Proteomic and transcriptomic profiling of Staphylococcus aureus surface LPXTG-proteins: correlation with agr genotypes and adherence phenotypes.

Staphylococcus aureus infections involve numerous adhesins and toxins, which expression depends on complex regulatory networks. Adhesins include a family of surface proteins covalently attached to the peptidoglycan via a conserved LPXTG motif. Here we determined the protein and mRNA expression of LPXTG-proteins of S. aureus Newman in time-course experiments, and their relation to fibrinogen adherence in vitro. Experiments were performed with mutants in the global accessory-gene regulator (agr), surface protein A (Spa), and fibrinogen-binding protein A (ClfA), as well as during growth in iron-rich or iron-poor media. Surface proteins were recovered by trypsin-shaving of live bacteria. Released peptides were analyzed by liquid chromatography coupled to tandem mass-spectrometry. To unambiguously identify peptides unique to LPXTG-proteins, the analytical conditions were refined using a reference library of S. aureus LPXTG-proteins heterogeneously expressed in surrogate Lactococcus lactis. Transcriptomes were determined by microarrays. Sixteen of the 18 LPXTG-proteins present in S. aureus Newman were detected by proteomics. Nine LPXTG-proteins showed a bell-shape agr-like expression that was abrogated in agr-negative mutants including Spa, fibronectin-binding protein A (FnBPA), ClfA, iron-binding IsdA, and IsdB, immunomodulator SasH, functionally uncharacterized SasD, biofilm-related SasG and methicillin resistance-related FmtB. However, only Spa and SasH modified their proteomic and mRNA profiles in parallel in the parent and its agr- mutant, whereas all other LPXTG-proteins modified their proteomic profiles independently of their mRNA. Moreover, ClfA became highly transcribed and active in fibrinogen-adherence tests during late growth (24 h), whereas it remained poorly detected by proteomics. On the other hand, iron-regulated IsdA-B-C increased their protein expression by >10-times in iron-poor conditions. Thus, proteomic, transcriptomic, and adherence-phenotype demonstrated differential profiles in S. aureus. Moreover, trypsin peptide signatures suggested differential protein domain exposures in various environments, which might be relevant for anti-adhesin vaccines. A comprehensive understanding of the S. aureus physiology should integrate all three approaches.

Correlation of Locally-Based Performance of Asphalts with Their Physicochemical Parameters, Progress Report 2, HR-298, 1989

This project was undertaken to study the relationships between the performance of locally available asphalts and their physicochemical properties under Iowa conditions with the ultimate objective of development of a locally and performance-based asphalt specification for durable pavements. Physical and physicochemical tests were performed on three sets of asphalt samples including: (a) twelve samples from local asphalt suppliers and their TFOT residues, (b) six core samples of known service records, and (c) a total of 79 asphalts from 10 pavement projects including original, lab aged and recovered asphalts from field mixes, as well as from lab aged mixes. Tests included standard rheological tests, HP-GPC and TMA. Some specific viscoelastic tests (at 5 deg C) were run on b samples and on some a samples. DSC and X-ray diffraction studies were performed on a and b samples. Furthermore, NMR techniques were applied to some a, b and c samples. Efforts were made to identify physicochemical properties which are correlated to physical properties known to affect field performance. The significant physicochemical parameters were used as a basis for an improved performance-based trial specification for Iowa to ensure more durable pavements.

Correlation of physical and chemical attributes of soil with sugarcane yield

The objective of this work was to evaluate the correlation between sugarcane yield and some physical and chemical attributes of soil. For this, a 42‑ha test area in Araras, SP, Brazil, was used. Soil properties were determined from samples collected at the beginning of the 2003/2004 harvest season, using a regular 100x100 m grid. Yield assessment was done with a yield monitor (Simprocana). Correlation analyses were performed between sugarcane yield and the following soil properties: pH, pH CaCl2, N, C, cone index, clay content, soil organic matter, P, K, Ca, Mg, H+AL, cation exchange capacity, and base saturation. Correlation coefficients were respectively ‑0.05, ‑0.29, 0.33, 0.41, ‑0.27, 0.22, 0.44, ‑0.24, trace, ‑0.06, 0.01, 0.32, 0.14, and 0.04. Correlations of chemical and physical attributes of soil with sugarcane yield are weak, and, per se, they are not able to explain sugarcane yield variation, which suggests that other variables, besides soil attributes, should be analysed.

Molecular characterization of a human matrix attachment region epigenetic regulator.

Matrix attachment regions (MAR) generally act as epigenetic regulatory sequences that increase gene expression, and they were proposed to partition chromosomes into loop-forming domains. However, their molecular mode of action remains poorly understood. Here, we assessed the possible contribution of the AT-rich core and adjacent transcription factor binding motifs to the transcription augmenting and anti-silencing effects of human MAR 1-68. Either flanking sequences together with the AT-rich core were required to obtain the full MAR effects. Shortened MAR derivatives retaining full MAR activity were constructed from combinations of the AT-rich sequence and multimerized transcription factor binding motifs, implying that both transcription factors and the AT-rich microsatellite sequence are required to mediate the MAR effect. Genomic analysis indicated that MAR AT-rich cores may be depleted of histones and enriched in RNA polymerase II, providing a molecular interpretation of their chromatin domain insulator and transcriptional augmentation activities.

Free-breathing T2 mapping at 3T for the monitoring of cardiac allograft rejection: initial results

rejection can lead to loss of function. Histological reading of endomyocardial biopsy remains the "gold standard" for guiding immunosuppression, despite its methodological limitations (sampling error and interobserver variability). The measurement of the T2 relaxation time has been suggested for detection of allograft rejection, on the pathophysiological basis that the T2 relaxation time prolongs with local edema resulting from acute allograft rejection. Using breath-held cardiac magnetic resonance T2 mapping at 1.5 T, Usman et al. (CircCardiovascImaging2012) detected moderate allograft rejection (grade 2R, ISHLT 2004). With modern immunosuppression grade 2R rejection has become a rare event, but the need remains for a technique that permits the discrimination of absent (grade 0R) and mild rejection (grade 1R). We therefore investigated whether an increase of magnetic field strength to 3T and the use of real-time navigator-gated respiration compensation allow for an increase in the sensitivity of T2 relaxation time detection that is necessary to achieve this discrimination. Methods: Eighteen patients received EMB (Tan et al., ArchPatholLabMed2007) and cardiac T2 mapping on the same day. Reading of T2 maps was blinded to the histological results. For final analysis, 3 cases with known 2R rejection at the time of T2 mapping were added, yielding 21 T2 mapping sessions. A respiration-navigator-gated radial gradient-recalled-echo pulse sequence (resolution 1.17 mm2, matrix 2562, trigger time 3 heartbeats, T2 preparation duration TET2 Prep = 60/30/0 ms) was applied to obtain 3 short-axis T2 maps (van Heeswijk et al., JACCCardiovascImaging2012), which were segmented according to AHA guidelines (Cerqueira et al, Circulation2001). The highest segmental T2 values were grouped according to histological rejection grade and differences were analyzed by Student's t-test, except for the non-blinded cases with 2R rejection. The degree of discrimination was determined using the Spearman's ranked correlation test. Results: The high-quality T2 maps allowed for visual differentiation of the rejection degrees (Figure 1), and the correlation of T2 mapping with the histological grade of acute cellular rejection was significant (Spearman's r = 0.56, p = 0.007). The 0R (n = 15) and 1R (n = 3) degrees demonstrated significantly different T2 values (46.9 ± 5.0 and 54.3 ± 3.0 ms, p = 0.02, Figure 2). Cases with 2R rejection showed clear T2 elevation (T2 = 60.3 ± 16.2 ms). Conclusions: This pilot study demonstrates that non-invasive free-breathing cardiac T2 mapping at 3T discriminates between no and mild cardiac allograft rejection. Confirmation of these encouraging results in a larger cohort should consider a study able to show equivalency or superiority of T2 mapping.

Selection of the most informative morphoagronomic descriptors for cassava germplasm

The objective of this work was to select the most informative morphoagronomic descriptors for cassava (Manihot esculenta) germplasm and to evaluate the ability of different methods to select the descriptors. Ninety-five accessions were characterized using 51 morphoagronomic descriptors. Data were subjected to a multiple correspondence analysis (MCA), whose information was used in the following four methods of descriptor selection: reverse order of the descriptor for the pth factorial axis of the MCA (Jolliffe); sequential, multiple correspondence analysis (SMCA); mean of the contribution orders of the descriptor in the first three factorial axes (C3PA); and C3PA method weighted by the respective eigenvalues of the full analysis (C3PAWeig). The correlations between the dissimilarity matrix with all descriptors and the most informative descriptors were high and significant (0.75, 0.77, 0.83, and 0.84 for C3PAWeig, C3PA, SMCA, and Jolliffe, respectively). The less informative descriptors were discarded, considering those common among the selection methods and relevant for the breeding interests. Therefore, 32 morphoagronomic descriptors with correlation between the dissimilarity matrices (r=0.81) were selected, due to their high capacity to discriminate cassava germplasm and to their ability to maintain some preliminary agronomic traits, useful for the initial characterization of the germplasm.

Genome-wide prediction of matrix attachment regions that increase gene expression in mammalian cells.

Gene transfer in eukaryotic cells and organisms suffers from epigenetic effects that result in low or unstable transgene expression and high clonal variability. Use of epigenetic regulators such as matrix attachment regions (MARs) is a promising approach to alleviate such unwanted effects. Dissection of a known MAR allowed the identification of sequence motifs that mediate elevated transgene expression. Bioinformatics analysis implied that these motifs adopt a curved DNA structure that positions nucleosomes and binds specific transcription factors. From these observations, we computed putative MARs from the human genome. Cloning of several predicted MARs indicated that they are much more potent than the previously known element, boosting the expression of recombinant proteins from cultured cells as well as mediating high and sustained expression in mice. Thus we computationally identified potent epigenetic regulators, opening new strategies toward high and stable transgene expression for research, therapeutic production or gene-based therapies.

Covariance Matrix Estimation and Mean-Variance Optimization in application to International Diversification

Tutkimus keskittyy kansainväliseen hajauttamiseen suomalaisen sijoittajan näkökulmasta. Tutkimuksen toinen tavoite on selvittää tehostavatko uudet kovarianssimatriisiestimaattorit minimivarianssiportfolion optimointiprosessia. Tavallisen otoskovarianssimatriisin lisäksi optimoinnissa käytetään kahta kutistusestimaattoria ja joustavaa monimuuttuja-GARCH(1,1)-mallia. Tutkimusaineisto koostuu Dow Jonesin toimialaindekseistä ja OMX-H:n portfolioindeksistä. Kansainvälinen hajautusstrategia on toteutettu käyttäen toimialalähestymistapaa ja portfoliota optimoidaan käyttäen kahtatoista komponenttia. Tutkimusaieisto kattaa vuodet 1996-2005 eli 120 kuukausittaista havaintoa. Muodostettujen portfolioiden suorituskykyä mitataan Sharpen indeksillä. Tutkimustulosten mukaan kansainvälisesti hajautettujen investointien ja kotimaisen portfolion riskikorjattujen tuottojen välillä ei ole tilastollisesti merkitsevää eroa. Myöskään uusien kovarianssimatriisiestimaattoreiden käytöstä ei synnytilastollisesti merkitsevää lisäarvoa verrattuna otoskovarianssimatrisiin perustuvaan portfolion optimointiin.

Colloid cysts of the third ventricle: correlation of MR and CT findings with histology and chemical analysis.

Eight patients with colloid cysts of the third ventricle were examined with CT and MR. In six, surgical resection was performed and the material was subjected to histologic evaluation; the concentrations of trace elements were determined by particle-induced X-ray emission. Stereotaxic aspiration was performed in two. The investigation showed that colloid cysts are often iso- or hypodense relative to brain on CT (5/8), but sometimes have a center of increased density. Increased density did not correlate with increased concentration of calcium or other metals but did not correlate with high cholesterol content. Colloid cysts appear more heterogeneous on MR (6/8) than on CT (3/8), despite a homogeneous appearance at histology. High signal on short TR/TE sequences is correlated with a high cholesterol content. A marked shortening of the T2 relaxation time is often noticed in the central part of the cyst. Analysis of trace elements showed that this phenomenon is not related to the presence of metals with paramagnetic effects. Our analysis of the contents of colloid cysts does not support the theory that differing metallic concentrations are responsible for differences in MR signal intensity or CT density. We did find that increased CT density and high MR signal correlated with high cholesterol content.

Different concentrations of Mg++ ions affect nuclear matrix protein distribution during thermal stabilization of isolated nuclei.

The nuclear matrix, a proteinaceous network believed to be a scaffolding structure determining higher-order organization of chromatin, is usually prepared from intact nuclei by a series of extraction steps. In most cell types investigated the nuclear matrix does not spontaneously resist these treatments but must be stabilized before the application of extracting agents. Incubation of isolated nuclei at 37C or 42C in buffers containing Mg++ has been widely employed as stabilizing agent. We have previously demonstrated that heat treatment induces changes in the distribution of three nuclear scaffold proteins in nuclei prepared in the absence of Mg++ ions. We studied whether different concentrations of Mg++ (2.0-5 mM) affect the spatial distribution of nuclear matrix proteins in nuclei isolated from K562 erythroleukemia cells and stabilized by heat at either 37C or 42C. Five proteins were studied, two of which were RNA metabolism-related proteins (a 105-kD component of splicing complexes and an RNP component), one a 126-kD constituent of a class of nuclear bodies, and two were components of the inner matrix network. The localization of proteins was determined by immunofluorescent staining and confocal scanning laser microscope. Mg++ induced significant changes of antigen distribution even at the lowest concentration employed, and these modifications were enhanced in parallel with increase in the concentration of the divalent cation. The different sensitivity to heat stabilization and Mg++ of these nuclear proteins might reflect a different degree of association with the nuclear scaffold and can be closely related to their functional or structural role.

Bond strength tests of dental adhesive systems and their correlation with clinical results : a meta-analysis

OBJECTIVE: To evaluate the variability of bond strength test results of adhesive systems (AS) and to correlate the results with clinical parameters of clinical studies investigating cervical restorations. MATERIALS AND METHODS: Regarding the clinical studies, the internal database which had previously been used for a meta-analysis on cervical restorations was updated with clinical studies published between 2008 and 2012 by searching the PubMed and SCOPUS databases. PubMed and the International Association for Dental Research abstracts online were searched for laboratory studies on microtensile, macrotensile and macroshear bond strength tests. The inclusion criteria were (1) dentin, (2) testing of at least four adhesive systems, (3) same diameter of composite and (4) 24h of water storage prior to testing. The clinical outcome variables were retention loss, marginal discoloration, detectable margins, and a clinical index comprising the three parameters by weighing them. Linear mixed models which included a random study effect were calculated for both, the laboratory and the clinical studies. The variability was assessed by calculating a ratio of variances, dividing the variance among the estimated bonding effects obtained in the linear mixed models by the sum of all variance components estimated in these models. RESULTS: Thirty-two laboratory studies fulfilled the inclusion criteria comprising 183 experiments. Of those, 86 used the microtensile test evaluating 22 adhesive systems (AS). Twenty-seven used the macrotensile test with 17 AS, and 70 used the macroshear test with 24 AS. For 28 AS the results from clinical studies were available. Microtensile and macrotensile (Spearman rho=0.66, p=0.007) were moderately correlated and also microtensile and macroshear (Spearman rho=0.51, p=0.03) but not macroshear and macrotensile (Spearman rho=0.34, p=0.22). The effect of the adhesive system was significant for microtensile and macroshear (p<0.001) but not for macrotensile. The effect of the adhesive system could explain 36% of the variability of the microtensile test, 27% of the macrotensile and 33% of the macroshear test. For the clinical trials, about 49% of the variability of retained restorations could be explained by the adhesive system. With respect to the correlation between bond strength tests and clinical parameters, only a moderate correlation between micro- and macrotensile test results and marginal discoloration was demonstrated. However, no correlation between these tests and a retention loss or marginal integrity was shown. The correlation improved when more studies were included compared to assessing only one study. SIGNIFICANCE: The high variability of bond strength test results highlights the need to establish individual acceptance levels for a given test institute. The weak correlation of bond-strength test results with clinical parameters leads to the conclusion that one should not rely solely on bond strength tests to predict the clinical performance of an adhesive system but one should conduct other laboratory tests like tests on the marginal adaptation of fillings in extracted teeth and the retention loss of restorations in non-retentive cavities after artificial aging.

Affinity distribution functions in multicomponent heterogeneous adsorption. Analytical inversion of isotherms to obtain affinity spectra

An analytical approach for the interpretation of multicomponent heterogeneous adsorption or complexation isotherms in terms of multidimensional affinity spectra is presented. Fourier transform, applied to analyze the corresponding integral equation, leads to an inversion formula which allows the computation of the multicomponent affinity spectrum underlying a given competitive isotherm. Although a different mathematical methodology is used, this procedure can be seen as the extension to multicomponent systems of the classical Sips’s work devoted to monocomponent systems. Furthermore, a methodology which yields analytical expressions for the main statistical properties (mean free energies of binding and covariance matrix) of multidimensional affinity spectra is reported. Thus, the level of binding correlation between the different components can be quantified. It has to be highlighted that the reported methodology does not require the knowledge of the affinity spectrum to calculate the means, variances, and covariance of the binding energies of the different components. Nonideal competitive consistent adsorption isotherm, widely used in metal/proton competitive complexation to environmental macromolecules, and Frumkin competitive isotherms are selected to illustrate the application of the reported results. Explicit analytical expressions for the affinity spectrum as well as for the matrix correlation are obtained for the NICCA case. © 2004 American Institute of Physics.