenger receptors (sCD5 and sCD6) in patients with primary Sjögren's syndrome (SS), and to analyse the correlation with clinical and immunological features of SS. METHODS: Ninety consecutive patients with primary SS were studied. All patients fulfilled four or more of the European diagnostic criteria for SS. sCD5 and sCD6 levels were determined using a specific enzyme-linked immunosorbent assay (ELISA) developed in our laboratory. RESULTS: Detectable levels of sCD5 were found in 39 (43%) SS patients. The mean+/-standard error values of sCD5 were 3.5+/-0.5 ng/ml for patients with SS and 1.9+/-0.1 ng/ml for healthy blood donors (P<0.001). We found higher levels of sCD5 in patients with hypocomplementaemia (6.5 vs 3.5 ng/ml, P=0.03) and cryoglobulinaemia (6.9 vs 2.6 ng/ml, P=0.001). On the other hand, detectable levels of sCD6 were found in 60 (67%) SS patients. The mean+/-standard error values of sCD6 were 25.5+/-7.8 ng/ml in SS patients and 5.27+/-1.40 ng/ml in healthy blood donors (P=0.01). When the sCD6 levels were compared according to the presence or absence of immunological features, patients with cryoglobulinaemia showed higher levels of circulating sCD6 (77.3 vs 17 ng/ml, P=0.01) than those without cryoglobulinaemia. CONCLUSION: Patients with primary SS showed higher levels of circulating sCD5 and sCD6 when compared with controls. Moreover, the existence of some immunological features (hypocomplementaemia and cryoglobulinaemia) was associated with high levels of both soluble scavenger receptors. These facts may reflect an enhanced lymphocytic activation in patients with primary SS.

Residues Y429 and Y463 of the human CD5 are targeted by protein tyrosine kinases.

The human CD5 lymphocyte cell surface co-receptor modulates activation and differentiation responses mediated by the antigen-specific receptor of T and B cells. CD5 is phosphorylated following lymphocyte activation; however, the exact sites and kinases involved are yet to be determined. Jurkat T cell transfectants expressing tyrosine-mutated CD5 molecules have been used to show that residues Y429 and Y463 are targeted in vivo by protein tyrosine kinases following cell stimulation with anti-CD3 mAb or pervanadate. This is in agreement with data from direct in vitro kinase assays using purified recombinant Lck and Fyn protein tyrosine kinases. The analysis of Lck- and CD3-deficient Jurkat cells shows that tyrosine phosphorylation of CD5 requires Lck activity. We propose that T cell activation mediates CD5 tyrosine phosphorylation at residues Y429 and Y463 mainly through the activation of Lck.