List_MAPS - Human listeriosis cases are due to the ingestion of contaminated foods with the pathogenic bacteria Listeria monocytogenes
ABSTRACT: Human listeriosis cases are due to the ingestion of contaminated foods with the pathogenic bacteria Listeria monocytogenes. The reduction of water availability in food workshops by decreasing the air relative humidity (RH) is one strategy to improve the control of bacterial contamination. This study aims to develop and implement an MSI approach on L. monocytogenes biofilms and proof of concept using a dehumidified stress condition. MSI allowed examining the distribution of low molecular weight proteins within the biofilms subjected to a dehumidification environment, mimicking the one present in a food workshop (10°C, 75% RH). Furthermore, a LC-MS/MS approach was made to link the dots between MSI and protein identification. Five identified proteins were assigned to registered MSI m/z, including two cold-shock proteins and a ligase involved in cell wall biogenesis.
Project description:Aberrant protease activity has been implicated in the etiology of various prevalent diseases including neurodegeneration and cancer, in particular metastasis. Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) has recently been established as a key technology for bioanalysis of multiple biomolecular classes such as proteins, lipids, and glycans. However, it has not yet been systematically explored for investigation of a tissue’s endogenous protease activity. In this study, we demonstrate that different tissues, spray-coated with substance P as a tracer, digest this peptide with different time-course profiles. Furthermore, we reveal that distinct cleavage products originating from substance P are generated transiently and that proteolysis can be attenuated by protease inhibitors in a concentration-dependent manner. To show the translational potential of the method, we analyzed protease activity of gastric carcinoma in mice. Our MSI and quantitative proteomics results reveal differential distribution of protease activity – with strongest activity being observed in mouse tumor tissue, suggesting the general applicability of the workflow in animal pharmacology and clinical studies.Aberrant protease activity has been implicated in the etiology of various prevalent diseases including neurodegeneration and cancer, in particular metastasis. Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) has recently been established as a key technology for bioanalysis of multiple biomolecular classes such as proteins, lipids, and glycans. However, it has not yet been systematically explored for investigation of a tissue’s endogenous protease activity. In this study, we demonstrate that different tissues, spray-coated with substance P as a tracer, digest this peptide with different time-course profiles. Furthermore, we reveal that distinct cleavage products originating from substance P are generated transiently and that proteolysis can be attenuated by protease inhibitors in a concentration-dependent manner. To show the translational potential of the method, we analyzed protease activity of gastric carcinoma in mice. Our MSI and quantitative proteomics results reveal differential distribution of protease activity – with strongest activity being observed in mouse tumor tissue, suggesting the general applicability of the workflow in animal pharmacology and clinical studies.
Project description:Introduction The identification of proteins involved in brain ischemia might allow the discovery of new putative biomarkers or potential therapeutic target of one of the most important neurological disorders. MALDI Imaging-Mass-Spectrometry (IMS) is a powerful technique that allows the visualization of protein distribution along a tissue without labeling. Our aim is to study the distribution of proteins along mouse brain after an ischemic insult and to identify relevant proteins involved in brain ischemia. Materials and methods We occluded the middle cerebral artery (60min) of C57BL/6J mice (n=4) with 24h of reperfusion. Brain slices were analyzed by MALDI-TOF and mass spectra from infarct (IC) and contralateral (CL) regions were compared using ClinProTools. Relevant m/z were selected after PCA analysis and their ion distribution maps were analyzed by FlexImagin3.0. Protein identification was conducted on mouse brain homogenates through a bottom-up approach consisting on complementary fractionations based on tricine gels and RP-HPLC. The identifications were confirmed by immunohistochemistry. Results We identified 102 m/z with different abundances between IC and CL (p<0.05), from which 21 m/z were selected by PCA as more relevant. Thirteen of them were found increased in the infarct region and 4 m/z showed a discrimination higher than 90% between IC and CL. After bottom-up identification, immunohistochemistry analysis confirmed increased ATP5i and COX6C expressions, and decreased UMP-CMP kinase in IC compared to CL. Conclusions We identified for the first time by MALDI-IMS several m/z peaks with different abundances between IC and CL. These proteins involved in brain ischemia might represent potential diagnostic biomarkers or target molecules for neurological recovery.
Project description:There are no widely-accepted prognostic markers currently available to predict outcomes in patients with triple-negative breast cancer (TNBC), and no targeted therapies with confirmed benefit. We have used MALDI mass spectrometry imaging (MSI) of tryptic peptides to compare regions of cancer and benign tissue in 10 formalin-fixed, paraffin-embedded sections of TNBC tumors. Proteins were identified by reference to a peptide library constructed by LC-MALDI-MS/MS analyses of the same tissues. The prognostic significance of proteins that distinguished between cancer and benign regions was estimated by Kaplan-Meier analysis of their gene expression from public databases. Among peptides that distinguished between cancer and benign tissue in at least 3 tissues with a ROC area under the curve >0.7, 14 represented proteins identified from the reference library, including proteins not previously associated with breast cancer. Initial network analysis using the STRING database showed no obvious functional relationships except among collagen subunits COL1A1, COL1A2, and COL63A, but manual curation, including the addition of EGFR to the analysis, revealed a unique network connecting 10 of the 14 proteins. Kaplan-Meier survival analysis to examine the relationship between tumor expression of genes encoding the 14 proteins, and recurrence-free survival (RFS) in patients with basal-like TNBC showed that, compared to low expression, high expression of nine of the genes was associated with significantly worse RFS, most with hazard ratios >2. In contrast, in estrogen receptor-positive tumors, high expression of these genes showed only low, or no, association with worse RFS. These proteins are proposed as putative markers of RFS in TNBC, and some may also be considered as possible targets for future therapies.
Project description:MALDI mass spectrometry imaging (MSI) enables label-free, spatially resolved analysis of a wide range of analytes in tissue sections. Quantitative analysis of MSI datasets is typically performed on single pixels or manually assigned regions of interest (ROI). However, many sparse, small objects such as Alzheimer’s disease (AD) brain deposits of amyloid peptides called plaques are neither single pixels nor ROI. Here, we propose a new approach to facilitate comparative computational evaluation of amyloid plaque-like objects by MSI: a fast PLAQUE PICKER tool that enables statistical evaluation of heterogeneous amyloid peptide composition. Comparing two AD mouse models, APP NL-G-F and APP PS1, we identified distinct heterogeneous plaque populations in the NL-G-F model, but only one class of plaques in the PS1 model. We propose quantitative metrics for the comparison of technical and biological MSI replicates.
Project description:Tryptic peptides and N-glycans were spatially mapped and visualised on formalin-fixed paraffin-embedded (FFPE) endometrial cancer tissue microarrays (TMAs) using an ultrafleXtreme MALDI-ToF/ToF MS instrument (Bruker Daltonics). FFPE egg white was placed either side of each TMA and used as an external control to monitor detector performance and sample preparation.
Project description:In osteoarthritis (OA), impairment of cartilage regeneration can be related to a defective chondrogenic differentiation of mesenchymal stromal cells (MSCs). Therefore, understanding the proteomic- and metabolomic-associated molecular events during the chondrogenesis of MSCs could provide alternative targets for therapeutic intervention. Here, a SILAC-based proteomic analysis identified 43 proteins related with metabolic pathways whose abundance was significantly altered during the chondrogenesis of OA human bone marrow MSCs (hBMSCs). Then, the level and distribution of metabolites was analyzed in these cells and healthy controls by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI), leading to the recognition of characteristic metabolomic profiles at the early stages of differentiation. Finally, integrative pathway analysis showed that UDP-glucuronic acid synthesis and amino sugar metabolism were downregulated in OA hBMSCs during chondrogenesis compared to healthy cells. Alterations in these metabolic pathways may disturb the production of hyaluronic acid (HA) and other relevant cartilage extracellular matrix (ECM) components. This work provides a novel integrative insight into the molecular alterations of osteoarthritic MSCs and potential therapeutic targets for OA drug development through the enhancement of chondrogenesis.
Project description:This dataset contains MS imaging raw data as well as the SCiLS file used to confirm the regions in dog sarcoma biopsies classified by the SpiderMass system after analysis of the consecutive tissue section. All the tissues were gathered during dog sarcoma surgeries, snap-frozen and analysed later in the lab. The images were acquired on a MALDI LTQ orbitrap XL hybrid instrument.
Project description:An integrated diagnosis using molecular features is recommended in the updated World Health Organization (WHO) classification. Our aim was to explore the role of MALDI-Mass spectrometry imaging (MSI) coupled to microproteomics in order to classify anaplastic glioma by integration of clinical data.
Project description:During aging, the kidney undergoes functional and physiological changes that are closely affiliated with chronic kidney disease (CKD). There is increasing evidence supporting the role of lipid or lipid-derived mediators in the pathogenesis of CKD and other aging-related diseases. To understand the role of lipids in various metabolic processes during kidney aging, we conducted MALDI imaging mass spectrometry (MALDI-IMS) analysis in kidneys harvested from young (2 months old, n=3) and old mice (24 months old, n=3). MALDI-IMS analysis showed an increase in ceramide level and a decrease in sphingomyelin (SM) and phosphatidylcholine (PC) levels in kidneys of old mice. The increased expression of cPLA2 and SMPD1 protein in aged kidney was confirmed by immunohistochemistry and western blot analysis. Our MALDI-IMS data showed the altered distribution of lipids in aged kidney as indicative of aging-related functional changes of the kidney. Combined analysis of MALDI-IMS and IHC confirmed lipidomic changes and expression levels of responsible enzymes as well as morphological changes.