ABSTRACT
OBJECTIVE: A survival risk assessment model associated with a lung adenocarcinoma (LUAD) microenvironment was established and evaluated to identify effective independent prognostic factors for LUAD. METHODS: The public data were downloaded from the TCGA database, and ESTIMATE prediction software was used to score immune cells and stromal cells for tumor purity prediction. The samples were divided into the high-score group and the low-score group by the median value of the immune score (or stromal score). The Wilcoxon test was used for differential analysis. GO and KEGG enrichment analysis of differentially expressed genes (DEGs) was performed using "clusterProfiler" of R package. Meanwhile, univariate and multivariate regression analysis was performed on DEGs to construct a multivariate Cox risk regression model with variable gene expression levels as independent prognostic factors affecting a tumor microenvironment (TME) and tumor immunity. RESULTS: This study found that LUAD patients with high immune cell (stromal cell) infiltration had better prognosis and were in earlier staging. Functional enrichment analysis revealed that most DEGs were related to the proliferation and activation of immune cells or stromal cells. A survival prediction model composed of 6 TME-related genes (CLEC17A, TAGAP, ABCC8, BCAN, FLT3, and CCR2) was established, and finally, the 6 feature genes closely related to the prognosis of LUAD were proved. The AUC value of the ROC curve in this model was 0.7, indicating that the model was reliable. CONCLUSION: Six genes related to the LUAD microenvironment have a predictive prognostic value in LUAD.
Subject(s)
Adenocarcinoma of Lung/diagnosis , Adenocarcinoma of Lung/genetics , Lung Neoplasms/diagnosis , Lung Neoplasms/genetics , Biomarkers, Tumor/genetics , Biomarkers, Tumor/metabolism , Brevican/genetics , Brevican/metabolism , GTPase-Activating Proteins/genetics , GTPase-Activating Proteins/metabolism , Humans , Lectins, C-Type/genetics , Lectins, C-Type/metabolism , Multivariate Analysis , Prognosis , Proportional Hazards Models , Receptors, CCR2/genetics , Receptors, CCR2/metabolism , Risk Assessment , Sulfonylurea Receptors/genetics , Sulfonylurea Receptors/metabolism , Tumor Microenvironment , fms-Like Tyrosine Kinase 3/genetics , fms-Like Tyrosine Kinase 3/metabolismABSTRACT
The synaptic transmission in the mammalian brain is not limited to the interplay between the pre- and the postsynapse of neurons, but involves also astrocytes as well as extracellular matrix (ECM) molecules. Glycoproteins, proteoglycans and hyaluronic acid of the ECM pervade the pericellular environment and condense to special superstructures termed perineuronal nets (PNN) that surround a subpopulation of CNS neurons. The present study focuses on the analysis of PNNs in a quadruple knockout mouse deficient for the ECM molecules tenascin-C (TnC), tenascin-R (TnR), neurocan and brevican. Here, we analysed the proportion of excitatory and inhibitory synapses and performed electrophysiological recordings of the spontaneous neuronal network activity of hippocampal neurons in vitro. While we found an increase in the number of excitatory synaptic molecules in the quadruple knockout cultures, the number of inhibitory synaptic molecules was significantly reduced. This observation was complemented with an enhancement of the neuronal network activity level. The in vivo analysis of PNNs in the hippocampus of the quadruple knockout mouse revealed a reduction of PNN size and complexity in the CA2 region. In addition, a microarray analysis of the postnatal day (P) 21 hippocampus was performed unravelling an altered gene expression in the quadruple knockout hippocampus.
Subject(s)
Brevican/metabolism , Excitatory Postsynaptic Potentials , Inhibitory Postsynaptic Potentials , Nerve Tissue Proteins/metabolism , Proteoglycans/metabolism , Tenascin/metabolism , Animals , Brevican/genetics , CA2 Region, Hippocampal/metabolism , CA2 Region, Hippocampal/physiology , Cells, Cultured , Female , Gene Deletion , Male , Mice , Nerve Tissue Proteins/genetics , Neurocan , Proteoglycans/genetics , Synapses/metabolism , Synapses/physiology , Tenascin/geneticsABSTRACT
It is a daily challenge for our brains to establish new memories via learning while providing stable storage of remote memories. In the adult vertebrate brain, bimodal regulation of the extracellular matrix (ECM) may regulate the delicate balance of learning-dependent plasticity and stable memory formation. Here, we trained adult male mice in a cortex-dependent auditory discrimination task and measured the abundance of ECM proteins brevican (BCN) and tenascin-R over the course of acquisition learning, consolidation, and long-term recall in two learning-relevant brain regions; the auditory cortex and hippocampus. Although early training led to a general downregulation of total ECM proteins, successful retrieval correlated with a region-specific and transient upregulation of BCN levels in the auditory cortex. No other parameter such as arousal or stress could account for the transient and region-specific BCN upregulation. This performance-dependent biphasic regulation of the ECM may assist transient plasticity to facilitate initial learning and subsequently promote the long-term consolidation of memory.SIGNIFICANCE STATEMENT The capacity to learn throughout life and at the same time guarantee lifelong storage and remote recall of established memories is a daily challenge. Emerging evidence suggests an important function of the extracellular matrix (ECM), a conglomerate of secreted proteins and polysaccharides in the adult vertebrate brain. We trained mice in an auditory long-term memory task and measured learning-related dynamic changes of the ECM protein brevican. Specifically, in the auditory cortex brevican is downregulated during initial learning and subsequently upregulated in exclusively those animals that have learned the task, suggesting a performance-dependent regulation in the service of memory consolidation and storage. Our data may provide novel therapeutic implications for several neuropsychiatric diseases involving dysregulation of the ECM.
Subject(s)
Auditory Cortex/metabolism , Brevican/genetics , Memory Consolidation , Animals , Auditory Cortex/physiology , Auditory Perception , Brevican/metabolism , Discrimination, Psychological , Hippocampus/metabolism , Hippocampus/physiology , Male , Mice , Mice, Inbred C57BL , Up-RegulationABSTRACT
BACKGROUND: Perineuronal nets (PNNs) are specialized aggregations of extracellular matrix (ECM) molecules surrounding specific neurons in the central nervous system (CNS). PNNs are supposed to control synaptic transmission and are frequently associated with neurons firing at high rates, including principal neurons of auditory brainstem nuclei. The origin of high-frequency activity of auditory brainstem neurons is the indefatigable sound-driven transmitter release of inner hair cells (IHCs) in the cochlea. RESULTS: Here, we show that synaptic poles of IHCs are ensheathed by basket-like ECM complexes formed by the same molecules that constitute PNNs of neurons in the CNS, including brevican, aggreccan, neurocan, hyaluronan, and proteoglycan link proteins 1 and 4 and tenascin-R. Genetic deletion of brevican, one of the main components, resulted in a massive degradation of ECM baskets at IHCs, a significant impairment in spatial coupling of pre- and postsynaptic elements and mild impairment of hearing. CONCLUSIONS: These ECM baskets potentially contribute to control of synaptic transmission at IHCs and might be functionally related to PNNs of neurons in the CNS.
Subject(s)
Brevican/genetics , Ear, Inner/physiology , Extracellular Matrix/metabolism , Synaptic Transmission/physiology , Animals , Brevican/metabolism , Female , Male , Mice , Mice, KnockoutABSTRACT
To accurately recapitulate the heterogeneity of human diseases, animal models require to recreate multiple complex genetic alterations. Here, we combine the RCAS-TVA system with the CRISPR-Cas9 genome editing tools for precise modeling of human tumors. We show that somatic deletion in neural stem cells of a variety of known tumor suppressor genes (Trp53, Cdkn2a, and Pten) leads to high-grade glioma formation. Moreover, by simultaneous delivery of pairs of guide RNAs we generate different gene fusions with oncogenic potential, either by chromosomal deletion (Bcan-Ntrk1) or by chromosomal translocation (Myb-Qk). Lastly, using homology-directed-repair, we also produce tumors carrying the homologous mutation to human BRAF V600E, frequently identified in a variety of tumors, including different types of gliomas. In summary, we have developed an extremely versatile mouse model for in vivo somatic genome editing, that will elicit the generation of more accurate cancer models particularly appropriate for pre-clinical testing.
Subject(s)
Brain Neoplasms/genetics , CRISPR-Cas Systems , Gene Editing , RNA, Guide, Kinetoplastida/genetics , Animals , Antigens, Neoplasm/genetics , Benzamides/pharmacology , Brain Neoplasms/drug therapy , Brevican/genetics , DNA Repair , False Positive Reactions , Gene Frequency , Gene Transfer Techniques , Glioma/metabolism , Humans , In Situ Hybridization, Fluorescence , Indazoles/pharmacology , Mice , Mice, SCID , Mice, Transgenic , Mutation , NIH 3T3 Cells , Receptor, trkA/geneticsABSTRACT
The widespread application of high-throughput sequencing methods is resulting in the identification of a rapidly growing number of novel gene fusions caused by tumour-specific chromosomal rearrangements, whose oncogenic potential remains unknown. Here we describe a strategy that builds upon recent advances in genome editing and combines ex vivo and in vivo chromosomal engineering to rapidly and effectively interrogate the oncogenic potential of genomic rearrangements identified in human brain cancers. We show that one such rearrangement, an microdeletion resulting in a fusion between Brevican (BCAN) and Neurotrophic Receptor Tyrosine Kinase 1 (NTRK1), is a potent oncogenic driver of high-grade gliomas and confers sensitivity to the experimental TRK inhibitor entrectinib. This work demonstrates that BCAN-NTRK1 is a bona fide human glioma driver and describes a general strategy to define the oncogenic potential of novel glioma-associated genomic rearrangements and to generate accurate preclinical models of this lethal human cancer.
Subject(s)
Benzamides/therapeutic use , Brevican/genetics , Glioma/genetics , Indazoles/therapeutic use , Oncogene Fusion , Receptor, trkA/genetics , Animals , Benzamides/pharmacology , CRISPR-Cas Systems , Drug Screening Assays, Antitumor , Feasibility Studies , Female , Gene Editing , Glioma/drug therapy , Humans , Indazoles/pharmacology , Mice, Inbred C57BL , Mice, Nude , Neoplasms, Experimental , Primary Cell Culture , Receptor, trkA/antagonists & inhibitorsABSTRACT
Hippocampal synaptic plasticity comprises a key cellular mechanism for information storage. In the hippocampus, both long-term potentiation (LTP) and long-term depression (LTD) are triggered by synaptic Ca2+ -elevations that are typically mediated by the opening of voltage-gated cation channels, such as N-methyl-d-aspartate receptors (NMDAR), in the postsynaptic density. The integrity of the post-synaptic density is ensured by the extracellular matrix (ECM). Here, we explored whether synaptic plasticity is affected in adult behaving mice that lack the ECM proteins brevican, neurocan, tenascin-C, and tenascin-R (KO). We observed that the profiles of synaptic potentiation and depression in the dentate gyrus (DG) were profoundly altered compared to plasticity profiles in wild-type littermates (WT). Specifically, synaptic depression was amplified in a frequency-dependent manner and although late-LTP (>24 hr) was expressed following strong afferent tetanization, the early component of LTP (<75 min post-tetanization) was absent. LTP (>4 hr) elicited by weaker tetanization was equivalent in WT and KO animals. Furthermore, this latter form of LTP was NMDAR-dependent in WT but not KO mice. Scrutiny of DG receptor expression revealed significantly lower levels of both the GluN2A and GluN2B subunits of the N-methyl-d-aspartate receptor, of the metabotropic glutamate receptor, mGlu5 and of the L-type calcium channel, Cav 1.3 in KO compared to WT animals. Homer 1a and of the P/Q-type calcium channel, Cav 1.2 were unchanged in KO mice. Taken together, findings suggest that in mice that lack multiple ECM proteins, synaptic plasticity is intact, but is fundamentally different.
Subject(s)
Extracellular Matrix/genetics , Extracellular Matrix/metabolism , Hippocampus/physiology , Neuronal Plasticity/physiology , Animals , Animals, Newborn , Brevican/genetics , Brevican/metabolism , Calcium Channels, L-Type/genetics , Calcium Channels, L-Type/metabolism , Electric Stimulation , Excitatory Amino Acid Antagonists/pharmacology , Male , Mice , Mice, Transgenic , Neurocan/genetics , Neurocan/metabolism , Neuronal Plasticity/genetics , Patch-Clamp Techniques , Piperazines/pharmacology , Receptors, N-Methyl-D-Aspartate/genetics , Receptors, N-Methyl-D-Aspartate/metabolism , Tenascin/genetics , Tenascin/metabolism , WakefulnessABSTRACT
Retinal ischemia occurs in a variety of eye diseases. Restrained blood flow induces retinal damage, which leads to progressive optic nerve degeneration and vision loss. Previous studies indicate that extracellular matrix (ECM) constituents play an important role in complex tissues, such as retina and optic nerve. They have great impact on de- and regeneration processes and represent major candidates of central nervous system glial scar formation. Nevertheless, the importance of the ECM during ischemic retina and optic nerve neurodegeneration is not fully understood yet. In this study, we analyzed remodeling of the extracellular glycoproteins fibronectin, laminin, tenascin-C and tenascin-R and the chondroitin sulfate proteoglycans (CSPGs) aggrecan, brevican and phosphacan/RPTPß/ζ in retinae and optic nerves of an ischemia/reperfusion rat model via quantitative real-time PCR, immunohistochemistry and Western blot. A variety of ECM constituents were dysregulated in the retina and optic nerve after ischemia. Regarding fibronectin, significantly elevated mRNA and protein levels were observed in the retina following ischemia, while laminin and tenascin-C showed enhanced immunoreactivity in the optic nerve after ischemia. Interestingly, CSPGs displayed significantly increased expression levels in the optic nerve. Our study demonstrates a dynamic expression of ECM molecules following retinal ischemia, which strengthens their regulatory role during neurodegeneration.
Subject(s)
Gene Expression Regulation , Optic Nerve/metabolism , Reperfusion Injury/genetics , Retina/metabolism , Retinal Degeneration/genetics , Aggrecans/genetics , Aggrecans/metabolism , Animals , Brevican/genetics , Brevican/metabolism , Disease Models, Animal , Fibronectins/genetics , Fibronectins/metabolism , Laminin/genetics , Laminin/metabolism , Male , Optic Nerve/blood supply , Optic Nerve/pathology , RNA, Messenger/genetics , RNA, Messenger/metabolism , Rats , Receptor-Like Protein Tyrosine Phosphatases, Class 5/genetics , Receptor-Like Protein Tyrosine Phosphatases, Class 5/metabolism , Reperfusion Injury/metabolism , Reperfusion Injury/pathology , Retina/pathology , Retinal Degeneration/metabolism , Retinal Degeneration/pathology , Signal Transduction , Tenascin/genetics , Tenascin/metabolismABSTRACT
OBJECTIVE: The primary focus of this study was to investigate the effects of local profound hypothermia and to explore the possible mechanism in adult rats with spinal cord injury. STUDY DESIGN AND METHODS: Spinal cord injury models were established by placing aneurysm clips on T10. An epidural perfusion device was applied to maintain a steady temperature (18â °C) for 120â min with gradual rewarming to 37â °C Total hypothermic duration lasted up to about 170â min. The expression of axon regeneration inhibitors was tested by Western blot and real-time PCR. Luxol Fast Blue (LFB) stain and Bielschowsky silver stain were used to observe spinal cord morphology. Motor function of the hind limbs (BBB score) was monitored for 21 days. RESULTS: The expressions of RhoA, ROCK-II, NG2, Neurocan, Brevican, and Nogo-A were downregulated by regional hypothermia (RH) after spinal cord injury. Subsequent observation showed that rats that had received RH had an alleviated demyelinating condition and a greater number of nerve fibers. Furthermore, the RH group achieved higher BBB scores than the spinal cord injury (SCI) group. CONCLUSIONS: Recovery of hind limb function in rats can be promoted by local profound hypothermia; this may be caused by the suppression of axon regeneration inhibitors.
Subject(s)
Hypothermia, Induced , Nerve Regeneration , Spinal Cord Injuries/therapy , Animals , Axons/metabolism , Axons/physiology , Brevican/genetics , Brevican/metabolism , Down-Regulation , Hindlimb/innervation , Hindlimb/physiology , Male , Neurocan/genetics , Neurocan/metabolism , Nogo Proteins/genetics , Nogo Proteins/metabolism , Rats , Rats, Sprague-Dawley , Recovery of Function , Spinal Cord Injuries/rehabilitation , rho-Associated Kinases/genetics , rho-Associated Kinases/metabolism , rhoA GTP-Binding Protein/genetics , rhoA GTP-Binding Protein/metabolismABSTRACT
Cocaine-associated environmental cues sustain relapse vulnerability by reactivating long-lasting memories of cocaine reward. During periods of abstinence, responding to cocaine cues can time-dependently intensify a phenomenon referred to as 'incubation of cocaine craving'. Here, we investigated the role of the extracellular matrix protein brevican in recent (1 day after training) and remote (3 weeks after training) expression of cocaine conditioned place preference (CPP). Wild-type and Brevican heterozygous knock-out mice, which express brevican at ~50% of wild-type levels, received three cocaine-context pairings using a relatively low dose of cocaine (5 mg/kg). In a drug-free CPP test, heterozygous mice showed enhanced preference for the cocaine-associated context at the remote time point compared with the recent time point. This progressive increase was not observed in wild-type mice and it did not generalize to contextual-fear memory. Virally mediated overexpression of brevican levels in the hippocampus, but not medial prefrontal cortex, of heterozygous mice prevented the progressive increase in cocaine CPP, but only when overexpression was induced before conditioning. Post-conditioning overexpression of brevican did not affect remote cocaine CPP, suggesting that brevican limited the increase in remote CPP by altering neuro-adaptive mechanisms during cocaine conditioning. We provide causal evidence that hippocampal brevican levels control time-dependent enhancement of cocaine CPP during abstinence, pointing to a novel substrate that regulates incubation of responding to cocaine-associated cues.
Subject(s)
Anesthetics, Local/pharmacokinetics , Brevican/metabolism , Cocaine/pharmacology , Conditioning, Operant/drug effects , Gene Expression Regulation/drug effects , Analysis of Variance , Animals , Brevican/genetics , Fear/drug effects , Gene Expression Regulation/genetics , Green Fluorescent Proteins/genetics , Green Fluorescent Proteins/metabolism , Maze Learning/drug effects , Mice , Mice, Inbred C57BL , Mice, Transgenic , Prefrontal Cortex/drug effects , Prefrontal Cortex/metabolism , Tenascin/metabolism , Time Factors , Transduction, GeneticABSTRACT
KEY POINTS: The proteoglycan brevican is a major component of the extracellular matrix of perineuronal nets and is highly enriched in the perisynaptic space suggesting a role for synaptic transmission. We have introduced the calyx of Held in the auditory brainstem as a model system to study the impact of brevican on dynamics and reliability of synaptic transmission. In vivo extracellular single-unit recordings at the calyx of Held in brevican-deficient mice yielded a significant increase in the action potential (AP) transmission delay and a prolongation of pre- and postsynaptic APs. The changes in dynamics of signal transmission were accompanied by the reduction of presynaptic vGlut1 and ultrastructural changes in the perisynaptic space. These data show that brevican is an important mediator of fast synaptic transmission at the calyx of Held. ABSTRACT: The extracellular matrix is an integral part of the neural tissue. Its most conspicuous manifestation in the brain are the perineuronal nets (PNs) which surround somata and proximal dendrites of distinct neuron types. The chondroitin sulfate proteoglycan brevican is a major component of PNs. In contrast to other PN-comprising proteoglycans (e.g. aggrecan and neurocan), brevican is mainly expressed in the perisynaptic space closely associated with both the pre- and postsynaptic membrane. This specific localization prompted the hypothesis that brevican might play a role in synaptic transmission. In the present study we specifically investigated the role of brevican in synaptic transmission at a central synapse, the calyx of Held in the medial nucleus of the trapezoid body, by the use of in vivo electrophysiology, immunohistochemistry, biochemistry and electron microscopy. In vivo extracellular single-unit recordings were acquired in brevican-deficient mice and the dynamics and reliability of synaptic transmission were compared to wild-type littermates. In knockout mice, the speed of pre-to-postsynaptic action potential (AP) transmission was reduced and the duration of the respective pre- and postsynaptic APs increased. The reliability of signal transmission, however, was not affected by the lack of brevican. The changes in dynamics of signal transmission were accompanied by the reduction of (i) presynaptic vGlut1 and (ii) the size of subsynaptic cavities. The present results suggest an essential role of brevican for the functionality of high-speed synaptic transmission at the calyx of Held.
Subject(s)
Brevican/physiology , Synaptic Transmission/physiology , Trapezoid Body/physiology , Acoustic Stimulation , Action Potentials , Animals , Brevican/genetics , Excitatory Amino Acid Transporter 2/metabolism , Extracellular Matrix , Female , Glucose Transporter Type 1/metabolism , Glucose Transporter Type 2/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Synapses/physiology , Trapezoid Body/metabolismABSTRACT
Growing evidence supports the important role of the tumor microenvironment (TME) in cancer biology. A defining aspect of the glioma TME is the unique composition and structure of its extracellular matrix (ECM), which enables tumor cells to overcome the inhibitory barriers of the adult central nervous system (CNS). In this way, the TME plays a role in glioma invasion and the cellular heterogeneity that distinguishes these tumors. Brain Enriched Hyaluronan Binding (BEHAB)/brevican (B/b), is a CNS-specific ECM constituent and is upregulated in the glioma TME. Previous studies have shown B/b exerts a pro-invasive function, suggesting it may represent a target to reduce glioma pathogenesis. Herein, we also provide evidence that B/b expression is enriched in the glioma initiating cell (GIC) niche. We demonstrate that B/b plays roles in the pathological progression, aggressiveness, and lethality of tumors derived from human GICs and traditional glioma cell lines. Interestingly, we found that B/b is not required to maintain the defining phenotypic properties of GICs and thereby acts primarily in late stages of glioma progression. This study suggests that the increased expression of B/b in the TME is a valuable therapeutic target for glioma.
Subject(s)
Brain Neoplasms/pathology , Brain/pathology , Brevican/antagonists & inhibitors , Glioma/pathology , Neoplastic Stem Cells/pathology , Adult , Animals , Brain Neoplasms/metabolism , Brain Neoplasms/mortality , Brevican/genetics , Brevican/metabolism , Cell Differentiation , Cell Survival , Extracellular Matrix , Female , Glioma/metabolism , Glioma/mortality , Humans , Neoplastic Stem Cells/metabolism , Prognosis , RNA, Small Interfering/genetics , Rats , Rats, Inbred Lew , Survival Rate , Tumor Cells, CulturedABSTRACT
Glioblastoma multiforme (GBM) is the most aggressive form of brain tumor, yet with no targeted therapy with substantial survival benefit. Recent studies on solid tumors showed that fusion genes often play driver roles and are promising targets for pharmaceutical intervention. To survey potential fusion genes in GBMs, we analysed RNA-Seq data from 162 GBM patients available through The Cancer Genome Atlas (TCGA), and found that 3' exons of neurotrophic tyrosine kinase receptor type 1 (NTRK1, encoding TrkA) are fused to 5' exons of the genes that are highly expressed in neuronal tissues, neurofascin (NFASC) and brevican (BCAN). The fusions preserved both the transmembrane and kinase domains of NTRK1 in frame. NTRK1 is a mediator of the pro-survival signaling of nerve growth factor (NGF) and is a known oncogene, found commonly altered in human cancer. While GBMs largely lacked NTRK1 expression, the fusion-positive GBMs expressed fusion transcripts in high abundance, and showed elevated NTRK1-pathway activity. Lentiviral transduction of the NFASC-NTRK1 fusion gene in NIH 3T3 cells increased proliferation in vitro, colony formation in soft agar, and tumor formation in mice, suggesting the possibility that the fusion contributed to the initiation or maintenance of the fusion-positive GBMs, and therefore may be a rational drug target.
Subject(s)
Brain Neoplasms/metabolism , Brevican/metabolism , Cell Adhesion Molecules/metabolism , Glioblastoma/metabolism , Oncogene Proteins, Fusion/metabolism , Receptor, trkA/metabolism , Amino Acid Sequence , Animals , Base Sequence , Brevican/chemistry , Brevican/genetics , Carcinogenesis/pathology , Cell Adhesion Molecules/chemistry , Cell Adhesion Molecules/genetics , Male , Mice , Mice, Inbred BALB C , Mice, Nude , Molecular Sequence Data , NIH 3T3 Cells , Nerve Growth Factors/chemistry , Nerve Growth Factors/genetics , Nerve Growth Factors/metabolism , Oncogene Proteins, Fusion/chemistry , Oncogene Proteins, Fusion/genetics , Receptor, trkA/chemistry , Receptor, trkA/geneticsABSTRACT
In Alzheimer's disease (AD), different types of neurons and different brain areas show differential patterns of vulnerability towards neurofibrillary degeneration, which provides the basis for a highly predictive profile of disease progression throughout the brain that now is widely accepted for neuropathological staging. In previous studies we could demonstrate that in AD cortical and subcortical neurons are constantly less frequently affected by neurofibrillary degeneration if they are enwrapped by a specialized form of the hyaluronan-based extracellular matrix (ECM), the so called 'perineuronal net' (PN). PNs are basically composed of large aggregating chondroitin sulphate proteoglycans connected to a hyaluronan backbone, stabilized by link proteins and cross-linked via tenascin-R (TN-R). Under experimental conditions in mice, PN-ensheathed neurons are better protected against iron-induced neurodegeneration than neurons without PN. Still, it remains unclear whether these neuroprotective effects are directly mediated by the PNs or are associated with some other mechanism in these neurons unrelated to PNs. To identify molecular components that essentially mediate the neuroprotective aspect on PN-ensheathed neurons, we comparatively analysed neuronal degeneration induced by a single injection of FeCl3 on four different mice knockout strains, each being deficient for a different component of PNs. Aggrecan, link protein and TN-R were identified to be essential for the neuroprotective properties of PN, whereas the contribution of brevican was negligible. Our findings indicate that the protection of PN-ensheathed neurons is directly mediated by the net structure and that both the high negative charge and the correct interaction of net components are essential for their neuroprotective function.
Subject(s)
Aggrecans/metabolism , Brain/drug effects , Chlorides/toxicity , Extracellular Matrix Proteins/metabolism , Extracellular Matrix/metabolism , Ferric Compounds/toxicity , Neurons/drug effects , Oxidative Stress/drug effects , Proteoglycans/metabolism , Tenascin/metabolism , Aggrecans/deficiency , Aggrecans/genetics , Animals , Brain/metabolism , Brain/pathology , Brevican/deficiency , Brevican/genetics , Chondroitin Sulfates/metabolism , Extracellular Matrix Proteins/deficiency , Extracellular Matrix Proteins/genetics , Female , Genotype , Hyaluronic Acid/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Nerve Degeneration , Neurons/metabolism , Neurons/pathology , Phenotype , Proteoglycans/deficiency , Proteoglycans/genetics , Tenascin/deficiency , Tenascin/geneticsABSTRACT
Aberrant DNA methylation constitutes a well-established epigenetic marker for breast cancer. Changes in methylation early in cancer development may be clinically relevant for cancer detection and prognosis-based therapeutic decisions. In the present study, a combination of methyl-CpG immunoprecipitation (MCIp) and human CpG island (CGI) arrays was applied to compare genome-wide DNA methylation profiles in 10 low-grade in situ and invasive breast cancers against 10 normal breast samples. In total, 214 CGIs were found to be hypermethylated in ≥6 of 10 tumors. Functional term enrichment analyses revealed an overrepresentation of homeobox genes and genes involved in transcription and regulation of transcription. Significant hypermethylation of 11 selected genes in tumor vs. normal tissue was validated in two independent sample sets (45 tumors and 11 controls, 43 tumors and 8 controls) using quantitative EpiTyper technology. In tumors, median methylation levels of BCAN, HOXD1, KCTD8, KLF11, NXPH1, POU4F1, SIM1, and TCF7L1 were ≥30% higher than in normal samples, representing potential biomarkers for tumor diagnosis. Using the 90th percentile of methylation levels in normal tissue as cutoff value, 62-92% of in situ samples (n=13), 72-97% of invasive samples from the first validation set (n=32), and 86-100% of invasive samples from the second validation set (n=43) were classified as hypermethylated. Hypermethylation of KLF11 and SIM1 might also be associated with increased risk of developing metastases. In summary, early methylation changes are frequent in the low-grade pathway of breast cancer and may be useful in the development of differential diagnostic and possibly also prognostic markers.
Subject(s)
Biomarkers, Tumor/genetics , Breast Neoplasms/genetics , DNA Methylation , Genome-Wide Association Study/methods , Adult , Aged , Aged, 80 and over , Apoptosis Regulatory Proteins , Basic Helix-Loop-Helix Transcription Factors/genetics , Breast Neoplasms/pathology , Brevican/genetics , Cell Cycle Proteins/genetics , Female , Gene Expression Profiling , Gene Expression Regulation, Neoplastic , Glycoproteins/genetics , Homeodomain Proteins/genetics , Humans , MCF-7 Cells , Middle Aged , Neoplasm Grading , Neoplasm Staging , Neuropeptides/genetics , Repressor Proteins/genetics , Transcription Factor 7-Like 1 Protein/genetics , Transcription Factor Brn-3A/geneticsABSTRACT
Brevican is a neural proteoglycan implicated in a multitude of physiological and pathophysiological plasticity processes in the brain. It localizes to neuronal surfaces and contributes to the formation of specific types of extracellular matrix like the perineuronal nets or the perisynaptic or axon initial segment-based matrix in mature neuronal tissue. Via a variable degree of chondroitin sulfate attachment, limited proteolytic cleavage by matrix metalloproteinases, differential splicing and Ca(2+)-dependent binding to interaction partners it acts as a regulator in synaptic plasticity, glioma invasion, post-lesion plasticity or Alzheimer's disease. This review briefly summarizes its gene and protein structure, biochemical interactions and neurobiological functions.
Subject(s)
Brain/metabolism , Brevican/metabolism , Proteoglycans/metabolism , Animals , Brevican/genetics , Brevican/pharmacology , Extracellular Matrix/metabolism , Humans , MiceABSTRACT
Episodic falling syndrome (EFS) is a canine paroxysmal hypertonicity disorder found in Cavalier King Charles spaniels. Episodes are triggered by exercise, stress or excitement and characterized by progressive hypertonicity throughout the thoracic and pelvic limbs, resulting in a characteristic 'deer-stalking' position and/or collapse. We used a genome-wide association strategy to map the EFS locus to a 3.48 Mb critical interval on canine chromosome 7. By prioritizing candidate genes on the basis of biological plausibility, we found that a 15.7 kb deletion in BCAN, encoding the brain-specific extracellular matrix proteoglycan brevican, is associated with EFS. This represents a compelling causal mutation for EFS, since brevican has an essential role in the formation of perineuronal nets governing synapse stability and nerve conduction velocity. Mapping of the deletion breakpoint enabled the development of Multiplex PCR and Multiplex Ligation-dependent Probe Amplification (MLPA) genotyping tests that can accurately distinguish normal, carrier and affected animals. Wider testing of a larger population of CKCS dogs without a history of EFS from the USA revealed that carriers are extremely common (12.9%). The development of molecular genetic tests for the EFS microdeletion will allow the implementation of directed breeding programs aimed at minimizing the number of animals with EFS and enable confirmatory diagnosis and pharmacotherapy of affected dogs.