Global proteomic analysis of extracellular matrix in mouse and human brain highlights relevance to cerebrovascular disease.
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ABSTRACT:
SUBMITTER: Pokhilko A
PROVIDER: S-EPMC8392779 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:The extracellular matrix (ECM), a key interface between the cerebrovasculature and other cells within the neuro-glial-vascular unit , provides structural stability and modulates cell behaviour and signalling. These functions are dependent on ECM protein composition. Since ECM defects may contribute to a broad range of disorders including cerebrovascular disease, it is crucially important to characterize ECM composition to better understand the mechanisms by which the ECM modulates brain health and disease. To date, molecular studies of the cerebrovascular ECM have been limited. We therefore generated ECM extracts from vascular enriched tissues of both mouse and human post-mortem brain. We used mass spectrometry with off-line high-pH reversed-phase fractionation to increase proteome depth and characterized the identified proteins. This identified a large number of proteins (>1000 mouse, >2000 human) in the ECM-enriched fractions, with > 66% of the identified proteins covered in both human and mouse samples. We now report 147 ECM proteins, including collagens (as major ECM components), laminins, fibronectin and nidogens, that can be considered as the core constituents of the human brain vascular matrisome. We also identified 12 novel proteins in the ECM-enriched fraction, that may have regulate or interact with the matrisome, including several key regulators of neurovascular interactions, such as BCAM, CDH5 and PARVB. Many of the identified brain vascular matrisome proteins are encoded by genes identified in stroke and cerebral small vessel disease genome-wide association studies, underscoring the importance of the vascular ECM in health and disease. This brain vascular matrisome represents a powerful resource for investigating the impact of aging and disease on the cerebrovasculature.
2021-03-11 | PXD020436 | Pride
Project description:BACKGROUND: Investigations performed in mice and humans have acknowledged obesity as a low-grade inflammatory disease. Several molecular mechanisms have been convincingly shown to be involved in activating inflammatory processes and altering cell composition in white adipose tissue (WAT). However, the overall importance of these alterations, and their long-term impact on the metabolic functions of the WAT and on its morphology, remain unclear. RESULTS: Here, we analyzed the transcriptomic signature of the subcutaneous WAT in obese human subjects, in stable weight conditions and after weight loss following bariatric surgery. An original integrative functional genomics approach was applied to quantify relations between relevant structural and functional themes annotating differentially expressed genes in order to construct a comprehensive map of transcriptional interactions defining the obese WAT. These analyses highlighted a significant up-regulation of genes and biological themes related to extracellular matrix (ECM) constituents, including members of the integrin family, and suggested that these elements could play a major mediating role in a chain of interactions that connect local inflammatory phenomena to the alteration of WAT metabolic functions in obese subjects. Tissue and cellular investigations, driven by the analysis of transcriptional interactions, revealed an increased amount of interstitial fibrosis in obese WAT, associated with an infiltration of different types of inflammatory cells, and suggest that phenotypic alterations of human pre-adipocytes, induced by a pro-inflammatory environment, may lead to an excessive synthesis of ECM components. CONCLUSION: This study opens new perspectives in understanding the biology of human WAT and its pathologic changes indicative of tissue deterioration associated with the development of obesity.
| S-EPMC2395253 | biostudies-literature
Project description:Degenerative aortic stenosis (AS) is the most common worldwide cause of valve replacement. The aortic valve is a thin, complex, layered connective tissue with compartmentalized extracellular matrix (ECM) produced by specialized cell types, which directs blood flow in one direction through the heart. There is evidence suggesting remodeling of such ECM during aortic stenosis development. Thus, a better characterization of the role of ECM proteins in this disease would increase our understanding of the underlying molecular mechanisms. Aortic valve samples were collected from 18 patients which underwent aortic valve replacement (50% males, mean age of 74 years) and 18 normal control valves were obtained from necropsies (40% males, mean age of 69 years). The proteome of the samples was analyzed by 2D-LC MS/MS iTRAQ methodology. The results showed an altered expression of 13 ECM proteins of which 3 (biglycan, periostin, prolargin) were validated by Western blotting and/or SRM analyses. These findings are substantiated by our previous results demonstrating differential ECM protein expression. The present study has demonstrated a differential ECM protein pattern in individuals with AS, therefore supporting previous evidence of a dynamic ECM remodeling in human aortic valves during AS development.
| S-EPMC4664895 | biostudies-literature
Project description:Because mouse models play a crucial role in biomedical research related to the human nervous system, understanding the similarities and differences between mouse and human brain is of fundamental importance. Studies comparing transcription in human and mouse have come to varied conclusions, in part because of their relatively small sample sizes or underpowered methodologies. To better characterize gene expression differences between mouse and human, we took a systems-biology approach by using weighted gene coexpression network analysis on more than 1,000 microarrays from brain. We find that global network properties of the brain transcriptome are highly preserved between species. Furthermore, all modules of highly coexpressed genes identified in mouse were identified in human, with those related to conserved cellular functions showing the strongest between-species preservation. Modules corresponding to glial and neuronal cells were sufficiently preserved between mouse and human to permit identification of cross species cell-class marker genes. We also identify several robust human-specific modules, including one strongly correlated with measures of Alzheimer disease progression across multiple data sets, whose hubs are poorly-characterized genes likely involved in Alzheimer disease. We present multiple lines of evidence suggesting links between neurodegenerative disease and glial cell types in human, including human-specific correlation of presenilin-1 with oligodendrocyte markers, and significant enrichment for known neurodegenerative disease genes in microglial modules. Together, this work identifies convergent and divergent pathways in mouse and human, and provides a systematic framework that will be useful for understanding the applicability of mouse models for human brain disorders.
| S-EPMC2906579 | biostudies-literature
Project description:BACKGROUND:Because of their low levels of expression and the inadequacy of current research tools, CB2 cannabinoid receptors (CB2R) have been difficult to study, particularly in the brain. This receptor is especially relevant in the context of neuroinflammation, so novel tools are needed to unveil its pathophysiological role(s). METHODS:We have generated a transgenic mouse model in which the expression of enhanced green fluorescent protein (EGFP) is under the control of the cnr2 gene promoter through the insertion of an Internal Ribosomal Entry Site followed by the EGFP coding region immediately 3' of the cnr2 gene and crossed these mice with mice expressing five familial Alzheimer's disease (AD) mutations (5xFAD). RESULTS:Expression of EGFP in control mice was below the level of detection in all regions of the central nervous system (CNS) that we examined. CB2R-dependent-EGFP expression was detected in the CNS of 3-month-old AD mice in areas of intense inflammation and amyloid deposition; expression was coincident with the appearance of plaques in the cortex, hippocampus, brain stem, and thalamus. The expression of EGFP increased as a function of plaque formation and subsequent microgliosis and was restricted to microglial cells located in close proximity to neuritic plaques. AD mice with CB2R deletion exhibited decreased neuritic plaques with no changes in IL1β expression. CONCLUSIONS:Using a novel reporter mouse line, we found no evidence for CB2R expression in the healthy CNS but clear up-regulation in the context of amyloid-triggered neuroinflammation. Data from CB2R null mice indicate that they play a complex role in the response to plaque formation.
| S-EPMC5968596 | biostudies-literature
Project description:In contrast to the dynamic intracellular environment, structural extracellular matrix (ECM) proteins with half-lives measured in decades, are susceptible to accumulating damage. Whilst conventional approaches such as histology, immunohistochemistry and mass spectrometry are able to identify age- and disease-related changes in protein abundance or distribution, these techniques are poorly suited to characterising molecular damage. We have previously shown that mass spectrometry can detect tissue-specific differences in the proteolytic susceptibility of protein regions within fibrillin-1 and collagen VI alpha-3. Here, we present a novel proteomic approach to detect damage-induced "peptide fingerprints" within complex multi-component ECM assemblies (fibrillin and collagen VI microfibrils) following their exposure to ultraviolet radiation (UVR) by broadband UVB or solar simulated radiation (SSR). These assemblies were chosen because, in chronically photoaged skin, fibrillin and collagen VI microfibril architectures are differentially susceptible to UVR. In this study, atomic force microscopy revealed that fibrillin microfibril ultrastructure was significantly altered by UVR exposure whereas the ultrastructure of collagen VI microfibrils was resistant. UVR-induced molecular damage was further characterised by proteolytic peptide generation with elastase followed by liquid chromatography tandem mass spectrometry (LC-MS/MS). Peptide mapping revealed that UVR exposure increased regional proteolytic susceptibility within the protein structures of fibrillin-1 and collagen VI alpha-3. This allowed the identification of UVR-induced molecular changes within these two key ECM assemblies. Additionally, similar changes were observed within protein regions of co-purifying, microfibril-associated receptors integrins αv and β1. This study demonstrates that LC-MS/MS mapping of peptides enables the characterisation of molecular post-translational damage (via direct irradiation and radiation-induced oxidative mechanisms) within a complex in vitro model system. This peptide fingerprinting approach reliably allows both the identification of UVR-induced molecular damage in and between proteins and the identification of specific protein domains with increased proteolytic susceptibility as a result of photo-denaturation. This has the potential to serve as a sensitive method of identifying accumulated molecular damage in vivo using conventional mass spectrometry data-sets.
| S-EPMC7852314 | biostudies-literature
Project description:Decellularized extracellular matrix (dECM) is a promising material for tissue engineering applications. Tissue-specific dECM is often seen as a favorable material that recapitulates a native-like microenvironment for cellular remodeling. However, the minute quantity of dECM derivable from small organs like the vocal fold (VF) hampers manufacturing scalability. Small intestinal submucosa (SIS), a commercial product with proven regenerative capacity, may be a viable option for VF applications. This study aims to compare dECM hydrogels derived from SIS or VF tissue with respect to protein content and functionality using mass spectrometry-based proteomics and in vitro studies. Proteomic analysis reveals that VF and SIS dECM share 75% of core matrisome proteins. Although VF dECM proteins have greater overlap with native VF, SIS dECM shows less cross-sample variability. Following decellularization, significant reductions of soluble collagen (61%), elastin (81%), and hyaluronan (44%) are noted in VF dECM. SIS dECM contains comparable elastin and hyaluronan but 67% greater soluble collagen than VF dECM. Cells deposit more neo-collagen on SIS than VF-dECM hydrogels, whereas neo-elastin (~50 μg/scaffold) and neo-hyaluronan (~ 6 μg/scaffold) are comparable between the two hydrogels. Overall, SIS dECM possesses reasonably similar proteomic profile and regenerative capacity to VF dECM. SIS dECM is considered a promising alternative for dECM-derived biomaterials for VF regeneration.
| S-EPMC10398787 | biostudies-literature
Project description:The cellular microenvironment is influenced explicitly by the extracellular matrix (ECM), the main tissue support biomaterial, as a decisive factor for tissue growth patterns. The recent emergence of hepatic microphysiological systems (MPS) provide the basic physiological emulation of the human liver for drug screening. However, engineering microfluidic devices with standardized surface coatings of ECM may improve MPS-based organ-specific emulation for improved drug screening. The influence of surface coatings of different ECM types on tissue development needs to be optimized. Additionally, an intensity-based image processing tool and transepithelial electrical resistance (TEER) sensor may assist in the analysis of tissue formation capacity under the influence of different ECM types. The current study highlights the role of ECM coatings for improved tissue formation, implying the additional role of image processing and TEER sensors. We studied hepatic tissue formation under the influence of multiple concentrations of Matrigel, collagen, fibronectin, and poly-L-lysine. Based on experimental data, a mathematical model was developed, and ECM concentrations were validated for better tissue development. TEER sensor and image processing data were used to evaluate the development of a hepatic MPS for human liver physiology modeling. Image analysis data for tissue formation was further strengthened by metabolic quantification of albumin, urea, and cytochrome P450. Standardized ECM type for MPS may improve clinical relevance for modeling hepatic tissue microenvironment, and image processing possibly enhance the tissue analysis of the MPS.
| S-EPMC8434375 | biostudies-literature
Project description:During inflammatory processes the extracellular matrix (ECM) is extensively remodeled, and many of the constituent components are released as proteolytically cleaved fragments. These degradative processes are better documented for inflammatory joint diseases than tendinopathy even though the pathogenesis has many similarities. The aims of this study were to investigate the proteomic composition of injured tendons during early and late disease stages to identify disease-specific cleavage patterns of the ECM protein cartilage oligomeric matrix protein (COMP). In addition to characterizing fragments released in naturally occurring disease, we hypothesized that stimulation of tendon explants with proinflammatory mediators in vitro would induce fragments of COMP analogous to natural disease. Therefore, normal tendon explants were stimulated with IL-1β and prostaglandin E2, and their effects on the release of COMP and its cleavage patterns were characterized. Analyses of injured tendons identified an altered proteomic composition of the ECM at all stages post injury, showing protein fragments that were specific to disease stage. IL-1β enhanced the proteolytic cleavage and release of COMP from tendon explants, whereas PGE2 had no catabolic effect. Of the cleavage fragments identified in early stage tendon disease, two fragments were generated by an IL-1-mediated mechanism. These fragments provide a platform for the development of neo-epitope assays specific to injury stage for tendon disease.
| S-EPMC3931053 | biostudies-literature
Project description:IntroductionExtracellular vesicles (EVs) from human Alzheimer's disease (AD) biospecimens contain amyloid beta (Aβ) peptide and tau. While AD EVs are known to affect brain disease pathobiology, their biochemical and molecular characterizations remain ill defined.MethodsEVs were isolated from the cortical gray matter of 20 AD and 18 control brains. Tau and Aβ levels were measured by immunoassay. Differentially expressed EV proteins were assessed by quantitative proteomics and machine learning.ResultsLevels of pS396 tau and Aβ1-42 were significantly elevated in AD EVs. High levels of neuron- and glia-specific factors are detected in control and AD EVs, respectively. Machine learning identified ANXA5, VGF, GPM6A, and ACTZ in AD EV compared to controls. They distinguished AD EVs from controls in the test sets with 88% accuracy.DiscussionIn addition to Aβ and tau, ANXA5, VGF, GPM6A, and ACTZ are new signature proteins in AD EVs.
| S-EPMC7293582 | biostudies-literature