Nicolas Guillemin

Serum proteome profiling in canine idiopathic dilated cardiomyopathy using TMT-based quantitative proteomics approach

Idiopathic dilated cardiomyopathy (iDCM) is a primary myocardial disorder with an unknown aetiology, characterized by reduced contractility and ventricular dilation of the left or both ventricles. Naturally occurring canine iDCM was used herein to identify serum proteomic signature of the disease compared to the healthy state, providing an insight into underlying mechanisms and revealing proteins with biomarker potential. To achieve this, we used high-throughput label-based quantitative LC-MS/MS proteomics approach and bioinformatics analysis of the in silico inferred interactome protein network created from the initial list of differential proteins. To complement the proteomic analysis, serum biochemical parameters and levels of know biomarkers of cardiac function were measured. Several proteins with biomarker potential were identified, such as inter-alpha-trypsin inhibitor heavy chain H4, microfibril-associated glycoprotein 4 and apolipoprotein A-IV, which were validated using an independent method (Western blotting) and showed high specificity and sensitivity according to the receiver operating characteristic curve analysis. Bioinformatics analysis revealed involvement of different pathways in iDCM, such as complement cascade activation, lipoprotein particles dynamics, elastic fibre formation, GPCR signalling and respiratory electron transport chain. SIGNIFICANCE Idiopathic dilated cardiomyopathy is a severe primary myocardial disease of unknown cause, affecting both humans and dogs. This study is a contribution to the canine heart disease research by means of proteomic and bioinformatic state of the art analyses, following similar approach in human iDCM research. Importantly, we used serum as non-invasive and easily accessible biological source of information and contributed to the scarce data on biofluid proteome research on this topic. Bioinformatics analysis revealed biological pathways modulated in canine iDCM with potential of further targeted research. Also, several proteins with biomarker potential have been identified and successfully validated.

Proteomics in Veterinary Medicine and Animal Science: Neglected Scientific Opportunities with Immediate Impact

Animal/veterinary proteomics is an evolving field which holds a great promise not only for fundamental and applied discoveries regarding biology and pathology of domestic species, but can also be implemented in comparative applications of human diseases research. Experimental proteomics in domestic animals have advantages over use of rodents, such as multiple sampling in time series and availability of biological samples in sufficient volume for multiple analyses, such that both experimental and natural disease processes can be investigated. While there are certain technical limitations in the expansion of the field, they can currently be circumvented and in the future mastered with a greater participation of proteomic experts, which will in turn drive the accessibility of species‐specific reagents, data volume expansion in bioinformatic databases, and increased funding. This Viewpoint highlights some comparative proteomics studies addressing important issues and encourages readers to expand their horizons of domestic animal proteomics research. It will hopefully inspire new fruitful collaborations between veterinary and animal scientists and proteomic specialists for research in these areas that can have immediate and direct impact on health, society, and the economy.

Surface Proteome Biotinylation Combined with Bioinformatic Tools as a Strategy for Predicting Pathogen Interacting Proteins

Constant advancements in methodology and mass spectrometry instrumentation, genome sequencing and bioinformatic tools have enabled the identification of numerous pathogen proteomes. Identifying the pathogen interacting proteins by means of high-throughput techniques is key for understanding pathogen invasion and survival mechanisms and in such a way proposing specific proteins as pharmaceutical targets. Herein we describe the methodology for the enrichment and identification of pathogen surface proteome using cell surface protein biotinylation followed by LC-MS/MS and bioinformatic analyses of such data. This strategy is to be employed for the determination of protein subcellular localization and prediction of potential pathogen interacting proteins.

Quantitative proteomics using tandem mass tags in relation to the acute phase protein response in chicken challenged with Escherichia coli lipopolysaccharide endotoxin

The inflammatory response in chickens (Gallus Gallus domesticus) is an integral part of the birds response to infection. Detailing proteomic changes occurring during infection would be beneficial to the poultry industry, offering opportunities for comparative pathophysiological analysis. The objective of this study was to quantify the changes in the plasma proteome in chickens challenged with lipopolysaccharide (LPS), a bacterial endotoxin known to stimulate the host innate immune system. Plasma from chicken (N = 6) challenged with Escherichia coli (LPS) (2 mg/kg body weight) was collected pre (0 h) and at 12, 24, 48, and 72 h post-injection along with plasma from a control group (N = 6) challenged with sterile saline. Samples were analysed by a quantitative Tandem Mass Tags approach using a Q-Exactive-Plus mass-spectrometer. Identification and relative quantification were performed using Proteome Discoverer, and data were analysed using R. Gene Ontology terms were analysed by Cytoscape based on the Gallus gallus database. Finally, 87 significantly regulated proteins were found, including serum-amyloid-A, ovotransferrin and alpha-1-acid-glycoprotein, showing a significant effect of time post-injection in the LPS-treated group. Different pathways related with protein activation cascade and heterotopic cell-cell adhesion were affected by LPS-challenge. LPS-challenged chickens demonstrate significant changes to the plasma proteome with both increases and decreases of individual proteins within 12 h of challenge. SIGNIFICANCE: The injection of chicken with bacterial lipopolysaccharide followed by sequential plasma and clinical analysis of the bird, is a long established and a widely used model for inflammation and infection studies. This study, utilising and combining proteomic and immunoassay analysis with bioinformatic analysis, revealed that several biological pathways are modulated during this early period of inflammation. In addition, proteins with biomarker potential were identified and successfully validated. This experimental model also demonstrated potential for pathophysiological mechanism investigation and as an inflammatory model for biomedical research. There is, despite plasma being an easily accessible biological matrix which is representative of the health status of the bird, scarce data on the chicken plasma proteome. This research makes a positive contribution to the current field, generating significant data for continuing comparative analysis.

Integrated dataset on acute phase protein response in chicken challenged with Escherichia coli lipopolysaccharide endotoxin

Data herein describe the quantitative changes in the plasma proteome in chickens challenged with lipopolysaccharide (LPS), a bacterial endotoxin known to stimulate the host innate immune system obtained by shotgun quantitative proteomic tandem mass tags approach using high-resolution Orbitrap technology. Statistical and bioinformatic analyses were performed to specify the effect of bacterial endotoxin. Plasma from chicken (N=6) challenged with Escherichia coli (LPS) (2 mg/kg body weight) was collected pre (0 h) and at 12, 24, 48, and 72 h post injection along with plasma from a control group (N=6) challenged with sterile saline. Protein identification and relative quantification were performed using Proteome Discoverer, and data were analysed using R. Gene Ontology terms were analysed by the Cytoscape application ClueGO based on Gallus gallus GO Biological Process database, and refined by REVIGO. Absolute quantification of several acute phase proteins, e.g. alpha-1-acid glycoprotein (AGP), serum amyloid A (SAA) and ovotrensferrin (OVT) was performed by immunoassays to validate the LC-MS results. The data contained within this article are directly related to our research article”Quantitative proteomics using tandem mass tags in relation to the acute phase protein response in chicken challenged with Escherichia coli lipopolysaccharide endotoxin” [1]. The raw mass spectrometric data generated in this study were deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD009399 (http://proteomecentral.proteomexchange.org/cgi/GetDataset?ID=PXD009399).

Changes in salivary analytes in canine parvovirus: A high-resolution quantitative proteomic study

Abstract The present study evaluated the changes in salivary proteome in parvoviral enteritis (PVE) in dogs through a high-throughput quantitative proteomic analysis. Saliva samples from healthy dogs and dogs with severe parvovirosis that survived or perished due to the disease were analysed and compared by Tandem Mass Tags (TMT) analysis. Proteomic analysis quantified 1516 peptides, and 287 (corresponding to 190 proteins) showed significantly different abundances between studied groups. Ten proteins were observed to change significantly between dogs that survived or perished due to PVE. Bioinformatics’ analysis revealed that saliva reflects the involvement of different pathways in PVE such as catalytic activity and binding, and indicates antimicrobial humoral response as a pathway with a major role in the development of the disease. These results indicate that saliva proteins reflect physiopathological changes that occur in PVE and could be a potential source of biomarkers for this disease.

The Challenges and Advances in Diagnosis of Vector-Borne Diseases: Where Do We Stand?

Vector-borne diseases (VBD) are of major importance to human and animal health. In recent years, VBD have been emerging or re-emerging in many geographical areas, alarming new disease threats and economic losses. The precise diagnosis of many of these diseases still remains a major challenge because of the lack of comprehensive data available on accurate and reliable diagnostic methods. Here, we conducted a systematic and in-depth review of the former, current, and upcoming techniques employed for the diagnosis of VBD.