Comparison of the glomerular filtration rate in children by the new revised Schwartz formula and a new generalized formula.

The most widely used formula for estimating glomerular filtration rate (eGFR) in children is the Schwartz formula. It was revised in 2009 using iohexol clearances with measured GFR (mGFR) ranging between 15 and 75 ml/min × 1.73 m(2). Here we assessed the accuracy of the Schwartz formula using the inulin clearance (iGFR) method to evaluate its accuracy for children with less renal impairment comparing 551 iGFRs of 392 children with their Schwartz eGFRs. Serum creatinine was measured using the compensated Jaffe method. In order to find the best relationship between iGFR and eGFR, a linear quadratic regression model was fitted and a more accurate formula was derived. This quadratic formula was: 0.68 × (Height (cm)/serum creatinine (mg/dl))-0.0008 × (height (cm)/serum creatinine (mg/dl))(2)+0.48 × age (years)-(21.53 in males or 25.68 in females). This formula was validated using a split-half cross-validation technique and also externally validated with a new cohort of 127 children. Results show that the Schwartz formula is accurate until a height (Ht)/serum creatinine value of 251, corresponding to an iGFR of 103 ml/min × 1.73 m(2), but significantly unreliable for higher values. For an accuracy of 20 percent, the quadratic formula was significantly better than the Schwartz formula for all patients and for patients with a Ht/serum creatinine of 251 or greater. Thus, the new quadratic formula could replace the revised Schwartz formula, which is accurate for children with moderate renal failure but not for those with less renal impairment or hyperfiltration.

A comparison of methods for differential expression analysis of RNA-seq data.

BACKGROUND: Finding genes that are differentially expressed between conditions is an integral part of understanding the molecular basis of phenotypic variation. In the past decades, DNA microarrays have been used extensively to quantify the abundance of mRNA corresponding to different genes, and more recently high-throughput sequencing of cDNA (RNA-seq) has emerged as a powerful competitor. As the cost of sequencing decreases, it is conceivable that the use of RNA-seq for differential expression analysis will increase rapidly. To exploit the possibilities and address the challenges posed by this relatively new type of data, a number of software packages have been developed especially for differential expression analysis of RNA-seq data. RESULTS: We conducted an extensive comparison of eleven methods for differential expression analysis of RNA-seq data. All methods are freely available within the R framework and take as input a matrix of counts, i.e. the number of reads mapping to each genomic feature of interest in each of a number of samples. We evaluate the methods based on both simulated data and real RNA-seq data. CONCLUSIONS: Very small sample sizes, which are still common in RNA-seq experiments, impose problems for all evaluated methods and any results obtained under such conditions should be interpreted with caution. For larger sample sizes, the methods combining a variance-stabilizing transformation with the 'limma' method for differential expression analysis perform well under many different conditions, as does the nonparametric SAMseq method.

Rate and processes of river network rearrangement during incipient faulting: The case of the Cahabon river, Guatemala

Deeply incised river networks are generally regarded as robust features that are not easily modified by erosion or tectonics. Although the reorganization of deeply incised drainage systems has been documented, the corresponding importance with regard to the overall landscape evolution of mountain ranges and the factors that permit such reorganizations are poorly understood. To address this problem, we have explored the rapid drainage reorganization that affected the Cahabon River in Guatemala during the Quaternary. Sediment-provenance analysis, field mapping, and electrical resistivity tomography (ERT) imaging are used to reconstruct the geometry of the valley before the river was captured. Dating of the abandoned valley sediments by the Be-10-Al-26 burial method and geomagnetic polarity analysis allow us to determine the age of the capture events and then to quantify several processes, such as the rate of tectonic deformation of the paleovalley, the rate of propagation of post-capture drainage reversal, and the rate at which canyons that formed at the capture sites have propagated along the paleovalley. Transtensional faulting started 1 to 3 million years ago, produced ground tilting and ground faulting along the Cahabon River, and thus generated differential uplift rate of 0.3 +/- 0.1 up to 0.7 +/- 0.4 mm . y(-1) along the river's course. The river responded to faulting by incising the areas of relative uplift and depositing a few tens of meters of sediment above the areas of relative subsidence. Then, the river experienced two captures and one avulsion between 700 ky and 100 ky. The captures breached high-standing ridges that separate the Cahabon River from its captors. Captures occurred at specific points where ridges are made permeable by fault damage zones and/or soluble rocks. Groundwater flow from the Cahabon River down to its captors likely increased the erosive power of the captors thus promoting focused erosion of the ridges. Valley-fill formation and capture occurred in close temporal succession, suggesting a genetic link between the two. We suggest that the aquifers accumulated within the valley-fills, increased the head along the subterraneous system connecting the Cahabon River to its captors, and promoted their development. Upon capture, the breached valley experienced widespread drainage reversal toward the capture sites. We attribute the generalized reversal to combined effects of groundwater sapping in the valley-fill, axial drainage obstruction by lateral fans, and tectonic tilting. Drainage reversal increased the size of the captured areas by a factor of 4 to 6. At the capture sites, 500 m deep canyons have been incised into the bedrock and are propagating upstream at a rate of 3 to 11 mm . y(-1) deepening at a rate of 0.7 to 1 5 mm . y(-1). At this rate, 1 to 2 million years will be necessary for headward erosion to completely erase the topographic expression of the paleovalley. It is concluded that the rapid reorganization of this drainage system was made possible by the way the river adjusted to the new tectonic strain field, which involved transient sedimentation along the river's course. If the river had escaped its early reorganization and had been given the time necessary to reach a new dynamic equilibrium, then the transient conditions that promoted capture would have vanished and its vulnerability to capture would have been strongly reduced.

High-throughput SELEX SAGE method for quantitative modeling of transcription-factor binding sites.

The ability to determine the location and relative strength of all transcription-factor binding sites in a genome is important both for a comprehensive understanding of gene regulation and for effective promoter engineering in biotechnological applications. Here we present a bioinformatically driven experimental method to accurately define the DNA-binding sequence specificity of transcription factors. A generalized profile was used as a predictive quantitative model for binding sites, and its parameters were estimated from in vitro-selected ligands using standard hidden Markov model training algorithms. Computer simulations showed that several thousand low- to medium-affinity sequences are required to generate a profile of desired accuracy. To produce data on this scale, we applied high-throughput genomics methods to the biochemical problem addressed here. A method combining systematic evolution of ligands by exponential enrichment (SELEX) and serial analysis of gene expression (SAGE) protocols was coupled to an automated quality-controlled sequence extraction procedure based on Phred quality scores. This allowed the sequencing of a database of more than 10,000 potential DNA ligands for the CTF/NFI transcription factor. The resulting binding-site model defines the sequence specificity of this protein with a high degree of accuracy not achieved earlier and thereby makes it possible to identify previously unknown regulatory sequences in genomic DNA. A covariance analysis of the selected sites revealed non-independent base preferences at different nucleotide positions, providing insight into the binding mechanism.

Termination of major ductile strike-slip shear and differential cooling along the Insubric line (Central Alps): U-Pb, Rb-Sr and 40Ar/39Ar ages of cross-cutting pegmatites

To constrain the age of strike-slip shear, related granitic magmatism, and cooling along the Insubric line, 29 size fractions of monazite and xenotime were dated by the U-Pb method, and a series of 25 Rb-Sr and Ar-40/Ar-39 ages were measured on different size fractions of muscovite and biotite. The three pegmatitic intrusions analyzed truncate high-grade metamorphic mylonite gneisses of the Simplon shear zone, a major Alpine structure produced in association with dextral strike-slip movements along the southern edge of the European plate, after collision with its Adriatic indenter. Pegmatites and aplites were produced between 29 and 25 Ma in direct relation to right-lateral shear along the Insubric line, by melting of continental crust having Sr-87/Sr-86 between 0.7199 and 0.7244 at the time of melting. High-temperature dextral strike-slip shear was active at 29.2 +/- 0.2 (2 sigma) Ma, and it terminated before 26.4 +/- 0.1 Ma. During dike injection, temperatures in the country rocks of the Isorno-Orselina and Monte Rosa structural units did not exceed approximate to 500 degrees C, leading to fast initial cooling, followed by slower cooling to approximate to 350 degrees C within several million years. In one case, initial cooling to approximate to 500 degrees C was significantly delayed by about 4 m.y., with final cooling to approximate to 300 degrees C at 20-19 Ma in all units. For the period between 29 and 19 Ma, cooling of the three sample localities was non-uniform in space and time, with significant variations on the kilometre scale. These differences are most likely due to strongly varying heat flow, and/or heterogeneous distribution of unroofing rates within the continuously deforming Insubric line. If entirely ascribed to differences in unroofing, corresponding rates would vary between 0.5 and 2.5 mm/y, for a thermal gradient of 30 degrees/km.

Differential protein labeling with thiol-reactive infrared DY-680 and DY-780 maleimides and analysis by two-dimensional gel electrophoresis.

Differential protein labeling with 2-DE separation is an effective method for distinguishing differences in the protein composition of two or more protein samples. Here, we report on a sensitive infrared-based labeling procedure, adding a novel tool to the many labeling possibilities. Defined amounts of newborn and adult mouse brain proteins and tubulin were exposed to maleimide-conjugated infrared dyes DY-680 and DY-780 followed by 1- and 2-DE. The procedure allows amounts of less than 5 microg of cysteine-labeled protein mixtures to be detected (together with unlabeled proteins) in a single 2-DE step with an LOD of individual proteins in the femtogram range; however, co-migration of unlabeled proteins and subsequent general protein stains are necessary for a precise comparison. Nevertheless, the most abundant thiol-labeled proteins, such as tubulin, were identified by MS, with cysteine-containing peptides influencing the accuracy of the identification score. Unfortunately, some infrared-labeled proteins were no longer detectable by Western blots. In conclusion, differential thiol labeling with infrared dyes provides an additional tool for detection of low-abundant cysteine-containing proteins and for rapid identification of differences in the protein composition of two sets of protein samples.

A treatment planning method for sequentially combining radiopharmaceutical therapy and external radiation therapy.

PURPOSE: Effective cancer treatment generally requires combination therapy. The combination of external beam therapy (XRT) with radiopharmaceutical therapy (RPT) requires accurate three-dimensional dose calculations to avoid toxicity and evaluate efficacy. We have developed and tested a treatment planning method, using the patient-specific three-dimensional dosimetry package 3D-RD, for sequentially combined RPT/XRT therapy designed to limit toxicity to organs at risk. METHODS AND MATERIALS: The biologic effective dose (BED) was used to translate voxelized RPT absorbed dose (D(RPT)) values into a normalized total dose (or equivalent 2-Gy-fraction XRT absorbed dose), NTD(RPT) map. The BED was calculated numerically using an algorithmic approach, which enabled a more accurate calculation of BED and NTD(RPT). A treatment plan from the combined Samarium-153 and external beam was designed that would deliver a tumoricidal dose while delivering no more than 50 Gy of NTD(sum) to the spinal cord of a patient with a paraspinal tumor. RESULTS: The average voxel NTD(RPT) to tumor from RPT was 22.6 Gy (range, 1-85 Gy); the maximum spinal cord voxel NTD(RPT) from RPT was 6.8 Gy. The combined therapy NTD(sum) to tumor was 71.5 Gy (range, 40-135 Gy) for a maximum voxel spinal cord NTD(sum) equal to the maximum tolerated dose of 50 Gy. CONCLUSIONS: A method that enables real-time treatment planning of combined RPT-XRT has been developed. By implementing a more generalized conversion between the dose values from the two modalities and an activity-based treatment of partial volume effects, the reliability of combination therapy treatment planning has been expanded.

Study of OSEM with different subsets in grating-based X-ray differential phase-contrast imaging.

Impressive developments in X-ray imaging are associated with X-ray phase contrast computed tomography based on grating interferometry, a technique that provides increased contrast compared with conventional absorption-based imaging. A new "single-step" method capable of separating phase information from other contributions has been recently proposed. This approach not only simplifies data-acquisition procedures, but, compared with the existing phase step approach, significantly reduces the dose delivered to a sample. However, the image reconstruction procedure is more demanding than for traditional methods and new algorithms have to be developed to take advantage of the "single-step" method. In the work discussed in this paper, a fast iterative image reconstruction method named OSEM (ordered subsets expectation maximization) was applied to experimental data to evaluate its performance and range of applicability. The OSEM algorithm with different subsets was also characterized by comparison of reconstruction image quality and convergence speed. Computer simulations and experimental results confirm the reliability of this new algorithm for phase-contrast computed tomography applications. Compared with the traditional filtered back projection algorithm, in particular in the presence of a noisy acquisition, it furnishes better images at a higher spatial resolution and with lower noise. We emphasize that the method is highly compatible with future X-ray phase contrast imaging clinical applications.

Detection of metabolite induction in fungal co-cultures on solid media by high-throughput differential ultra-high pressure liquid chromatography-time-of-flight mass spectrometry fingerprinting.

Access to new biological sources is a key element of natural product research. A particularly large number of biologically active molecules have been found to originate from microorganisms. Very recently, the use of fungal co-culture to activate the silent genes involved in metabolite biosynthesis was found to be a successful method for the induction of new compounds. However, the detection and identification of the induced metabolites in the confrontation zone where fungi interact remain very challenging. To tackle this issue, a high-throughput UHPLC-TOF-MS-based metabolomic approach has been developed for the screening of fungal co-cultures in solid media at the petri dish level. The metabolites that were overexpressed because of fungal interactions were highlighted by comparing the LC-MS data obtained from the co-cultures and their corresponding mono-cultures. This comparison was achieved by subjecting automatically generated peak lists to statistical treatments. This strategy has been applied to more than 600 co-culture experiments that mainly involved fungal strains from the Fusarium genera, although experiments were also completed with a selection of several other filamentous fungi. This strategy was found to provide satisfactory repeatability and was used to detect the biomarkers of fungal induction in a large panel of filamentous fungi. This study demonstrates that co-culture results in consistent induction of potentially new metabolites.

Screening for wound-induced oxylipins in Arabidopsis thaliana by differential HPLC-APCI/MS profiling of crude leaf extracts and subsequent characterisation by capillary-scale NMR

A simple non-targeted differential HPLC-APCI/MS approach has been developed in order to survey metabolome modifications that occur in the leaves of Arabidopsis thaliana following wound-induced stress. The wound-induced accumulation of metabolites, particularly oxylipins, was evaluated by HPLC-MS analysis of crude leaf extracts. A generic, rapid and reproducible pressure liquid extraction procedure was developed for the analysis of restricted leaf samples without the need for specific sample preparation. The presence of various oxylipins was determined by head-to-head comparison of the HPLC-MS data, filtered with a component detection algorithm, and automatically compared with the aid of software searching for small differences in similar HPLC-MS profiles. Repeatability was verified in several specimens belonging to different series. Wound-inducible jasmonates were efficiently highlighted by this non-targeted approach without the need for complex sample preparation as is the case for the 'oxylipin signature' procedure based on GC-MS. Furthermore this HPLC-MS screening technique allowed the isolation of induced compounds for further characterisation by capillary-scale NMR (CapNMR) after HPLC scale-up. In this paper, the screening method is described and applied to illustrate its potential for monitoring polar and non-polar stress-induced constituents as well as its use in combination with CapNMR for the structural assignment of wound-induced compounds of interest

Investigation of discrete imaging models and iterative image reconstruction in differential X-ray phase-contrast tomography.

Differential X-ray phase-contrast tomography (DPCT) refers to a class of promising methods for reconstructing the X-ray refractive index distribution of materials that present weak X-ray absorption contrast. The tomographic projection data in DPCT, from which an estimate of the refractive index distribution is reconstructed, correspond to one-dimensional (1D) derivatives of the two-dimensional (2D) Radon transform of the refractive index distribution. There is an important need for the development of iterative image reconstruction methods for DPCT that can yield useful images from few-view projection data, thereby mitigating the long data-acquisition times and large radiation doses associated with use of analytic reconstruction methods. In this work, we analyze the numerical and statistical properties of two classes of discrete imaging models that form the basis for iterative image reconstruction in DPCT. We also investigate the use of one of the models with a modern image reconstruction algorithm for performing few-view image reconstruction of a tissue specimen.