Structure and function in rhodopsin: Kinetic studies of retinal binding to purified opsin mutants in defined phospholipid–detergent mixtures serve as probes of the retinal binding pocket

In the current standard procedure for preparation of mammalian rhodopsin mutants, transfected COS-1 cells expressing the mutant opsin genes are treated with 5 μM 11-cis-retinal before detergent solubilization for purification. We found that binding of 11-cis-retinal to opsin mutants with single amino acid changes at Trp-265 (W265F,Y,A) and a retinitis pigmentosa mutant (A164V) was far from complete and required much higher concentrations of 11-cis-retinal. By isolation of the expressed opsins in a stable form, kinetic studies of retinal binding to the opsins in vitro have been carried out by using defined phospholipid–detergent mixtures. The results show wide variation in the rates of 11-cis-retinal binding. Thus, the in vitro reconstitution procedure serves as a probe of the retinal-binding pocket in the opsins. Further, a method is described for purification and characterization of the rhodopsin mutants after retinal binding to the opsins in vitro.

A methodology to estimate time varying user responses to travel time and travel time reliability in a road pricing environment

Road pricing has emerged as an effective means of managing road traffic demand while simultaneously raising additional revenues to transportation agencies. Research on the factors that govern travel decisions has shown that user preferences may be a function of the demographic characteristics of the individuals and the perceived trip attributes. However, it is not clear what are the actual trip attributes considered in the travel decision- making process, how these attributes are perceived by travelers, and how the set of trip attributes change as a function of the time of the day or from day to day. In this study, operational Intelligent Transportation Systems (ITS) archives are mined and the aggregated preferences for a priced system are extracted at a fine time aggregation level for an extended number of days. The resulting information is related to corresponding time-varying trip attributes such as travel time, travel time reliability, charged toll, and other parameters. The time-varying user preferences and trip attributes are linked together by means of a binary choice model (Logit) with a linear utility function on trip attributes. The trip attributes weights in the utility function are then dynamically estimated for each time of day by means of an adaptive, limited-memory discrete Kalman filter (ALMF). The relationship between traveler choices and travel time is assessed using different rules to capture the logic that best represents the traveler perception and the effect of the real-time information on the observed preferences. The impact of travel time reliability on traveler choices is investigated considering its multiple definitions. It can be concluded based on the results that using the ALMF algorithm allows a robust estimation of time-varying weights in the utility function at fine time aggregation levels. The high correlations among the trip attributes severely constrain the simultaneous estimation of their weights in the utility function. Despite the data limitations, it is found that, the ALMF algorithm can provide stable estimates of the choice parameters for some periods of the day. Finally, it is found that the daily variation of the user sensitivities for different periods of the day resembles a well-defined normal distribution.

The Metabolic Syndrome And Sexual Function In Climacteric Women: A Cross-sectional Study.

Studies have associated the metabolic syndrome with poor sexual function; the results, however, are controversial. To evaluate the relationship between the metabolic syndrome and sexual function and to identify the factors associated with poor sexual function. A secondary analysis of a cross-sectional cohort study including 256 women of 40-60 years of age receiving care at the outpatient department of a university teaching hospital. A specific questionnaire was applied to collect sociodemographic and behavioral data, and the Short Personal Experience Questionnaire was used to evaluate sexual function, with a score ≤ 7 being indicative of poor sexual function. Anthropometric measurements, blood pressure, fasting glucose, high-density lipoprotein, total cholesterol, triglycerides, follicle-stimulating hormone and thyroid stimulating hormone levels were determined. The prevalence of the metabolic syndrome, as defined by the International Diabetes Federation, was 62.1%, and the prevalence of poor sexual function was 31.4%. The only factor related to female sexual function that was associated with the metabolic syndrome was sexual dysfunction in the woman's partner. The factors associated with poor sexual function in the bivariate analysis were age >50 years (P=0.003), not having a partner (P<0.001), being postmenopausal (P=0.046), the presence of hot flashes (P=0.02), poor self-perception of health (P=0.04), partner's age ≥ 50 years, and time with partner ≥ 21 years. Reported active (P=0.02) and passive (P=0.01) oral sex was associated with an absence of sexual dysfunction. In the multiple regression analysis, the only factor associated with poor sexual function was being 50 years of age or more. The prevalence of the metabolic syndrome was high and was not associated with poor sexual function in this sample of menopausal women. The only factor associated with poor sexual function was being over 50 years of age.

Thyroid Function After Unilateral Total Lobectomy

Objective: To evaluate the incidence of postoperative hypothyroidism among patients who underwent unilateral total lobectomy and identify related factors. Design: Retrospective medical record analysis. Setting: Oncological center and private clinic. Patients: From March 1996 to July 2005, 228 euthyroid patients underwent unilateral total lobectomy for benign diseases; 168 had all the information required for inclusion in this study. Main Outcome Measures: Serum levels of thyrotropin and antithyroidal antibodies were assessed, as well as ultrasonographic evaluation of the remaining thyroid lobe and review of all histological specimens, with emphasis on lymphocytic infiltration. Hypothyroidism was defined as thyrotropin level greater than 5.5 mU/L. Results: Most patients were female (88%), with a median (range) age of 45 (16-72) years. Hypothyroidism occurred in 61 cases (32.8%), during a median follow-up period of 29 months (range, 6-108 months). Statistically related factors included higher preoperative thyrotropin levels (2.1 mU/L among hypothyroid patients vs 1.2 mU/L in euthyroid patients; P<.001), smaller thyroid remnant volume (3.9 mL vs; 6.0 mL, respectively; P = .003); right vs left lobectomy (P = .006), and higher thyroperoxidase antibody serum levels (P = .009). Conclusions: Postoperative hypothyroidism appeared in 32.8% of the cases in this series, especially among patients with elevated preoperative thyrotropin and postoperative thyroperoxidase antibody levels, after right lobectomy and when a smaller thyroid remnant was left. After confirmation with larger prospective series, these results may support the indication for early postoperative hormone supplementation in these instances.

Genetic association and cellular function of MC1R variant alleles in human pigmentation

We have examined MC1R variant allele frequencies in the general population of South East Queensland and in a collection of adolescent dizygotic and monozygotic twins and family members to define statistical associations with hair and skin color, freckling, and mole count. Results of these studies are consistent with a linear recessive allelic model with multiplicative penetrance in the inheritance of red hair. Four alleles, D84E, R151C, R160W, and D294H, are strongly associated with red hair and fair skin with multinomial regression analysis showing odds ratios of 63, 118, 50, and 94, respectively. An additional three low-penetrance alleles V60L, V92M, and R163Q have odds ratios 6, 5, and 2 relative to the wild-type allele. To address the cellular effects of MC1R variant alleles in signal transduction, we expressed these receptors in permanently transfected HEK293 cells. Measurement of receptor activity via induction of a cAMP-responsive luciferase reporter gene found that the R151C and R160W receptors were active in the presence of NDP-MSH ligand, but at much reduced levels compared with that seen with the wild-type receptor. The ability to stimulate phosphorylation of the cAMP response element binding protein (CREB) transcription factor was also apparent in all stimulated MC1R variant allele-expressing HEK293 cell extracts as assessed by immunoblotting. In contrast, human melanoma cell lines showed wide variation in the their ability to undergo cAMP-mediated CREB phosphorylation. Culture of human melanocytes of known MC1R genotype may provide the best experimental approach to examine the functional consequences for each MC1R variant allele. With this objective, we have established more than 300 melanocyte cell strains of defined MC1R genotype.

The role of BPH, lower urinary tract symptoms, and PSA levels on erectile function of Brazilian men who undergo prostate cancer screening

Introduction. Lower urinary tract symptoms (LUTS) and erectile dysfunction (ED) are common problems in middle-aged and older men. Recently, epidemiologic studies have shown significant associations between severity of LUTS and male sexual dysfunction. Aim. We analyzed the role of prostate enlargement, LUTS, and prostate specific antigen (PSA) levels in the erectile function of Brazilian men who underwent prostate cancer (PCa) screening. Method. We analyzed data from 1,008 consecutive patients enrolled in a PCa screening program. Benign prostatic hyperplasia (BPH) was defined as a prostate weight greater than 30 g as defined by digital rectal examination. For statistical analysis, we used the chi-squared and analysis of variance tests. The odds ratios (OR) for correlation of ED with prostate volume LUTS and PSA were estimated using logistic regression models. Main Outcome Measure. The American Urological Association (AUA) symptom score for LUTS and the International Index of Erectile Function. Results. Mean patient age was 61.2 years (45-87) and median PSA value was 1.9 ng/mL. BPH was identified in 48.5% of patients. Mild, moderate, and severe LUTS were found in 52.3%, 30.9%, and 16.8% of cases, respectively. ED was classified as absent, mild, mild to moderate, moderate, and severe in 18.6%, 23.1%, 18.6%, 15.2%, and 24.5%, respectively. While only 5.4% of the patients with no ED presented severe LUTS, this finding was observed in 27.1% of patients with severe ED (P<0.001). Univariate logistic regression analysis demonstrated that age, prostate volume, AUA symptom score, and PSA levels were significant predictors of ED. However, when controlled for patient age, only LUTS remained as an independent predictor of ED. Conclusions. Controlling for patient age, LUTS are independent risk factors for the development of ED among Brazilian men who undergo PCa screening.

Gag-Pol supplied in trans is efficiently packaged and supports viral function in human immunodeficiency virus type 1

The intracellular trafficking and subsequent incorporation of Gag-Pol into human immunodeficiency virus type 1 (HIV-1) remains poorly defined. Gag-Pol is encoded by the same mRNA as Gag and is generated by ribosomal frameshifting. The multimerization of Gag and Gag-Pol is an essential step in the formation of infectious viral particles. In this study, we examined whether the interaction between Gag and Gag-Pol is initiated during protein translation in order to facilitate the trafficking and subsequent packaging of Gag-Pol into the virion. A conditional cotransfection system was developed in which virion formation required the coexpression of two HIV-1-based plasmids, one that produces both Gag and Gag-Pol and one that only produces Gag-Pol. The Gag-Pol proteins were either immunotagged with a His epitope or functionally tagged with a mutation (K65R) in reverse transcriptase that is associated with drug resistance. Gag-Pol packaging was assessed to determine whether the Gag-Pol incorporated into the virion was preferentially packaged from the plasmid that expressed both Gag and Gag-Pol or whether it could be packaged from either plasmid. Our data show that translation of Gag and Gag-Pol from the same mRNA is not critical for virion packaging of the Gag-Pol polyprotein or for viral function.

Determinants of tissue Doppler measures of regional diastolic function during dobutamine stress echocardiography

Background Diastolic dysfunction induced by ischemia may alter transmitral blood flow, but this reflects global ventricular function, and pseudonormalization may occur with increased preload. Tissue Doppler may assess regional diastolic function and is relatively load-independent, but limited data exist regarding its application to stress testing. We sought to examine the stress response of regional diastolic parameters to dobutomine echocardiography (DbE). Methods Sixty-three patients underwent study with DbE: 20 with low probability of coronary artery disease (CAD) and 43 with CAD who underwent angiography. A standard DbE protocol was used, and segments were categorized as ischemic, scar, or normal. Color tissue Doppler was acquired at baseline and peak stress, and waveforms in the basal and mid segments were used to measure early filling (Em), late filling (Am), and E deceleration time. Significant CAD was defined by stenoses >50% vessel diameter. Results Diastolic parameters had limited feasibility because of merging of Em and Am waves at high heart rates and limited reproducibility. Nonetheless, compared with normal segments, segments subtended with significant stenoses showed a lower Em velocity at rest (6.2 +/- 2.6 cm/s vs 4.8 +/- 2.2 cm/s, P < .0001) and peak (7.5 +/- 4.2 cm/s vs 5.1 +/- 3.6 cm/s, P < .0001), Abnormal segments also showed a shorter E deceleration time (51 +/- 27 ms vs 41 +/- 27 ms, P = .0001) at base and peak. No changes were documented in Am. The same pattern was seen with segments identified as ischemic with wall motion score. However, in the absence of ischemia, segments of patients with left ventricular hypertrophy showed a lower Em velocity, with blunted Em responses to stress. Conclusion Regional diastolic function is sensitive to ischemia. However, a number of practical limitations limit the applicability of diastolic parameters for the quantification of stress echocardiography.

CD40 and dendritic cell function

CD40 has emerged as a key signaling pathway for the function of B cells, monocytes, and dendritic cells (DC) in the immune system, and plays a major role in inflammatory pathways of nonhemopoletic cells. CD40 is expressed by monocytes and DC and is up-regulated when DC migrate from the periphery to draining lymph nodes (DLN) in response to microbial challenge. CD154 signaling by MHC-restricted, activated CD4* T cells induces differentiation of DC, as defined by an increased surface expression of MHC, costimulatory, and adhesion molecules. Thus, CD40 functions in the adaptive immune response as a trigger for the expression of costimulatory molecules for efficient T-cell activation. CD40 ligation of DC also has the capacity to induce high levels of the cytokine IL-12, which polarizes CD4(+) T cells toward a T helper 1 (Th1) type, enhances proliferation of CD8(+) T cells, and activates NK cells. CD40 may also play an important role in the decision between tolerance and immunity and the generation of regulatory CD4(+) T cells that are thought to maintain peripheral self-tolerance in vivo.

Relationship of ventricular longitudinal function to contractile reserve in patients with mitral regurgitation

Background Latent left ventricular (LV) dysfunction in patients with valvular or myocardial disease may be identified by loss of contractile reserve (CR) at exercise echocardiography. Contraction in the LV longitudinal axis may be more sensitive than radial contraction to minor disturbances of LV function. We sought to determine whether tissue Doppler measurement of longitudinal function could be used to identify CR. Methods Exercise echocardiography was performed in 86 patients (20 women, age 53 +/- 18 years), 72 with asymptomatic or minimally symptomatic mitral regurgitation, and 14 normal controls. Pulsed-wave tissue Doppler imaging (DTI) was used to measure maximum annular systolic velocity at rest and stress. Inducible ischemia was excluded by analysis of wall motion by an experienced observer. CR was defined by greater than or equal to5% improvement of stress compared with rest ejection fraction (EF). Exercise capacity was assessed from expired gas analysis. Results CR was present in 34 patients with mitral regurgitation (47%); peak EF in patients with and without CR was 74% +/- 11% versus 54% +/- 15% (P

Usefulness of quantitative echocardiographic techniques to predict recovery of regional and global left ventricular function after acute myocardial infarction

The left ventricular response to dobutamine may be quantified using tissue Doppler measurement of myocardial velocity or displacement or 3-dimensional echocardiography to measure ventricular volume and ejection fraction. This study sought to explore the accuracy of these methods for predicting segmental and global responses to therapy. Standard dobutamine and 3-dimensional echocardiography were performed in 92 consecutive patients with abnormal left ventricular function at rest. Recovery of function was defined by comparison with follow-up echocardiography at rest 5 months later. Segments that showed improved regional function at follow-up showed a higher increment in peak tissue Doppler velocity with dobutamine therapy than in nonviable segments (1.2 +/- 0.4 vs 0.3 +/- 0.2 cm/s, p = 0.001). Similarly, patients who showed a > 5% improvement of ejection fraction at follow-up showed a greater displacement response to dobutamine (6.9 +/- 3.2 vs 2.1 +/- 2.3 mm, p = 0.001), as well as a higher rate of ejection fraction, response to dobutamine (9 +/- 3% vs 2 +/- 2%, p = 0.001). The optimal cutoff values for predicting subsequent recovery of function at rest were an increment of peak velocity > 1 cm/s, >5 mm of displacement, and a >5% improvement of ejection fraction with low-dose dobutamine. (C) 2003 by Excerpta Medica, Inc.

Lightweight user modeling – a case study

In the context of previous publications, we propose a new lightweight UM process, intended to work as a tourism recommender system in a commercial environment. The new process tackles issues like cold start, gray sheep and over specialization through a rich user model and the application of a gradual forgetting function to the collected user action history. Also, significant performance improvements were achieved regarding the previously proposed UM process.

Complex question answering on semi-structured repositories: a user centric process enhanced with context

A Teia Mundial (Web) foi prevista como uma rede de documentos de hipertexto interligados de forma a criar uma espaço de informação onde humanos e máquinas poderiam comunicar. No entanto, a informação contida na Web tradicional foi/é armazenada de forma não estruturada o que leva a que apenas os humanos a possam consumir convenientemente. Consequentemente, a procura de informações na Web sintáctica é uma tarefa principalmente executada pelos humanos e nesse sentido nem sempre é fácil de concretizar. Neste contexto, tornou-se essencial a evolução para uma Web mais estruturada e mais significativa onde é dado significado bem definido à informação de forma a permitir a cooperação entre humanos e máquinas. Esta Web é usualmente referida como Web Semântica. Além disso, a Web Semântica é totalmente alcançável apenas se os dados de diferentes fontes forem ligados criando assim um repositório de Dados Abertos Ligados (LOD). Com o aparecimento de uma nova Web de Dados (Abertos) Ligados (i.e. a Web Semântica), novas oportunidades e desafios surgiram. Pergunta Resposta (QA) sobre informação semântica é actualmente uma área de investigação activa que tenta tirar vantagens do uso das tecnologias ligadas à Web Semântica para melhorar a tarefa de responder a questões. O principal objectivo do projecto World Search passa por explorar a Web Semântica para criar mecanismos que suportem os utilizadores de domínios de aplicação específicos a responder a questões complexas com base em dados oriundos de diferentes repositórios. No entanto, a avaliação feita ao estado da arte permite concluir que as aplicações existentes não suportam os utilizadores na resposta a questões complexas. Nesse sentido, o trabalho desenvolvido neste documento foca-se em estudar/desenvolver metodologias/processos que permitam ajudar os utilizadores a encontrar respostas exactas/corretas para questões complexas que não podem ser respondidas fazendo uso dos sistemas tradicionais. Tal inclui: (i) Ultrapassar a dificuldade dos utilizadores visionarem o esquema subjacente aos repositórios de conhecimento; (ii) Fazer a ponte entre a linguagem natural expressa pelos utilizadores e a linguagem (formal) entendível pelos repositórios; (iii) Processar e retornar informações relevantes que respondem apropriadamente às questões dos utilizadores. Para esse efeito, são identificadas um conjunto de funcionalidades que são consideradas necessárias para suportar o utilizador na resposta a questões complexas. É também fornecida uma descrição formal dessas funcionalidades. A proposta é materializada num protótipo que implementa as funcionalidades previamente descritas. As experiências realizadas com o protótipo desenvolvido demonstram que os utilizadores efectivamente beneficiam das funcionalidades apresentadas: ▪ Pois estas permitem que os utilizadores naveguem eficientemente sobre os repositórios de informação; ▪ O fosso entre as conceptualizações dos diferentes intervenientes é minimizado; ▪ Os utilizadores conseguem responder a questões complexas que não conseguiam responder com os sistemas tradicionais. Em suma, este documento apresenta uma proposta que comprovadamente permite, de forma orientada pelo utilizador, responder a questões complexas em repositórios semiestruturados.

Spectrum sensing through software defined radio

Dissertação apresentada para obtenção do Grau de Mestre em Engenharia Electrotécnica e de Computadores, pela Universidade Nova de Lisboa, Faculdade de Ciências e Tecnologia

Differential top-antitop cross-section measurements as a function of observables constructed from final-state particles using pp collisions at s√=7 TeV in the ATLAS detector

Various differential cross-sections are measured in top-quark pair (tt¯) events produced in proton--proton collisions at a centre-of-mass energy of s√=7 TeV at the LHC with the ATLAS detector. These differential cross-sections are presented in a data set corresponding to an integrated luminosity of 4.6 fb−1. The differential cross-sections are presented in terms of kinematic variables of a top-quark proxy referred to as the pseudo-top-quark whose dependence on theoretical models is minimal. The pseudo-top-quark can be defined in terms of either reconstructed detector objects or stable particles in an analogous way. The measurements are performed on tt¯ events in the lepton+jets channel, requiring exactly one charged lepton and at least four jets with at least two of them tagged as originating from a b-quark. The hadronic and leptonic pseudo-top-quarks are defined via the leptonic or hadronic decay mode of the W boson produced by the top-quark decay in events with a single charged lepton.The cross-section is measured as a function of the transverse momentum and rapidity of both the hadronic and leptonic pseudo-top-quark as well as the transverse momentum, rapidity and invariant mass of the pseudo-top-quark pair system. The measurements are corrected for detector effects and are presented within a kinematic range that closely matches the detector acceptance. Differential cross-section measurements of the pseudo-top-quark variables are compared with several Monte Carlo models that implement next-to-leading order or leading-order multi-leg matrix-element calculations.