Development and validation of a fast RP-HPLC method to determine the analogue of the thyroid hormone, 3,5,3 '-triiodothyroacetic acid (TRIAC), in polymeric nanoparticles

The objective of this work was to develop and validate a rapid Reversed-Phase High-Performance Liquid Chromatography method for the quantification of 3,5,3 '-triiodothyroacetic acid (TRIAC) in nanoparticles delivery system prepared in different polymeric matrices. Special attention was given to developing a reliable reproductive technique for the pretreatment of the samples. Chromatographic runs were performed on an Agilent 1200 Series HPLC with a RP Phenomenex (R) Gemini C18 (150 x 4, 6 mm i.d., 5 mu m) column using acetonitrile and triethylamine buffer 0.1% (TEA) (40 : 60 v/v) as a mobile phase in an isocratic elution, pH 5.6 at a flow rate of 1 ml min(-1). TRIAC was detected at a wavelength of 220 nm. The injection volume was 20 mu l and the column temperature was maintained at 35 degrees C. The validation characteristics included accuracy, precision, specificity, linearity, recovery, and robustness. The standard curve was found to have a linear relationship (r(2) - 0.9996) over the analytical range of 5-100 mu g ml(-1) . The detection and quantitation limits were 1.3 and 3.8 mu g ml(-1), respectively. The recovery and loaded TRIAC in colloidal system delivery was nearly 100% and 98%, respectively. The method was successfully applied in polycaprolactone, polyhydroxybutyrate, and polymethylmethacrylate nanoparticles.

Gene Expression Complex Networks: Synthesis, Identification, and Analysis

Thanks to recent advances in molecular biology, allied to an ever increasing amount of experimental data, the functional state of thousands of genes can now be extracted simultaneously by using methods such as cDNA microarrays and RNA-Seq. Particularly important related investigations are the modeling and identification of gene regulatory networks from expression data sets. Such a knowledge is fundamental for many applications, such as disease treatment, therapeutic intervention strategies and drugs design, as well as for planning high-throughput new experiments. Methods have been developed for gene networks modeling and identification from expression profiles. However, an important open problem regards how to validate such approaches and its results. This work presents an objective approach for validation of gene network modeling and identification which comprises the following three main aspects: (1) Artificial Gene Networks (AGNs) model generation through theoretical models of complex networks, which is used to simulate temporal expression data; (2) a computational method for gene network identification from the simulated data, which is founded on a feature selection approach where a target gene is fixed and the expression profile is observed for all other genes in order to identify a relevant subset of predictors; and (3) validation of the identified AGN-based network through comparison with the original network. The proposed framework allows several types of AGNs to be generated and used in order to simulate temporal expression data. The results of the network identification method can then be compared to the original network in order to estimate its properties and accuracy. Some of the most important theoretical models of complex networks have been assessed: the uniformly-random Erdos-Renyi (ER), the small-world Watts-Strogatz (WS), the scale-free Barabasi-Albert (BA), and geographical networks (GG). The experimental results indicate that the inference method was sensitive to average degree k variation, decreasing its network recovery rate with the increase of k. The signal size was important for the inference method to get better accuracy in the network identification rate, presenting very good results with small expression profiles. However, the adopted inference method was not sensible to recognize distinct structures of interaction among genes, presenting a similar behavior when applied to different network topologies. In summary, the proposed framework, though simple, was adequate for the validation of the inferred networks by identifying some properties of the evaluated method, which can be extended to other inference methods.

Inference of gene regulatory networks from time series by Tsallis entropy

Background: The inference of gene regulatory networks (GRNs) from large-scale expression profiles is one of the most challenging problems of Systems Biology nowadays. Many techniques and models have been proposed for this task. However, it is not generally possible to recover the original topology with great accuracy, mainly due to the short time series data in face of the high complexity of the networks and the intrinsic noise of the expression measurements. In order to improve the accuracy of GRNs inference methods based on entropy (mutual information), a new criterion function is here proposed. Results: In this paper we introduce the use of generalized entropy proposed by Tsallis, for the inference of GRNs from time series expression profiles. The inference process is based on a feature selection approach and the conditional entropy is applied as criterion function. In order to assess the proposed methodology, the algorithm is applied to recover the network topology from temporal expressions generated by an artificial gene network (AGN) model as well as from the DREAM challenge. The adopted AGN is based on theoretical models of complex networks and its gene transference function is obtained from random drawing on the set of possible Boolean functions, thus creating its dynamics. On the other hand, DREAM time series data presents variation of network size and its topologies are based on real networks. The dynamics are generated by continuous differential equations with noise and perturbation. By adopting both data sources, it is possible to estimate the average quality of the inference with respect to different network topologies, transfer functions and network sizes. Conclusions: A remarkable improvement of accuracy was observed in the experimental results by reducing the number of false connections in the inferred topology by the non-Shannon entropy. The obtained best free parameter of the Tsallis entropy was on average in the range 2.5 <= q <= 3.5 (hence, subextensive entropy), which opens new perspectives for GRNs inference methods based on information theory and for investigation of the nonextensivity of such networks. The inference algorithm and criterion function proposed here were implemented and included in the DimReduction software, which is freely available at http://sourceforge.net/projects/dimreduction and http://code.google.com/p/dimreduction/.

Application of electron paramagnetic resonance Spectroscopy for validation of the novel (AN plus DN) solvent polarity scale

Based on solvation studies of polymers, the sum (1: 1) of the electron acceptor (AN) and electron donor (DN) values of solvents has been proposed as an alternative polarity scale. To test this, the electron paramagnetic resonance isotropic hyperfine splitting constant, a parameter known to be dependent on the polarity/proticity of the medium, was correlated with the (AN+DN) term using three paramagnetic probes. The linear regression coefficient calculated for 15 different solvents was approximately 0.9, quite similar to those of other well-known polarity parameters, attesting to the validity of the (AN+DN) term as a novel ""two-parameter"" solvent polarity scale.

Validation of dot blot hybridization and denaturing high performance liquid chromatography as reliable methods for TP53 codon 72 genotyping in molecular epidemiologic studies

Background: Mutations in TP53 are common events during carcinogenesis. In addition to gene mutations, several reports have focused on TP53 polymorphisms as risk factors for malignant disease. Many studies have highlighted that the status of the TP53 codon 72 polymorphism could influence cancer susceptibility. However, the results have been inconsistent and various methodological features can contribute to departures from Hardy-Weinberg equilibrium, a condition that may influence the disease risk estimates. The most widely accepted method of detecting genotyping error is to confirm genotypes by sequencing and/or via a separate method. Results: We developed two new genotyping methods for TP53 codon 72 polymorphism detection: Denaturing High Performance Liquid Chromatography (DHPLC) and Dot Blot hybridization. These methods were compared with Restriction Fragment Length Polymorphism (RFLP) using two different restriction enzymes. We observed high agreement among all methodologies assayed. Dot-blot hybridization and DHPLC results were more highly concordant with each other than when either of these methods was compared with RFLP. Conclusions: Although variations may occur, our results indicate that DHPLC and Dot Blot hybridization can be used as reliable screening methods for TP53 codon 72 polymorphism detection, especially in molecular epidemiologic studies, where high throughput methodologies are required.

Comparison of univariate and multivariate calibration for the determination of micronutrients in pellets of plant materials by laser induced breakdown spectrometry

The application of laser induced breakdown spectrometry (LIBS) aiming the direct analysis of plant materials is a great challenge that still needs efforts for its development and validation. In this way, a series of experimental approaches has been carried out in order to show that LIBS can be used as an alternative method to wet acid digestions based methods for analysis of agricultural and environmental samples. The large amount of information provided by LIBS spectra for these complex samples increases the difficulties for selecting the most appropriated wavelengths for each analyte. Some applications have suggested that improvements in both accuracy and precision can be achieved by the application of multivariate calibration in LIBS data when compared to the univariate regression developed with line emission intensities. In the present work, the performance of univariate and multivariate calibration, based on partial least squares regression (PLSR), was compared for analysis of pellets of plant materials made from an appropriate mixture of cryogenically ground samples with cellulose as the binding agent. The development of a specific PLSR model for each analyte and the selection of spectral regions containing only lines of the analyte of interest were the best conditions for the analysis. In this particular application, these models showed a similar performance. but PLSR seemed to be more robust due to a lower occurrence of outliers in comparison to the univariate method. Data suggests that efforts dealing with sample presentation and fitness of standards for LIBS analysis must be done in order to fulfill the boundary conditions for matrix independent development and validation. (C) 2009 Elsevier B.V. All rights reserved.

Optimization and validation of a LIBS method for the determination of macro and micronutrients in sugar cane leaves

Laser induced breakdown spectrometry (LIBS) was applied for the determination of macro (P, K, Ca, Mg) and micronutrients (B, Cu, Fe, Mn and Zn) in sugar cane leaves, which is one of the most economically important crops in Brazil. Operational conditions were previously optimized by a neuro-genetic approach, by using a laser Nd:YAG at 1064 nm with 110 mJ per pulse focused on a pellet surface prepared with ground plant samples. Emission intensities were measured after 2.0 mu s delay time, with 4.5 mu s integration time gate and 25 accumulated laser pulses. Measurements of LIBS spectra were based on triplicate and each replicate consisted of an average of ten spectra collected in different sites (craters) of the pellet. Quantitative determinations were carried out by using univariate calibration and chemometric methods, such as PLSR and iPLS. The calibration models were obtained by using 26 laboratory samples and the validation was carried out by using 15 test samples. For comparative purpose, these samples were also microwave-assisted digested and further analyzed by ICP OES. In general, most results obtained by LIBS did not differ significantly from ICP OES data by applying a t-test at 95% confidence level. Both LIBS multivariate and univariate calibration methods produced similar results, except for Fe where better results were achieved by the multivariate approach. Repeatability precision varied from 0.7 to 15% and 1.3 to 20% from measurements obtained by multivariate and univariate calibration, respectively. It is demonstrated that LIBS is a powerful tool for analysis of pellets of plant materials for determination of macro and micronutrients by choosing calibration and validation samples with similar matrix composition.

Contuse lesion of the rat spinal cord of moderate intensity leads to a higher time-dependent secondary neurodegeneration than severe one - An open-window for experimental neuroprotective interventions

Secondary neurodegeneration takes place in the surrounding tissue of spinal cord trauma and modifies substantially the prognosis, considering the small diameter of its transversal axis. We analyzed neuronal and glial responses in rat spinal cord after different degree of contusion promoted by the NYU Impactor. Rats were submitted to vertebrae laminectomy and received moderate or severe contusions. Control animals were sham operated. After 7 and 30 days post surgery, stereological analysis of Nissl staining cellular profiles showed a time progression of the lesion volume after moderate injury, but not after severe injury. The number of neurons was not altered cranial to injury. However, same degree of diminution was seen in the caudal cord 30 days after both severe and moderate injuries. Microdensitometric image analysis demonstrated a microglial reaction in the white matter 30 days after a moderate contusion and showed a widespread astroglial reaction in the white and gray matters 7 days after both severities. Astroglial activation lasted close to lesion and in areas related to Wallerian degeneration. Data showed a more protracted secondary degeneration in rat spinal cord after mild contusion, which offered an opportunity for neuroprotective approaches. Temporal and regional glial responses corroborated to diverse glial cell function in lesioned spinal cord. (C) 2007 Elsevier Ltd. All rights reserved.

Exercise prevents the effects of experimental arthritis on the metabolism and function of immune cells

Active lymphocytes (LY) and macrophages (M Phi) are involved in the pathophysiology of rheumatoid arthritis (RA) Due to its anti-inflammatory effect. physical exercise may be beneficial in RA by acting on the immune system (IS) Thus, female Wistar rats with type II collagen-induced arthritis (CIA) were submitted to swimming training (6 weeks. 5 days/week. 60 min/day) and some biochemical and immune parameters, such as the metabolism of glucose and glutamine and function of LY and M. were evaluated In addition, plasma levels of some hormones and of interleukin-2 (IL-2) were also determined Results demonstrate that CIA increased lymphocyte proliferation (1.9- and 1 7-fold, respectively, in response to concanavalin A (ConA) and lipopolysaccharide (LPS)), as well as macrophage H(2)O(2) production (1 6-fold), in comparison to control Exercise training prevented the activation of immune cells, induced by CIA. and established a pattern of substrate utilization similar to that described as normal for these cells. Exercise also promoted an elevation of plasma levels of corticosterone (22 2%), progesterone (1 7-fold) and IL-2 (2 6-fold) Our data suggest that chronic exercise is able to counterbalance the effects of CIA on cells of the IS. reinforcing the proposal that the benefits of exercise may not be restricted to aerobic capacity and/or strength improvement Copyright (C) 2010 John Wiley & Sons, Ltd

Fatigue in Brazilian cancer patients, caregivers, and nursing students: a psychometric validation study of the Piper Fatigue Scale-Revised

The objective of this study was to validate the Piper Fatigue Scale-Revised (PFS-R) for use in Brazilian culture. Translation of the PFS-R into Portuguese and validity and reliability tests were performed. Convenience samples in Brazil we as follows: 584 cancer patients (mean age 57 +/- 13 years; 51.3% female); 184 caregivers (mean age 50 +/- 12.7 years; 65.8% female); and 189 undergraduate nursing students (mean age 21.6 +/- 2.8 years; 96.2% female); Instruments used were as follows: Brazilian PFS, Beck Depression Inventory (BDI), and Karnofsky Performance Scale (KPS). The 22 items of the Brazilian PFS loaded well (factor loading > 0.35) on three dimensions identified by factor analysis (behavioral, affective, and sensorial-psychological). These dimensions explained 65% of the variance. Internal consistency reliability was very good (Cronbach`s alpha ranged from 0.841 to 0.943 for the total scale and its dimensions). Cancer patients and their caregivers completed the Brazilian PFS twice for test-retest reliability and results showed good stability (Pearson`s r a parts per thousand yenaEuro parts per thousand 0,60, p < 0,001). Correlations among the Brazilian PFS and other scales were significant, in hypothesized directions, and mostly moderate contributing to divergent (Brazilian PFS x KPS) and convergent validity (Brazilian PFS x BDI). Mild, moderate, and severe fatigue in patients were reported by 73 (12.5%), 167 (28.6%), and 83 (14.2%), respectively. Surprisingly, students had the highest mean total fatigue scores; no significant differences were observed between patients and caregivers showing poor discriminant validity. While the Brazilian PFS is a reliable and valid instrument to measure fatigue in Brazilian cancer patients, further work is needed to evaluate the discriminant validity of the scale in Brazil.

Evaluation of biologic occupational risk control practices: Quality indicators development and validation

Background: There is growing demand for the adoption of qualification systems for health care practices. This study is aimed at describing the development and validation of indicators for evaluation of biologic occupational risk control programs. Methods: The study involved 3 stages: (1) setting up a research team, (2) development of indicators, and (3) validation of the indicators by a team of specialists recruited to validate each attribute of the developed indicators. The content validation method was used for the validation, and a psychometric scale was developed for the specialists` assessment. A consensus technique was used, and every attribute that obtained a Content Validity Index of at least 0.75 was approved. Results: Eight indicators were developed for the evaluation of the biologic occupational risk prevention program, with emphasis on accidents caused by sharp instruments and occupational tuberculosis prevention. The indicators included evaluation of the structure, process, and results at the prevention and biologic risk control levels. The majority of indicators achieved a favorable consensus regarding all validated attributes. Conclusion: The developed indicators were considered validated, and the method used for construction and validation proved to be effective.

Experimental chronic low-frequency resistance training produces skeletal muscle hypertrophy in the absence of muscle damage and metabolic stress markers

Volitional animal resistance training constitutes an important approach to modeling human resistance training. However, the lack of standardization protocol poses a frequent impediment to the production of skeletal muscle hypertrophy and the study of related physiological variables (i.e., cellular damage/inflammation or metabolic stress). Therefore, the purposes of the present study were: (1) to test whether a long-term and low frequency experimental resistance training program is capable of producing absolute increases in muscle mass; (2) to examine whether cellular damage/inflammation or metabolic stress is involved in the process of hypertrophy. In order to test this hypothesis, animals were assigned to a sedentary control (C, n = 8) or a resistance trained group (RT, n = 7). Trained rats performed 2 exercise sessions per week (16 repetitions per day) during 12 weeks. Our results demonstrated that the resistance training strategy employed was capable of producing absolute mass gain in both soleus and plantaris muscles (12%, p<0.05). Furthermore, muscle tumor necrosis factor (TNF-alpha) protein expression (soleus muscle) was reduced by 24% (p<0.01) in trained group when compared to sedentary one. Finally, serum creatine kinase (CK) activity and serum lactate concentrations were not affected in either group. Such information may have practical applications if reproduced in situations where skeletal muscle hypertrophy is desired but high mechanical stimuli of skeletal muscle and inflammation are not. Copyright (C) 2010 John Wiley & Sons, Ltd.

Electrodeposition of copper on titanium wires: Taguchi experimental design approach

Electrodeposition of thin copper layer was carried out on titanium wires in acidic sulphate bath. The influence of titanium surface preparation, cathodic current density, copper sulphate and sulphuric acid concentrations, electrical charge density and stirring of the solution on the adhesion of the electrodeposits was studied using the Taguchi statistical method. A L(16) orthogonal array with the six factors of control at two levels each and three interactions was employed. The analysis of variance of the mean adhesion response and signal-to-noise ratio showed the great influence of cathodic current density on adhesion. on the contrary, the other factors as well as the three investigated interactions revealed low or no significant effect. From this study optimized electrolysis conditions were defined. The copper electrocoating improved the electrical conductivity of the titanium wire. This shows that copper electrocoated titanium wires could be employed for both electrical purpose and mechanical reinforcement in superconducting magnets. (C) 2008 Elsevier B.V. All rights reserved.

The cultural adaptation and validation of the ""Burn Specific Health Scale-Revised"" (BSHS-R): Version for Brazilian burn victims

Background: The Burns Specific Health Scale-Revised (BSHS-R) is of easy application, can be self-administered, and it is considered a good scale to evaluate various important life aspects of burn victims. Objectives: To translate and culturally adapt the BSHS-R into the Brazilian-Portuguese language and to evaluate the internal consistency and convergent validity of the translated BSHS-R. Methods: The cultural adaptation of the BSHS-R included translation and back-translation, discussions with professionals and patients to ensure conceptual equivalence, semantic evaluation, and pre-test of the instrument. The Final Brazilian-Portuguese Version (FBPV) of the BSHS-R was tested on a group of 115 burn patients for internal consistency and validity of construct (using the Rosenberg Self-Esteem Scale (RSES) and the Beck Depression Inventory (BDI)). Results: All values of Cronbach`s alpha were greater than. 8, demonstrating that the internal consistency of the FBPV was very high. Self-esteem was highly correlated with affect and body image (r = .59, p < .001), and with interpersonal relationships (T = .51, p < .001). Correlations between the domains of the FBPV and the BDI were all negative but larger in magnitude than the correlations with RSES. Depression was highly correlated with affect and body image (r = -77, p < .001), and with interpersonal relationships (r = -67, p < .001). Conclusions: The results showed that the adapted version of the BSHS-R into Brazilian-Portuguese fulfills the validity and reliability criteria required from an instrument of health status assessment for burn patients. (C) 2008 Elsevier Ltd and ISBI. All rights reserved.

Identification and Validation of Brazilian Chronic Pain Descriptors

Appropriate pain assessment is very important for managing chronic pain. Given the cultural differences in verbally expressing pain and in psychosocial problems, specific tools are needed. The goal of this study was to identify and validate Brazilian pain descriptors. A purposive sample of health professionals and chronic pain patients was recruited. Four studies were conducted using direct and indirect psychophysical methods: category estimation, magnitude estimation, and magnitude estimation and tine-length. Results showed the descriptors which best describe chronic pain in Brazilian culture and demonstrated that there is not a significant correlation between patients and health professionals and that the psychophysical scale of judgment of pain descriptors is valid, stable, and consistent. Results reinforced that the translations of word descriptors and research tools into another language may be inappropriate, owing to differences in perception and communication and the inadequacy of exact translations to reflect the intended meaning. Given the complexity of the chronic pain, personal suffering involved, and the need for accurate assessment of chronic pain using descriptors stemming from Brazilian culture and language, it is essential to investigate the most adequate words to describe chronic pain. Although it requires more refinement, the Brazilian chronic pain descriptors can be used further to develop a multidimensional pain assessment tool that is culturally sensitive. (C) 2009 by the American Society for Pain Management Nursing