Characterizing and modeling preferential flow using magnetic resonance imaging and multifractal theory.

bitstream/item/89956/1/Proci-07.00331.PD

Uso de ressonância magnética nuclear na caracterização de ácidos húmicos de solos sob diferentes manejos.

bitstream/CNPDIA/9813/1/PA08_96.pd

High-content screen in human pluripotent cells identifies miRNA-regulated pathways controlling pluripotency and differentiation

Background: By post-transcriptionally regulating multiple target transcripts, microRNAs (miRNAs or miR) play important biological functions. H1 embryonic stem cells (hESCs) and NTera-2 embryonal carcinoma cells (ECCs) are two of the most widely used human pluripotent model cell lines, sharing several characteristics, including the expression of miRNAs associated to the pluripotent state or with differentiation. However, how each of these miRNAs functionally impacts the biological properties of these cells has not been systematically evaluated. Methods: We investigated the effects of 31 miRNAs on NTera-2 and H1 hESCs, by transfecting miRNA mimics. Following 3-4 days of culture, cells were stained for the pluripotency marker OCT4 and the G2 cell-cycle marker Cyclin B1, and nuclei and cytoplasm were co-stained with Hoechst and Cell Mask Blue, respectively. By using automated quantitative fluorescence microscopy (i.e., high-content screening (HCS)), we obtained several morphological and marker intensity measurements, in both cell compartments, allowing the generation of a multiparametric miR-induced phenotypic profile describing changes related to proliferation, cell cycle, pluripotency, and differentiation. Results: Despite the overall similarities between both cell types, some miRNAs elicited cell-specific effects, while some related miRNAs induced contrasting effects in the same cell. By identifying transcripts predicted to be commonly targeted by miRNAs inducing similar effects (profiles grouped by hierarchical clustering), we were able to uncover potentially modulated signaling pathways and biological processes, likely mediating the effects of the microRNAs on the distinct groups identified. Specifically, we show that miR-363 contributes to pluripotency maintenance, at least in part, by targeting NOTCH1 and PSEN1 and inhibiting Notch-induced differentiation, a mechanism that could be implicated in na\uefve and primed pluripotent states. Conclusions: We present the first multiparametric high-content microRNA functional screening in human pluripotent cells. Integration of this type of data with similar data obtained from siRNA screenings (using the same HCS assay) could provide a large-scale functional approach to identify and validate microRNA-mediated regulatory mechanisms controlling pluripotency and differentiation

Principal-Oscillation-Pattern Analysis of Gene Expression

Principal-oscillation-pattern (POP) analysis is a multivariate and systematic technique for identifying the dynamic characteristics of a system from time-series data. In this study, we demonstrate the first application of POP analysis to genome-wide time-series gene-expression data. We use POP analysis to infer oscillation patterns in gene expression. Typically, a genomic system matrix cannot be directly estimated because the number of genes is usually much larger than the number of time points in a genomic study. Thus, we first identify the POPs of the eigen-genomic system that consists of the first few significant eigengenes obtained by singular value decomposition. By using the linear relationship between eigengenes and genes, we then infer the POPs of the genes. Both simulation data and real-world data are used in this study to demonstrate the applicability of POP analysis to genomic data. We show that POP analysis not only compares favorably with experiments and existing computational methods, but that it also provides complementary information relative to other approaches

Decreased CD90 expression in human mesenchymal stem cells by applying mechanical stimulation

BACKGROUND: Mesenchymal stem cells (MSC) are multipotent cells which can differentiate along osteogenic, chondrogenic, and adipogenic lineages. The present study was designed to investigate the influence of mechanical force as a specific physiological stress on the differentiation of (MSC) to osteoblast-like cells. METHODS: Human MSC were cultured in osteoinductive medium with or without cyclic uniaxial mechanical stimulation (2000 μstrain, 200 cycles per day, 1 Hz). Cultured cells were analysed for expression of collagen type I, osteocalcin, osteonectin, and CD90. To evaluate the biomineral formation the content of bound calcium in the cultures was determined. RESULTS: After 14 days in culture immunfluorescence staining revealed enhancement of collagen type I and osteonectin expression in response to mechanical stimulation. In contrast, mechanically stimulated cultures stained negative for CD90. In stimulated and unstimulated cultures an increase in the calcium content over time was observed. After 21 days in culture the calcium content in mechanical stimulated cultures was significantly higher compared to unstimulated control cultures. CONCLUSION: These results demonstrate the influence of mechanical force on the differentiation of human MSC into osteoblast-like cells in vitro. While significant enhancement of the biomineral formation by mechanical stimulation is not detected before 21 days, effects on the extracellular matrix became already obvious after 14 days. The decrease of CD90 expression in mechanically stimulated cultures compared to unstimulated control cultures suggests that CD90 is only transiently expressed expression during the differentiation of MSC to osteoblast-like cells in culture

New Mutations in Chronic Lymphocytic Leukemia Identified by Target Enrichment and Deep Sequencing

Chronic lymphocytic leukemia (CLL) is a heterogeneous disease without a well-defined genetic alteration responsible for the onset of the disease. Several lines of evidence coincide in identifying stimulatory and growth signals delivered by B-cell receptor (BCR), and co-receptors together with NFkB pathway, as being the driving force in B-cell survival in CLL. However, the molecular mechanism responsible for this activation has not been identified. Based on the hypothesis that BCR activation may depend on somatic mutations of the BCR and related pathways we have performed a complete mutational screening of 301 selected genes associated with BCR signaling and related pathways using massive parallel sequencing technology in 10 CLL cases. Four mutated genes in coding regions (KRAS, SMARCA2, NFKBIE and PRKD3) have been confirmed by capillary sequencing. In conclusion, this study identifies new genes mutated in CLL, all of them in cases with progressive disease, and demonstrates that next-generation sequencing technologies applied to selected genes or pathways of interest are powerful tools for identifying novel mutational changes

DIGE Proteome Analysis Reveals Suitability of Ischemic Cardiac In Vitro Model for Studying Cellular Response to Acute Ischemia and Regeneration

Proteomic analysis of myocardial tissue from patient population is suited to yield insights into cellular and molecular mechanisms taking place in cardiovascular diseases. However, it has been limited by small sized biopsies and complicated by high variances between patients. Therefore, there is a high demand for suitable model systems with the capability to simulate ischemic and cardiotoxic effects in vitro, under defined conditions. In this context, we established an in vitro ischemia/reperfusion cardiac disease model based on the contractile HL-1 cell line. To identify pathways involved in the cellular alterations induced by ischemia and thereby defining disease-specific biomarkers and potential target structures for new drug candidates we used fluorescence 2D-difference gel electrophoresis. By comparing spot density changes in ischemic and reperfusion samples we detected several protein spots that were differentially abundant. Using MALDI-TOF/TOF-MS and ESI-MS the proteins were identified and subsequently grouped by functionality. Most prominent were changes in apoptosis signalling, cell structure and energy-metabolism. Alterations were confirmed by analysis of human biopsies from patients with ischemic cardiomyopathy

C-13 NMR and conductivity measurements of ethylene oxide-epychloridrine copolymer doped with LiClO4

The ethylene oxide-epychloridrine copolymer has interesting properties as an ionic conducting polymer, principally due to its low glass transition temperature (T-g = -40 degrees C). The effect of dopant concentration (LiClO4) was studied by C-13 NMR spectra and conductivity measurements. These studies allowed us to identify the different carbon atoms in the copolymer and their interaction with lithium cations and solvent molecules. The analysis of C-13 spectra and conductivity measurements show that the solvent participates of the solid electrolyte, modifying the charge transport process by the change in the diffusion coefficient.854173021922

NMR and conductivity studies of ethylene oxide epichloridrine copolymer doped with LiClO4

The copolymer ethylene oxide and epichloridrine doped with lithium perchlorate shows interesting properties as an ionic conducting polymer mainly because of its amorphous structure at room temperature and its low glass transition temperature (T-g = -40 degrees C). C-13, Li-7 and H-1 nuclear magnetic resonance studies were performed in order to analyze the interactions between the polymer matrix and the dopant salt with solvent molecules. These studies and conductivity measurements were used to investigate the influence of the preparation method and the properties of the solvent on the physicochemical characteristics of the polymer electrolyte. Experimental results show that solvent molecules are retained in the solid polymer electrolyte, strongly modifying its ion transport dynamic, This fact can be related to the increase in the ionic conductivity when solvent molecules rest inside the polymer matrix, producing a plasticizing effect.101183469347