Employment of a noninvasive magnetic method for evaluation of gastrointestinal transit in rats

ABSTRACT: AC Biosusceptometry (ACB) was previously employed towards recording gastrointestinal motility. Our data show a reliable and successful evaluation of gastrointestinal transit of liquid and solid meals in rats, considering the methods scarcity and number of experiments needed to endorsement of drugs and medicinal plants. ACB permits real time and simultaneous experiments using the same animal, preserving the physiological conditions employing both meals with simplicity and accuracy.     

A Software Toolkit and Interface for Performing Stable Isotope Labeling and Top3 Quantification Using Progenesis LC-MS

Abstract Numerous software packages exist to provide support for quantifying peptides and proteins from mass spectrometry (MS) data. However, many support only a subset of experimental methods or instrument types, meaning that laboratories often have to use multiple software packages. The Progenesis LC-MS software package from Nonlinear Dynamics is a software solution for label-free quantitation. However, many laboratories using Progenesis also wish to employ stable isotope-based methods that are not natively supported in Progenesis. We have developed a Java programming interface that can use the output files produced by Progenesis, allowing the basic MS features quantified across replicates to be used in a range of different experimental methods. We have developed post-processing software (the Progenesis Post-Processor) to embed Progenesis in the analysis of stable isotope labeling data and top3 pseudo-absolute quantitation. We have also created export ability to the new data standard, mzQuantML, produced by the Proteomics Standards Initiative to facilitate the development and standardization process. The software is provided to users with a simple graphical user interface for accessing the different features. The underlying programming interface may also be used by Java developers to develop other routines for analyzing data produced by Progenesis.     

The mzQuantML Data Standard for Mass Spectrometry–based Quantitative Studies in Proteomics

The range of heterogeneous approaches available for quantifying protein abundance via mass spectrometry (MS)¹ leads to considerable challenges in modeling, archiving, exchanging, or submitting experimental data sets as supplemental material to journals. To date, there has been no widely accepted format for capturing the evidence trail of how quantitative analysis has been performed by software, for transferring data between software packages, or for submitting to public databases. In the context of the Proteomics Standards Initiative, we have developed the mzQuantML data standard. The standard can represent quantitative data about regions in two-dimensional retention time versus mass/charge space (called features), peptides, and proteins and protein groups (where there is ambiguity regarding peptide-to-protein inference), and it offers limited support for small molecule (metabolomic) data. The format has structures for representing replicate MS runs, grouping of replicates (for example, as study variables), and capturing the parameters used by software packages to arrive at these values. The format has the capability to reference other standards such as mzML and mzIdentML, and thus the evidence trail for the MS workflow as a whole can now be described. Several software implementations are available, and we encourage other bioinformatics groups to use mzQuantML as an input, internal, or output format for quantitative software and for structuring local repositories. All project resources are available in the public domain from the HUPO Proteomics Standards Initiative http://www.psidev.info/mzquantml.The Proteomics Standards Initiative (PSI) has been working for ten years to improve the reporting and standardization of proteomics data. The PSI has published minimum reporting guidelines, called MIAPE (Minimum Information about a Proteomics Experiment) documents, for MS-based proteomics (1) and molecular interactions (2), as well as data standards for raw/processed MS data in mzML (3), peptide and protein identifications in mzIdentML (4), transitions for selected reaction monitoring analysis in TraML (5), and molecular interactions in PSI-MI format (6). Standards are particularly important for quantitative proteomics research, because the associated bioinformatics analysis is highly challenging as a result of the range of different experimental techniques for deriving abundance values for proteins using MS. The techniques can be broadly divided into those based on (i) differential labeling, in which a metabolic label or chemical tag is applied to cells, peptides, or proteins, samples are mixed, and intensity signals for peptide ions are compared within single MS runs; or (ii) label-free methods in which MS runs occur in parallel and bioinformatics methods are used to extract intensity signals, ensuring that like-for-like signals are compared between runs (7). In most label-based and label-free approaches, peptide ratios or abundance values must be summarized in order for one to arrive at relative protein abundance values, taking into account ambiguity in peptide-to-protein inference. Absolute protein abundance values can typically be derived only using internal standards spiked into samples of known abundance (8, 9). The PSI has recently developed a MIAPE-Quant document defining and describing the minimal information necessary in order to judge or repeat a quantitative proteomics experiment.Software packages tend to report peptide or protein abundance values in a bespoke format, often as tab or comma separated values, for import into spreadsheet software. In complementary work, the PSI has developed a standard format for capturing these final results in a standardized tab separated value format, called mzTab, suitable for post-processing and visualization in end-user tools such as Microsoft Excel or the R programming language. The final results of a quantitative analysis are sufficient for many purposes, such as performing statistical analysis to determine differential expression or cluster analysis to find co-expressed proteins. However, mzTab (or similar bespoke formats) was not designed to hold a trace of how the peptide and protein abundance values were calculated from MS data (i.e. metadata is lost that might be crucial for other tasks). For example, most quantitative software packages detect and quantify so-called “features” (representing all ions collected for a given peptide) in two-dimensional MS data, where the two dimensions are retention time from liquid chromatography (LC) and mass over charge (m/z). Without capturing the two-dimensional coordinates of the features, it is not possible to write visualization software showing exactly what the software has quantified; researchers have to trust that the software has accurately quantified all ions from isotopes of a given peptide, excluding any overlapping ions derived from other peptides. The history of proteomics research has been one in which studies of highly variable quality have been published. There is also little quality control or benchmarking performed on quantitative software (10), meaning it is difficult to make quality judgments on a set of peptide and protein abundance values. The PSI has recently developed mzML, which can capture raw or processed MS data in a vendor neutral format, and the mzIdentML standard, to capture search engine results and the important metadata (such as software parameters), such that peptide and protein identification data can be interpreted consistently. These two standards are now being used for data sharing and to support open source software development, so that informatics groups can focus on algorithmic development rather than file format conversions. Until now, there has been no widely used open source format or data standard for capturing metadata and data relating to the quantitation step of analysis pipelines. In this work, we report the mzQuantML standard from the PSI, which has recently completed the PSI standardization process (11), from which version 1.0 was released. We believe that quantitative proteomics research will benefit from improved capabilities for tracing what manipulations have happened to data at each stage of the analysis process. The mzQuantML standard has been designed to store quantitative values calculated for features, peptides, proteins, and/or protein groups (where there is ambiguity in protein inference), plus associated software parameters. It has also been designed to accommodate small molecule data to improve interoperability with metabolomics investigations. The format can represent experimental replicates and grouping of replicates, and it has been designed via an open and transparent process.     

A critical appraisal of techniques, software packages, and standards for quantitative proteomic analysis

Abstract New methods for performing quantitative proteome analyses based on differential labeling protocols or label-free techniques are reported in the literature on an almost monthly basis. In parallel, a correspondingly vast number of software tools for the analysis of quantitative proteomics data has also been described in the literature and produced by private companies. In this article we focus on the review of some of the most popular techniques in the field and present a critical appraisal of several software packages available to process and analyze the data produced. We also describe the importance of community standards to support the wide range of software, which may assist researchers in the analysis of data using different platforms and protocols. It is intended that this review will serve bench scientists both as a useful reference and a guide to the selection and use of different pipelines to perform quantitative proteomics data analysis. We have produced a web-based tool ( http://www.proteosuite.org/?q=other_resources ) to help researchers find appropriate software for their local instrumentation, available file formats, and quantitative methodology.     

Chronic exposure of diesel exhaust particles induces alveolar enlargement in mice

Background

Diesel exhaust particles (DEPs) are deposited into the respiratory tract and are thought to be a risk factor for the development of diseases of the respiratory system. In healthy individuals, the timing and mechanisms of respiratory tract injuries caused by chronic exposure to air pollution remain to be clarified.

Methods

We evaluated the effects of chronic exposure to DEP at doses below those found in a typical bus corridor in Sao Paulo (150 μg/m³). Male BALB/c mice were divided into mice receiving a nasal instillation: saline (saline; n = 30) and 30 μg/10 μL of DEP (DEP; n = 30). Nasal instillations were performed five days a week, over a period of 90 days. Bronchoalveolar lavage (BAL) was performed, and the concentrations of interleukin (IL)-4, IL-10, IL-13 and interferon-gamma (INF-γ) were determined by ELISA-immunoassay. Assessment of respiratory mechanics was performed. The gene expression of Muc5ac in lung was evaluated by RT-PCR. The presence of IL-13, MAC2+ macrophages, CD3+, CD4+, CD8+ T cells and CD20+ B cells in tissues was analysed by immunohistochemistry. Bronchial thickness and the collagen/elastic fibers density were evaluated by morphometry. We measured the mean linear intercept (Lm), a measure of alveolar distension, and the mean airspace diameter (D0) and statistical distribution (D2).

Results

DEP decreased IFN-γ levels in BAL (p = 0.03), but did not significantly alter IL-4, IL-10 and IL-13 levels. MAC2+ macrophage, CD4+ T cell and CD20+ B cell numbers were not altered; however, numbers of CD3+ T cells (p ≤ 0.001) and CD8+ T cells (p ≤ 0.001) increased in the parenchyma. Although IL-13 (p = 0.008) expression decreased in the bronchiolar epithelium, Muc5ac gene expression was not altered in the lung of DEP-exposed animals. Although respiratory mechanics, elastic and collagen density were not modified, the mean linear intercept (Lm) was increased in the DEP-exposed animals (p ≤ 0.001), and the index D2 was statistically different (p = 0.038) from the control animals.

Conclusion

Our data suggest that nasal instillation of low doses of DEP over a period of 90 days results in alveolar enlargement in the pulmonary parenchyma of healthy mice.

Electronic supplementary material

The online version of this article (doi:10.1186/s12931-015-0172-z) contains supplementary material, which is available to authorized users.     

A new species of Mesocoelium (Digenea: Mesocoeliidae) found in Rhinella marina (Amphibia: Bufonidae) from Brazilian Amazonia

Mesocoelium lanfrediaesp. nov. (Digenea: Mesocoeliidae) inhabits the small intestine of Rhinella marina (Amphibia: Bufonidae) and is described here, with illustrations provided by light, scanning electron microscopy and molecular approachs. M. lanfrediae sp. nov. presents the typical characteristics of the genus, but is morphometrically and morphologically different from the species described previously. The main diagnostic characteristics of M. lanfrediae sp. nov. are (i) seven pairs of regularly-distributed spherical papillae on the oral sucker, (ii) ventral sucker outlined by four pairs of papillae distributed in a uniform pattern and interspersed with numerous spines, which are larger at the posterior margin and (iii) small, rounded tegumentary papillae around the opening of the oral sucker, which are morphologically different from those of the oral sucker itself, some of which are randomly disposed in the ventrolateral tegumentary region of the anterior third of the body. Addionally, based on SSU rDNA, a phylogenetic analysis including Brachycoeliidae and Mesocoeliidae taxa available on GenBank established the close relationship between M. lanfrediae sp. nov. and Mesocoelium sp.     

Biosynthesis of chlorophyll P-680 of photosystem II in greening leaves of plants adapted to heat shock

In etiolated pea and maize leaves illuminated after incubation at 38 degreesC, a new dark reaction was shown manifested in the bathochromic shift of spectral bands and accompanied by esterification of the product of protochlorophyllide photochemical reduction--Chld 684/676: Chld 684/676 --> Chl 688/680. After completion of the reaction a rapid (20-30 sec) quenching of the fluorescence of the reaction product (Chl 688/680) was observed. The reaction Chld 684/676 --> Chl 688/680 is inhibited under anaerobic conditions and in the presence of cyanide; the reaction accompanied by Chl 688/680 fluorescence quenching is not observed in pea mutants with impaired function of photosystem II reaction centers. The spectral properties of the formed Chl form with the absorption maximum at 680 nm, fluorescence quenching, and simultaneous synthesis of pheophytin suggest that the reaction is connected with the chlorophyll of photosystem II reaction center--P-680.