Bayesian probit regression model for the diagnosis of pulmonary fibrosis: proof-of-principle

Background

The accurate diagnosis of idiopathic pulmonary fibrosis (IPF) is a major clinical challenge. We developed a model to diagnose IPF by applying Bayesian probit regression (BPR) modelling to gene expression profiles of whole lung tissue.

Methods

Whole lung tissue was obtained from patients with idiopathic pulmonary fibrosis (IPF) undergoing surgical lung biopsy or lung transplantation. Controls were obtained from normal organ donors. We performed cluster analyses to explore differences in our dataset. No significant difference was found between samples obtained from different lobes of the same patient. A significant difference was found between samples obtained at biopsy versus explant. Following preliminary analysis of the complete dataset, we selected three subsets for the development of diagnostic gene signatures: the first signature was developed from all IPF samples (as compared to controls); the second signature was developed from the subset of IPF samples obtained at biopsy; the third signature was developed from IPF explants. To assess the validity of each signature, we used an independent cohort of IPF and normal samples. Each signature was used to predict phenotype (IPF versus normal) in samples from the validation cohort. We compared the models' predictions to the true phenotype of each validation sample, and then calculated sensitivity, specificity and accuracy.

Results

Surprisingly, we found that all three signatures were reasonably valid predictors of diagnosis, with small differences in test sensitivity, specificity and overall accuracy.

Conclusions

This study represents the first use of BPR on whole lung tissue; previously, BPR was primarily used to develop predictive models for cancer. This also represents the first report of an independently validated IPF gene expression signature. In summary, BPR is a promising tool for the development of gene expression signatures from non-neoplastic lung tissue. In the future, BPR might be used to develop definitive diagnostic gene signatures for IPF, prognostic gene signatures for IPF or gene signatures for other non-neoplastic lung disorders such as bronchiolitis obliterans.

     

Optimization of fecal sample preparation for untargeted LC-HRMS based metabolomics

Introduction

Collecting feces is easy. It offers direct outcome to endogenous and microbial metabolites.

Objectives

In a context of lack of consensus about fecal sample preparation, especially in animal species, we developed a robust protocol allowing untargeted LC-HRMS fingerprinting.

Methods

The conditions of extraction (quantity, preparation, solvents, dilutions) were investigated in bovine feces.

Results

A rapid and simple protocol involving feces extraction with methanol (1/3, M/V) followed by centrifugation and a step filtration (10 kDa) was developed.

Conclusion

The workflow generated repeatable and informative fingerprints for robust metabolome characterization.

     

Clinical relevance of glycerophospholipid,sphingomyelin and glutathione metabolism in the pathogenesis of pharyngolaryngeal cancer in smokers: the Korean Cancer Prevention Study-II

Introduction

Although smoking is a major risk factor for pharyngolaryngeal cancer, most smokers do not develop pharyngolaryngeal cancer.

Objectives

In the prospective Korean Cancer Prevention Study-II (KCPS-II), we investigated the application of metabolomics to differentiate smokers with incident pharyngolaryngeal cancer (pharyngolaryngeal cancer group) from smokers who remained free from cancer (controls) during a mean follow-up period of 7 years and aimed to discover valuable early biomarkers of pharyngolaryngeal cancer.

Methods

We used baseline serum samples from 30 smoking men with incident pharyngolaryngeal cancer and 59 age-matched cancer-free smoking men. Metabolic alterations associated with the incidence of pharyngolaryngeal cancer were investigated by performing metabolomics on baseline serum samples using ultra-performance liquid chromatography-linear-trap quadrupole-Orbitrap mass spectrometry.

Results

Compared to the control group, the pharyngolaryngeal cancer group showed significantly higher oxidized LDL levels. Seventeen metabolites were differentially abundant between the two groups. At baseline, compared to controls, smokers with incident pharyngolaryngeal cancer during follow-up showed significantly higher levels of pyroglutamic acid (glutathione metabolism) but lower levels of lysophosphatidylcholines (lysoPCs) C14:0, C15:0, C16:0, C17:0, C18:0, and C20:5; glycerophosphocholine; PC C36:5; lysoPEs C16:0, C20:1, and C22:0 (glycerophospholipid metabolism); SM (d18:0/16:1); and SM (d18:1/18:1) (sphingomyelin metabolism). Furthermore, smokers with incident pharyngolaryngeal cancer showed significantly higher levels of oleamide and lower levels of tryptophan and linoleyl carnitine at baseline than cancer-free smokers.

Conclusion

This prospective study showed the clinical relevance of dysregulated metabolism of glutathione, glycerophospholipids and sphingolipids to the pathogenesis of pharyngolaryngeal cancer among smokers. These data suggest that the dysregulation of these metabolic processes may be a key mechanism underlying pharyngolaryngeal cancer progression and development.

     

Using multitask classification methods to investigate the kinase-specific phosphorylation sites

Background

Identification of phosphorylation sites by computational methods is becoming increasingly important because it reduces labor-intensive and costly experiments and can improve our understanding of the common properties and underlying mechanisms of protein phosphorylation.

Methods

A multitask learning framework for learning four kinase families simultaneously, instead of studying each kinase family of phosphorylation sites separately, is presented in the study. The framework includes two multitask classification methods: the Multi-Task Least Squares Support Vector Machines (MTLS-SVMs) and the Multi-Task Feature Selection (MT-Feat3).

Results

Using the multitask learning framework, we successfully identify 18 common features shared by four kinase families of phosphorylation sites. The reliability of selected features is demonstrated by the consistent performance in two multi-task learning methods.

Conclusions

The selected features can be used to build efficient multitask classifiers with good performance, suggesting they are important to protein phosphorylation across 4 kinase families.

     

Potential benefit of bosentan therapy in borderline or less severe pulmonary hypertension secondary to idiopathic pulmonary fibrosis—an interim analysis of results from a prospective,single-center,randomized, parallel-group study

Background

No drugs have been approved for the treatment of patients with pulmonary hypertension (PH) secondary to idiopathic pulmonary fibrosis (IPF), particularly those with idiopathic honeycomb lung. This study was conducted to investigate the long-term efficacy and safety of bosentan for PH based on changes in prognosis and respiratory failure.

Methods

IPF patients with borderline or less severe PH and completely organized honeycomb lung were randomized (1:1) to bosentan or no treatment for PH for 2 years and assessed at baseline and every 6 months for respiratory failure, activities of daily living (ADL), lung and heart functions by right cardiac catheterization, and other parameters. An interim analysis was performed, however, following detection of a significant survival benefit favoring bosentan therapy.

Results

Significant differences were noted for the bosentan-treated (n?=?12) vs. untreated (n?=?12) groups in hospital-free survival (603.44?±?50.074 days vs. 358.87?±?68.65 days; hazard ratio [HR], 0.19; P?=?0.017) and overall survival (671 days vs. 433.78?±?66.98 days; HR, 0.10; P?=?0.0082). Again, significant improvements were noted for the bosentan-treated group from baseline to month 6 or 12 in several indices in ADL, pulmonary circulation, and %DLCO. Without requiring O₂ inhalation, bosentan was associated with no increase but a trend toward a decrease in adverse events and an improvement in respiratory status.

Conclusions

Bosentan tended to improve prognosis and ADL without worsening respiratory failure in IPF patients with borderline or less severe PH and completely organized honeycomb lung alone.

Trial registration

This study was registered on December 18, 2010 with UMIN-CTR Clinical Trial as UMIN000004749 to investigate the long-term influence of bosentan on cardiac function, as well as its cardioprotective efficacy and safety, in patients with pulmonary hypertension secondary to concurrent COPD and IPF, respectively.

     

M2 macrophages promote myofibroblast differentiation of LR-MSCs and are associated with pulmonary fibrogenesis

Background

Idiopathic pulmonary fibrosis (IPF) is a devastating disease characterized by the histopathological pattern of usual interstitial pneumonia and is associated with a high mortality rate. Recently, lung resident mesenchymal stem cells (LR-MSCs) have been identified as an important contributor to myofibroblast activation in pulmonary fibrosis. Macrophages are also believed to play a critical role in pulmonary fibrosis. However, the underlying connections between LR-MSCs and macrophages in the pathogenesis of pulmonary fibrosis are still elusive.

Methods

In this study, we investigated the interaction between LR-MSCs and macrophages using a bleomycin-induced mouse pulmonary fibrosis model and a coculture system.

Results

Here, we show that blocking pulmonary macrophage infiltration attenuated bleomycin-induced pulmonary fibrosis. In addition, as determined by flow cytometry, we discovered that the recruited macrophages in fibrotic lungs of bleomycin-treated mice were mainly M2 macrophages. In particular, we found that M2, rather than M1 macrophages, promoted myofibroblast differentiation of LR-MSCs. Moreover, we demonstrated that suppression of the Wnt/β-catenin signaling pathway could attenuate myofibroblast differentiation of LR-MSCs induced by M2 macrophages and bleomycin-induced pulmonary fibrosis. Tissue samples from IPF patients confirmed the infiltration of M2 macrophages and activation of Wnt/β-catenin signaling pathway.

Conclusion

In summary, this study furthered our understanding of the pulmonary fibrosis pathogenesis and highlighted M2 macrophages as a critical target for treating pulmonary fibrosis.

     

Pseudo current density maps of electrophysiological heart,nerve or brain function and their physical basis

Background

In recent years the visualization of biomagnetic measurement data by so-called pseudo current density maps or Hosaka-Cohen (HC) transformations became popular.

Methods

The physical basis of these intuitive maps is clarified by means of analytically solvable problems.

Results

Examples in magnetocardiography, magnetoencephalography and magnetoneurography demonstrate the usefulness of this method.

Conclusion

Hardware realizations of the HC-transformation and some similar transformations are discussed which could advantageously support cross-platform comparability of biomagnetic measurements.

     

Crystal structure of <Emphasis Type="Italic">Escherichia coli</Emphasis> protein ybgI,a toroidal structure with a dinuclear metal site

Background

The protein encoded by the gene ybgI was chosen as a target for a structural genomics project emphasizing the relation of protein structure to function.

Results

The structure of the ybgI protein is a toroid composed of six polypeptide chains forming a trimer of dimers. Each polypeptide chain binds two metal ions on the inside of the toroid.

Conclusion

The toroidal structure is comparable to that of some proteins that are involved in DNA metabolism. The di-nuclear metal site could imply that the specific function of this protein is as a hydrolase-oxidase enzyme.

     

Untargeted metabolomics suffers from incomplete raw data processing

Introduction

Untargeted metabolomics is a powerful tool for biological discoveries. To analyze the complex raw data, significant advances in computational approaches have been made, yet it is not clear how exhaustive and reliable the data analysis results are.

Objectives

Assessment of the quality of raw data processing in untargeted metabolomics.

Methods

Five published untargeted metabolomics studies, were reanalyzed.

Results

Omissions of at least 50 relevant compounds from the original results as well as examples of representative mistakes were reported for each study.

Conclusion

Incomplete raw data processing shows unexplored potential of current and legacy data.

     

Expression of Toll-like receptor 2 is up-regulated in monocytes from patients with chronic obstructive pulmonary disease

Background

Chronic obstructive pulmonary disease (COPD) is characterised by pulmonary and systemic inflammation which flare-up during episodes of acute exacerbation (AECOPD). Given the role of Toll-like receptors (TLRs) in the induction of inflammatory responses we investigated the involvement of TLRs in COPD pathogenesis.

Methods

The expression of TLR-2, TLR-4 and CD14 in monocytes was analyzed by flow cytometry. To study the functional responses of these receptors, monocytes were stimulated with peptidoglycan or lipopolysaccharide and the amounts of TNFα and IL-6 secreted were determined by ELISA.

Results

We found that the expression of TLR-2 was up-regulated in peripheral blood monocytes from COPD patients, either clinically stable or during AECOPD, as compared to never smokers or smokers with normal lung function. Upon stimulation with TLR-2 ligand monocytes from COPD patients secreted increased amounts of cytokines than similarly stimulated monocytes from never smokers and smokers. In contrast, the expressions of TLR-4 and CD14 were not significantly different between groups, and the response to lipopolysaccharide (a TLR-4 ligand) stimulation was not significantly different either. At discharge from hospital TLR-2 expression was down-regulated in peripheral blood monocytes from AECOPD patients. This could be due to the treatment with systemic steroids because, in vitro, steroids down-regulated TLR-2 expression in a dose-dependent manner. Finally, we demonstrated that IL-6, whose plasma levels are elevated in patients, up-regulated in vitro TLR-2 expression in monocytes from never smokers.

Conclusion

Our results reveal abnormalities in TLRs expression in COPD patients and highlight its potential relationship with systemic inflammation in these patients.

     

Metabolomic profiling of maternal hair suggests rapid development of intrahepatic cholestasis of pregnancy

Introduction

Intrahepatic cholestasis of pregnancy (ICP) is a common maternal liver disease; development can result in devastating consequences, including sudden fetal death and stillbirth. Currently, recognition of ICP only occurs following onset of clinical symptoms.

Objective

Investigate the maternal hair metabolome for predictive biomarkers of ICP.

Methods

The maternal hair metabolome (gestational age of sampling between 17 and 41 weeks) of 38 Chinese women with ICP and 46 pregnant controls was analysed using gas chromatography–mass spectrometry.

Results

Of 105 metabolites detected in hair, none were significantly associated with ICP.

Conclusion

Hair samples represent accumulative environmental exposure over time. Samples collected at the onset of ICP did not reveal any metabolic shifts, suggesting rapid development of the disease.

     

KniMet: a pipeline for the processing of chromatography–mass spectrometry metabolomics data

Introduction

Data processing is one of the biggest problems in metabolomics, given the high number of samples analyzed and the need of multiple software packages for each step of the processing workflow.

Objectives

Merge in the same platform the steps required for metabolomics data processing.

Methods

KniMet is a workflow for the processing of mass spectrometry-metabolomics data based on the KNIME Analytics platform.

Results

The approach includes key steps to follow in metabolomics data processing: feature filtering, missing value imputation, normalization, batch correction and annotation.

Conclusion

KniMet provides the user with a local, modular and customizable workflow for the processing of both GC–MS and LC–MS open profiling data.

     

Quantitative analysis of tetrahydrofolate metabolites from <Emphasis Type="Italic">clostridium autoethanogenum</Emphasis>

Introduction

Quantification of tetrahydrofolates (THFs), important metabolites in the Wood–Ljungdahl pathway (WLP) of acetogens, is challenging given their sensitivity to oxygen.

Objective

To develop a simple anaerobic protocol to enable reliable THFs quantification from bioreactors.

Methods

Anaerobic cultures were mixed with anaerobic acetonitrile for extraction. Targeted LC–MS/MS was used for quantification.

Results

Tetrahydrofolates can only be quantified if sampled anaerobically. THF levels showed a strong correlation to acetyl-CoA, the end product of the WLP.

Conclusion

Our method is useful for relative quantification of THFs across different growth conditions. Absolute quantification of THFs requires the use of labelled standards.

     

Accumulation of Fascin<Superscript>+</Superscript> cells during experimental autoimmune neuritis

Background

Experimental autoimmune neuritis (EAN) is a well-known animal model of human demyelinating polyneuropathies and is characterized by inflammation and demyelination in the peripheral nervous system. Fascin is an evolutionarily highly conserved cytoskeletal protein of 55 kDa containing two actin binding domains that cross-link filamentous actin to hexagonal bundles.

Methods

Here we have studied by immunohistochemistry the spatiotemporal accumulation of Fascin?+?cells in sciatic nerves of EAN rats.

Results

A robust accumulation of Fascin?+?cell was observed in the peripheral nervous system of EAN which was correlated with the severity of neurological signs in EAN.

Conclusion

Our results suggest a pathological role of Fascin in EAN.

Virtual slides

The virtual slides for this article can be found here: http://www.diagnosticphatology.diagnomx.eu/vs/6734593451114811

     

A methodology for elucidating regulatory mechanisms leading to changes in lipid profiles

Introduction

It is difficult to elucidate the metabolic and regulatory factors causing lipidome perturbations.

Objectives

This work simplifies this process.

Methods

A method has been developed to query an online holistic lipid metabolic network (of 7923 metabolites) to extract the pathways that connect the input list of lipids.

Results

The output enables pathway visualisation and the querying of other databases to identify potential regulators. When used to a study a plasma lipidome dataset of polycystic ovary syndrome, 14 enzymes were identified, of which 3 are linked to ELAVL1—an mRNA stabiliser.

Conclusion

This method provides a simplified approach to identifying potential regulators causing lipid-profile perturbations.

     

Minimal variation of the plasma lipidome after delayed processing of neonatal cord blood

Background

Cord blood lipids are potential disease biomarkers. We aimed to determine if their concentrations were affected by delayed blood processing.

Method

Refrigerated cord blood from six healthy newborns was centrifuged every 12 h for 4 days. Plasma lipids were analysed by liquid chromatography/mass spectroscopy.

Results

Of 262 lipids identified, only eight varied significantly over time. These comprised three dihexosylceramides, two phosphatidylserines and two phosphatidylethanolamines whose relative concentrations increased and one sphingomyelin that decreased.

Conclusion

Delay in separation of plasma from refrigerated cord blood has minimal effect overall on the plasma lipidome.

     

Metabolic response of porcine colon explants to in vitro infection by <Emphasis Type="Italic">Brachyspira hyodysenteriae</Emphasis>: a leap into disease pathophysiology

Introduction

Swine dysentery caused by Brachyspira hyodysenteriae is a production limiting disease in pig farming. Currently antimicrobial therapy is the only treatment and control method available.

Objective

The aim of this study was to characterize the metabolic response of porcine colon explants to infection by B. hyodysenteriae.

Methods

Porcine colon explants exposed to B. hyodysenteriae were analyzed for histopathological, metabolic and pro-inflammatory gene expression changes.

Results

Significant epithelial necrosis, increased levels of l-citrulline and IL-1α were observed on explants infected with B. hyodysenteriae.

Conclusions

The spirochete induces necrosis in vitro likely through an inflammatory process mediated by IL-1α and NO.

     

Nucleotide binding domain 1 pharmacophore modeling for visualization and analysis of P-glycoprotein–flavonoid molecular interactions

Background

P-glycoprotein (P-gp) is a 170-kDa membrane protein. It provides a barrier function and help to excrete toxins from the body as a transporter. Some bioflavonoids have been shown to block P-gp activity.

Objective

To evaluate the important amino acid residues within nucleotide binding domain 1 (NBD1) of P-gp that play a key role in molecular interactions with flavonoids using structure-based pharmacophore model.

Methods

In the molecular docking with NBD1 models, a putative binding site of flavonoids was proposed and compared with the site for ATP. The binding modes for ligands were achieved using LigandScout to generate the P-gp–flavonoid pharmacophore models.

Results

The binding pocket for flavonoids was investigated and found these inhibitors compete with the ATP for binding site in NBD1 including the NBD1 amino acid residues identified by the in silico techniques to be involved in the hydrogen bonding and van der Waals (hydrophobic) interactions with flavonoids.

Conclusion

These flavonoids occupy with the same binding site of ATP in NBD1 proffering that they may act as an ATP competitive inhibitor.

     

Extraction of natural products from bark of <Emphasis Type="Italic">Betula pendula</Emphasis> using ionic liquids

Introduction

Aqueous–methanol mixtures have successfully been applied to extract a broad range of metabolites from plant tissue. However, a certain amount of material remains insoluble.

Objectives

To enlarge the metabolic compendium, two ionic liquids were selected to extract the methanol insoluble part of trunk from Betula pendula.

Methods

The extracted compounds were analyzed by LC/MS and GC/MS.

Results

The results show that 1-butyl-3-methylimidazolium acetate (IL-Ac) predominantly resulted in fatty acids, whereas 1-ethyl-3-methylimidazolium tosylate (IL-Tos) mostly yielded phenolic structures. Interestingly, bark yielded more ionic liquid soluble metabolites compared to interior wood.

Conclusion

From this one can conclude that the application of ionic liquids may expand the metabolic snapshot.