Quantitation of alternatively spliced NMDA receptor NR1 isoform mRNA transcripts in human brain by competitive RT-PCR

The N-methyl-D-aspartate (NMDA)-selective subtype of ionotropic glutamate receptor is of importance in neuronal differentiation and synapse consolidation, activity-dependent forms of synaptic plasticity, and excitatory amino acid-mediated neuronal toxicity [Neurosci. Res. Program, Bull. 19 (1981) 1; Lab. Invest. 68 (1993) 372]. NMDA receptors exist in vivo as tetrameric or pentameric complexes comprising proteins from two families of homologous subunits, designated NR1 and NR2(A-D) [Biochem. Biophys. Res. Commun. 185 (1992) 826]. The gene coding for the human NR1 subunit (hNR1) is composed of 21 exons, three of which (4, 20 and 21) can be differentially spliced to generate a total of eight distinct subunit variants. We detail here a competitive RT-PCR (cRT-PCR) protocol to quantify endogenous levels of hNR1 splice variants in autopsied human brain. Quantitation of each hNR1 splice variant is performed using standard curve methodology in which a known amount of synthetic ribonucleic acid competitor (internal standard) is co-amplified against total RNA. This method can be used for the quantitation of hNR1 mRNA levels in response to acute or chronic disease states, in particular in the glutamatergic-associated neuronal loss observed in Alzheimer's disease [J. Neurochem. 78 (2001) 175]. Furthermore, alterations in hNR1 mRNA expression may be reflected at the translational level, resulting in functional changes in the NMDA receptor. (C) 2003 Elsevier Science B.V. All rights reserved.

Effect of changing data collection parameters on statistical motor unit number estimates

The effect of number of samples and selection of data for analysis on the calculation of surface motor unit potential (SMUP) size in the statistical method of motor unit number estimates (MUNE) was determined in 10 normal subjects and 10 with amyotrophic lateral sclerosis (ALS). We recorded 500 sequential compound muscle action potentials (CMAPs) at three different stable stimulus intensities (10���50% of maximal CMAP). Estimated mean SMUP sizes were calculated using Poisson statistical assumptions from the variance of 500 sequential CMAP obtained at each stimulus intensity. The results with the 500 data points were compared with smaller subsets from the same data set. The results using a range of 50���80% of the 500 data points were compared with the full 500. The effect of restricting analysis to data between 5���20% of the CMAP and to standard deviation limits was also assessed. No differences in mean SMUP size were found with stimulus intensity or use of different ranges of data. Consistency was improved with a greater sample number. Data within 5% of CMAP size gave both increased consistency and reduced mean SMUP size in many subjects, but excluded valid responses present at that stimulus intensity. These changes were more prominent in ALS patients in whom the presence of isolated SMUP responses was a striking difference from normal subjects. Noise, spurious data, and large SMUP limited the Poisson assumptions. When these factors are considered, consistent statistical MUNE can be calculated from a continuous sequence of data points. A 2 to 2.5 SD or 10% window are reasonable methods of limiting data for analysis. Muscle Nerve 27: 320���331, 2003

Immunological changes after cancer treatment and participation in an exercise program

Purpose: The purpose of this investigation was to evaluate the impact of undertaking peripheral blood stem cell transplantation (PBST) on T-cell number and function, and to determine the role of a mixed type, moderate intensity exercise program in facilitating the recovery of T-cell number and function. Methods: Immunological measures of white blood cell, lymphocyte, CD3(+), CD4(+), and CD8(+) counts, and CD3(+) cell function were assessed pretransplant (PI), immediately posttransplant (PII), and 1 month (II), 2 months (12) and 3 months (PIII) posttransplant. After PII, 12 patients were divided equally into a control group (CG) or exercise intervention group (EG). Results: Lower total T-cell, helper T-cell, and suppressor T-cell counts (P < 0.01), as well as lower T-cell function (P < 0.01), when compared with normative data, were found at PI. More specifically, 88% of the group had CD3(+), CD4(+), and CD8(+) counts that were more than 40%, 20%, and 50% below normal at PI, respectively. Undertaking a PBST caused further adverse changes to the total leukocyte, lymphocyte, CD3(+), CD4(+) and CD8(+) count. and the helper/suppressor ratio. Although CD8(+) counts had returned to normal by PIII, CD3(+), CD4(+), and the CD4(+)/CD8(+) ratio remained significantly lower than normative data (P < 0.01), with 66%, 100%, and 100% of the subject group reporting counts and ratios, respectively, below the normal range. Conclusion: The PBST patients were immunocompromised before undertaking the transplant, and the transplant procedure imposed further adverse changes to the leukocyte and lymphocyte counts. The leukocyte and CD8(+) counts returned to normal within 3 months posttransplant; however, the other immunological parameters assessed demonstrated a delayed recovery. Although participation in the exercise program did not facilitate a faster immune cell recovery, neither did the exercise program hinder or delay recovery.

Molecular basis of sulfonylurea herbicide inhibition of acetohydroxyacid synthase

Acetohydroxyacid synthase (AHAS) (acetolactate synthase, EC 4.1.3.18) catalyzes the first step in branchedchain amino acid biosynthesis and is the target for sulfonylurea and imidazolinone herbicides. These compounds are potent and selective inhibitors, but their binding site on AHAS has not been elucidated. Here we report the 2.8 Angstrom resolution crystal structure of yeast AHAS in complex with a sulfonylurea herbicide, chlorimuron ethyl. The inhibitor, which has a K-i of 3.3 nM blocks access to the active site and contacts multiple residues where mutation results in herbicide resistance. The structure provides a starting point for the rational design of further herbicidal compounds.

High rate of detection of primary aldosteronism, including surgically treatable forms, after 'non-selective' screening of hypertensive patients

Background Wide testing of the aldosterone: renin ratio among hypertensive individuals has revealed primary aldosteronism to be common, with most patients normokalaemic. Some investigators, however, have reported aldosterone-producing adenoma to be rare among patients so detected. Objective To test the hypothesis that differences among reported studies in the rate of detection of aldosterone-producing adenoma (as opposed to bilateral adrenal hyperplasia) reflect differences in the procedures used for diagnosis of primary aldosteronism, and the methods used to identify aldosterone-producing adenomas. Methods In the newly established Princess Alexandra Hospital Hypertension Unit (PAHHU), we used procedures developed by Greenslopes Hospital Hypertension Unit (which reports that more than 30% of patients with primary aldosteronism have aldosterone-producing adenomas) to diagnose primary aldosteronism and determine the subtype. All patients with an increased aldosterone: renin ratio (measured after correction for hypokalaemia and while the patient was not receiving interfering medications) underwent fludrocortisone suppression testing to confirm or exclude primary aldosteronism; if they were positive, they underwent genetic testing to exclude glucocorticoid-remediable aldosteronism before adrenal venous sampling was used to differentiate lateralizing from bilateral primary aldosteronism. Results This approach allowed PAHHU to diagnose, within 2 years, 54 patients [only seven (13%) hypokalaemic] with primary aldosteronism. All tested negative for glucocorticoid-remediable aldosteronism. Aldosterone production was lateralized to one adrenal in 15 patients (31%; only six hypokalaemic) and was bilateral in 34 (69%; all normokalaemic) of 49 patients who underwent adrenal venous sampling. Among patients with lateralizing adrenal hyperplasia, computed tomography revealed an ipsilateral mass in only six and a contralateral lesion in one. Fourteen patients underwent unilateral adrenalectomy, which cured the hypertension in seven and improved it in the remainder. In patients with bilateral primary aldlosteronism, hypertension responded to spironolactone (112.5-50 mg/ day) or amiloride (2.5-10 mg/day). Conclusion When performed with careful regard to confounding factors, measurement of the aldosterone: renin ratio in all hypertensive individuals, followed by fludrocortisone suppression testing to confirm or exclude primary aldosteronism and adrenal venous sampling to determine the subtype, can result in the detection of significant numbers of patients with specifically treatable or potentially curable hypertension. (C) 2003 Lippincott Williams Wilkins.

Selective entrainment of sediment graded by size and density under waves

The influence of near-bed sorting processes on heavy mineral content in suspension is discussed. Sediment concentrations above a rippled bed of mixed quartz and heavy mineral sand were measured under regular nonbreaking waves in the laboratory. Using the traditional gradient diffusion process, settling velocity would be expected to strongly affect sediment distribution. This was not observed during present trials. In fact, the vertical gradients of time-averaged suspension concentrations were found to be similar for the light and heavy minerals, despite their different settling velocities. This behavior implies a convective rather than diffusive distribution mechanism. Between the nonmoving bed and the lowest suspension sampling point, fight and heavy mineral concentration differs by two orders of magnitude. This discrimination against the heavy minerals in the pickup process is due largely to selective entrainment at the ripple face. Bed-form dynamics and the nature of quartz suspension profiles are found to be little affected by the trialed proportion of overall heavy minerals in the bed (3.8-22.1%).

Conselhos municipais de pol??ticas p??blicas: uma an??lise explorat??ria

Os Conselhos Gestores s??o f??runs h??bridos onde se viabiliza a participa????o popular no desenho das pol??ticas p??blicas, nas negocia????es de interesses da coletividade, na elabora????o de programas e projetos sociais e na fiscaliza????o das a????es governamentais. O significado e a dimens??o dos Conselhos para a democracia participativa s??o enormes, justificando a import??ncia da realiza????o deste estudo, que se baseou na an??lise explorat??ria de 12 Conselhos locais do munic??pio mineiro de Vi??osa, com o objetivo de avaliar o desempenho dos mesmos. A avalia????o do desempenho levou em considera????o os mesmos par??metros dimensionais, tanto para a coleta de dados prim??rios quanto para a coleta secund??ria. A coleta de dados prim??rios foi realizada a partir da aplica????o de question??rios semi-estruturados aos presidentes dos Conselhos em an??lise. Utilizou-se o programa SPSS, vers??o 15.0, para an??lise estat??stica das quest??es objetivas dos question??rios e a t??cnica de an??lise de conte??do para exame das quest??es abertas. Ap??s essa etapa, realizou-se um workshop com os presidentes dos Conselhos, visando ?? obten????o de novos dados e informa????es. Os Conselhos municipais analisados apresentaram resultados pouco discrepantes entre a teoria aplicada ao tema e a sua efetiva pr??tica no campo p??blico. Contudo, a pesquisa revelou que o grau de desenvolvimento institucional dos Conselhos n??o ?? homog??neo, ou seja, h?? alguns melhores gestores que outros. A escolha independente e democr??tica dos membros n??o-governamentais dos Conselhos em Vi??osa revelou o seu grau de independ??ncia do Poder Executivo. Por??m, a capacita????o de conselheiros precisa ser implementada para evitar uma desfigura????o dos Conselhos. No que tange ?? governan??a, destacouse em Vi??osa o seu grau e formas de aplicabilidade observadas na gest??o de v??rias pol??ticas. Da mesma forma, foi observado um n??vel de governabilidade que confere legitimidade ??s pol??ticas implementadas. Por fim, verificou-se que a efetividade da accountability societal tem sido instrumentalizada pelos Conselhos no ??mbito das diversas pol??ticas setoriais. J?? os dados secund??rios da an??lise foram obtidos por meio das s??nteses dos relat??rios de fiscaliza????o de munic??pios do Estado de Minas Gerais, realizadas pela Controladoria Geral da Uni??o. Foram identificados todos os registros pertinentes aos Conselhos Gestores Municipais do Estado de Minas Gerais, no per??odo de 2003 a 2006. Verificou-se que os maiores problemas se referem aos Conselhos Municipais de Sa??de e Assist??ncia Social. Entre as in??meras falhas apontadas, destacam-se problemas graves na constitui????o e operacionaliza????o dos Conselhos, resultando na inoper??ncia desses mecanismos nos munic??pios fiscalizados. A avalia????o do desempenho dos Conselhos locais de Vi??osa possibilitou concluir que esses mecanismos apresentaram um desempenho bem satisfat??rio na gest??o e controle das pol??ticas p??blicas no munic??pio.

Museu e escola : espa��os de sentidos

Esta pesquisa procurou compreender como o Museu de Arte do Esp��rito Santo (MAES) atua na produ����o de sentidos dos estudantes, a partir das rela����es estabelecidas com a escola, e, nesse proceder, como o museu compreende a arte e a escola como destinat��ria de suas a����es. Os sujeitos da pesquisa foram 98 crian��as, do 5�� ano do Ensino Fundamental I, de uma escola privada de Vit��ria. A pesquisa teve como corpus os discursos verbo-visuais produzidos pelas crian��as nessa rela����o museu e escola, bem como as performances discursivas do Programa de A����o Educativa do Museu, no que concerne �� forma����o de educadores e ao programa ciclo de palestras. Como metodologia de pesquisa, utilizou-se o estudo explorat��rio, e o aporte te��rico foi a semi��tica discursiva. A coleta de dados ocorreu no museu e na escola. No Museu, a partir de eventos que nortearam a exposi����o ���Meu Pa��s Tropical���, da artista alem��, Heidi Lieberman, e das a����es educativas propostas pelo MAES; na Escola, com o mapeamento inicial para compreender como e quais os sentidos eram apontados pelas crian��as quanto ao conhecimento delas sobre a arte e sobre um museu de arte (antes da realiza����o de sua primeira visita a esse espa��o, o que elas sabem e qual a compreens��o que elas t��m da arte), bem como, nas produ����es pl��sticas e verbo-visuais produzidas por elas ap��s a visita �� institui����o museal. Os resultados da pesquisa apontam que a metodologia anal��tica da semi��tica discursiva contribuiu, de forma significativa, para desvelar os sentidos apontados pelas crian��as no museu e na escola. Por meio dos discursos verbo-visuais produzidos por elas, compreendeu-se que uma educa����o pelo sens��vel, em conson��ncia com rela����es de bons encontros no museu e na escola, pode propiciar uma experi��ncia singular e significativa, que aguce a sensibilidade e reencante a educa����o.

Genetic divergence in cocoa progenies for backcrossing program to witches' broom disease resistance

Backcrossing has been little used in cacao breeding, particularly due to the long time required to transfer genes and recover the genetic background of the recurrent parent. The objective of this study was to select individuals, resulting from the backcross CEPEC-42 x SIC-19, genetically related to the recurrent parent SIC-19 by using RAPD molecular markers, among those with resistance to witches' broom. Of the 31 plants that clustered with SIC-19, 18 from the replanted material remained free of the disease in the field, with good vegetative aspect and, therefore can be used for backcross to reach the desired objective.

Thermal skin reference values in healthy late pregnancy

Digital thermal imaging has been employed in medicine for over 50 years. However, its use has been focused on vascular, musculoskeletal and neurological conditions, while other potential applications,such as obstetrics, have been much less explored. This paper presents a study conducted during 2011 at the Hospital of Braga on a group of healthy pregnant women in the last third of gestation. The analysis focused on characterizing typical pregnant women steady temperature profiles in specific defined regions of interest (ROI), and determining if the thermal symmetry values for late pregnant healthy women are in line with the values for non-pregnant healthy women. A temperature distribution pattern was found in the defined ROI. The obtained thermal symmetry value had a maximum of 0.370.2 1C, and there was no evidence for the influence of age (p40.05) in the observed group. The influence of the BMI requires further investigation since one ROI (P2 right) presented a p��0.059, close to the threshold of statistical evidence in the influence of BMI. The study group presented symmetry values in line with non-pregnant reference values, but the profiles in temperature distribution are different. Assumptions can therefore now be used with higher confidence when assessing abnormalities in specific pathologic states during pregnancy using the defined ROI. This work represents a first contribution towards establishing guidelines for future research in this specific field of study.

Optimizing electrode positioning in 3-lead ECG chest devices

Although the 12-lead electrocardiogram has become an essential medical and research tool, many current and envisaged applications would benefit from simpler devices, using 3-lead ECG configuration. This is particularly true for Ambient Assisted Living (in a broad perspective). However, the chest anatomy of female patients, namely during pregnancy, can hamper the adequate placement of a 3-lead ECG device and, very often, electrodes are placed below the chest rather than at the precise thoracic landmarks. Thus, the aim of this study was to compare the effect of electrode positioning on the ECG signal of pregnant women and provide guidelines for device development. The effect of breast tissue on the ECG signal was investigated by relating breast size with the signal-to-noise ratio, root mean square and R-wave amplitude. Results show that the 3-lead ECG should be placed on the breast rather than under the breast and indicate positive correlation between breast size and signal-to-noise ratio.

The need for standardized tests to evaluate the reliability of data transport in wireless medical systems

Wireless medical systems are comprised of four stages, namely the medical device, the data transport, the data collection and the data evaluation stages. Whereas the performance of the first stage is highly regulated, the others are not. This paper concentrates on the data transport stage and argues that it is necessary to establish standardized tests to be used by medical device manufacturers to provide comparable results concerning the communication performance of the wireless networks used to transport medical data. Besides, it suggests test parameters and procedures to be used to produce comparable communication performance results.

Program slicing by calculation

Program slicing is a well known family of techniques used to identify code fragments which depend on or are depended upon specific program entities. They are particularly useful in the areas of reverse engineering, program understanding, testing and software maintenance. Most slicing methods, usually oriented towards the imperative or object paradigms, are based on some sort of graph structure representing program dependencies. Slicing techniques amount, therefore, to (sophisticated) graph transversal algorithms. This paper proposes a completely different approach to the slicing problem for functional programs. Instead of extracting program information to build an underlying dependencies��� structure, we resort to standard program calculation strategies, based on the so-called Bird-Meertens formalism. The slicing criterion is specified either as a projection or a hiding function which, once composed with the original program, leads to the identification of the intended slice. Going through a number of examples, the paper suggests this approach may be an interesting, even if not completely general, alternative to slicing functional programs

Component identification through program slicing

This paper reports on the development of specific slicing techniques for functional programs and their use for the identification of possible coherent components from monolithic code. An associated tool is also introduced. This piece of research is part of a broader project on program understanding and re-engineering of legacy code supported by formal methods