Microvesicles: Novel Biomarkers for Neurological Disorders

Microvesicles (MVs) are released by most cell types in physiological conditions, but their number is often increased upon cellular activation or neoplastic transformation. This suggests that their detection may be helpful in pathological conditions to have information on activated cell types and, possibly, on the nature of the activation. This could be of paramount importance in districts and tissues that are not accessible to direct examination, such as the central nervous system. Increased release of MVs has been described to be associated to the acute or active phase of several neurological disorders. While the subcellular origin of MVs (exosome or ectosomes) is basically never addressed in these studies because of technical limitations, the cell of origin is always identified. Endothelium- or platelet-derived MVs, detected in plasma or serum, are linked to neurological pathologies with a vascular or ischemic pathogenic component, and may represent a very useful marker to support therapeutic choices in stroke. In neuroinflammatory disorders, such as multiple sclerosis, MVs of oligodendroglial, or microglial origin have been described in the cerebrospinal fluid and may carry, in perspective, additional information on the biological alterations in their cell of origin. Little specific evidence is available in neurodegenerative disorders and, specifically, MVs of neural origin have never been investigated in these pathologies. Few data have been reported for neuroinfection and brain trauma. In brain tumors, despite the limited number of studies performed, results are very promising and potentially close to clinical translation. We here review all currently available data on the detection of MVs in neurological diseases, limiting our search to exclusively human studies. Current literature and our own data indicate that MVs detection may represent a very promising strategy to gain pathogenic information, identify therapeutic targets, and select specific biomarkers for neurological disorders

Role of extracellular vesicles in early synaptic dysfunction in AD

AbstractBackgroundAlzheimer's disease (AD) is a progressive neurodegenerative disorder associated with amyloid‐β (Aβ) and tau protein accumulation. Synaptic dysfunction is an early mechanism in AD which involves progressively larger areas of the brain over time. However how synaptic dysfunction starts and propagates is unknown. The hypothesis we are testing is that extracellular vesicles (EVs) released by microglia exposed to and carrying Aβ42 (Aβ‐EVs) may be responsible for these early events in AD.MethodCombining optical manipulation and time lapse imaging to place single EVs on RFP‐positive cultured neuron dendrites and test their effects on the synapse, we show that Aβ‐EVs rapidly alter dendritic spine morphology (a structural correlate of synaptic strength) locally at the site of interaction.ResultAβ‐EVs induce a significant increase in the density of immature protrusions around the contact site (60 µm).ConclusionEmploying the same methodologies, we are currently monitoring Aβ‐EV‐neuron dynamics at the neuronal surface, to test whether Aβ‐EVs may propagate dendritic spine alterations to adjacent regions over time, contributing to the spreading of synaptic deficits. Our data provide evidence of the involvement of microglial EVs in early synaptic dysfunction in AD, paving the way for novel therapeutic strategies

The histone demethylase PHF8 regulates astrocyte differentiation and function

Astrocyte differentiation; Chromatin transcription; SynapseDiferenciación de astrocitos; Transcripción de la cromatina; SinapsisDiferenciació d'astròcits; Transcripció de la cromatina; SinapsiEpigenetic factors have been shown to play a crucial role in X-linked intellectual disability (XLID). Here, we investigate the contribution of the XLID-associated histone demethylase PHF8 to astrocyte differentiation and function. Using genome-wide analyses and biochemical assays in mouse astrocytic cultures, we reveal a regulatory crosstalk between PHF8 and the Notch signaling pathway that balances the expression of the master astrocytic gene Nfia. Moreover, PHF8 regulates key synaptic genes in astrocytes by maintaining low levels of H4K20me3. Accordingly, astrocytic-PHF8 depletion has a striking effect on neuronal synapse formation and maturation in vitro. These data reveal that PHF8 is crucial in astrocyte development to maintain chromatin homeostasis and limit heterochromatin formation at synaptogenic genes. Our studies provide insights into the involvement of epigenetics in intellectual disability.This study was supported by grants from the Spanish Ministry of Economy (Ministerio de Economía, Industria y Competitividad, Gobierno de España; BFU2015-69248-P and PGC2018-096082-B-I00 to M.A.M.-B.) and Ministerio de Ciencia e Innovación (PID2019-111217RB-I00 to X.C) and a Travelling Fellowship from Boehringer Ingelheim to S.I. Open access funding provided by Consejo Superior de Investigaciones Científicas. Deposited in PMC for immediate release

TRPV1 channels are critical brain inflammation detectors and neuropathic pain biomarkers in mice

The capsaicin receptor TRPV1 has been widely characterized in the sensory system as a key component of pain and inflammation. A large amount of evidence shows that TRPV1 is also functional in the brain although its role is still debated. Here we report that TRPV1 is highly expressed in microglial cells rather than neurons of the anterior cingulate cortex and other brain areas. We found that stimulation of microglial TRPV1 controls cortical microglia activation per se and indirectly enhances glutamatergic transmission in neurons by promoting extracellular microglial microvesicles shedding. Conversely, in the cortex of mice suffering from neuropathic pain, TRPV1 is also present in neurons affecting their intrinsic electrical properties and synaptic strength. Altogether, these findings identify brain TRPV1 as potential detector of harmful stimuli and a key player of microglia to neuron communication

a new approach to follow a single extracellular vesicle cell interaction using optical tweezers

Extracellular vesicles (EVs) are spherical membrane structures released by most cells. These highly conserved mediators of intercellular communication carry proteins, lipids, and nucleic acids, and..

Microglial extracellular vesicles induce Alzheimer’s diseaserelated cortico-hippocampal network dysfunction.

β-Amyloid is one of the main pathological hallmarks of Alzheimer’s disease and plays a major role in synaptic dysfunction. It has been demonstrated that β-amyloid can elicit aberrant excitatory activity in cortical-hippocampal networks, which is associated with behavioural abnormalities. However, the mechanism of the spreading of β-amyloid action within a specific circuitry has not been elucidated yet. We have previously demonstrated that the motion of microglia-derived large extracellular vesicles carrying β-amyloid, at the neuronal surface, is crucial for the initiation and propagation of synaptic dysfunction along the entorhinal–hippocampal circuit. Here, using chronic EEG recordings, we show that a single injection of extracellular vesicles carrying β-amyloid into the mouse entorhinal cortex could trigger alterations in the cortical and hippocampal activity that are reminiscent of those found in Alzheimer’s disease mouse models and human patients. The development of EEG abnormalities was associated with progressive memory impairment as assessed by an associative (object-place context recognition) and non-associative (object recognition) task. Importantly, when the motility of extracellular vesicles, carrying β-amyloid, was inhibited, the effect on network stability and memory function was significantly reduced. Our model proposes a new biological mechanism based on the extracellular vesicles–mediated progression of β-amyloid pathology and offers the opportunity to test pharmacological treatments targeting the early stages of Alzheimer’s disease

Acid sphingomyelinase activity triggers microparticle release from glial cells

We have earlier shown that microglia, the immune cells of the CNS, release microparticles from cell plasma membrane after ATP stimulation. These vesicles contain and release IL-1β, a crucial cytokine in CNS inflammatory events. In this study, we show that microparticles are also released by astrocytes and we get insights into the mechanism of their shedding. We show that, on activation of the ATP receptor P2X7, microparticle shedding is associated with rapid activation of acid sphingomyelinase, which moves to plasma membrane outer leaflet. ATP-induced shedding and IL-1β release are markedly reduced by the inhibition of acid sphingomyelinase, and completely blocked in glial cultures from acid sphingomyelinase knockout mice. We also show that p38 MAPK cascade is relevant for the whole process, as specific kinase inhibitors strongly reduce acid sphingomyelinase activation, microparticle shedding and IL-1β release. Our results represent the first demonstration that activation of acid sphingomyelinase is necessary and sufficient for microparticle release from glial cells and define key molecular effectors of microparticle formation and IL-1β release, thus, opening new strategies for the treatment of neuroinflammatory diseases

Ectonucleotidase activity and immunosuppression in astrocyte-CD4 T cell bidirectional signaling

Astrocytes play a crucial role in neuroinflammation as part of the glia limitans, which regulates infiltration of the brain parenchyma by leukocytes. The signaling pathways and molecular events, which result from the interaction of activated T cells with astrocytes are poorly defined. Here we show that astrocytes promote the expression and enzymatic activity of CD39 and CD73 ectonucleotidases in recently activated CD4 cells by a contact dependent mechanism that is independent of T cell receptor interaction with class II major histocompatibility complex (MHC). Transforming growth factor-β (TGF-β) is robustly upregulated and sufficient to promote ectonucleotidases expression. T cell adhesion to astrocyte results in differentiation to an immunosuppressive phenotype defined by expression of the transcription factor Rorγt, which characterizes the CD4 T helper 17 subset. CD39 activity in T cells in turn inhibits spontaneous calcium oscillations in astrocytes that correlated with enhanced and reduced transcription of CCL2 chemokine and Sonic hedgehog (Shh), respectively. We hypothesize this TCR-independent interaction promote an immunosuppressive program in T cells to control possible brain injury by deregulated T cell activation during neuroinflammation. On the other hand, the increased secretion of CCL2 with concomitant reduction of Shh might promote leukocytes extravasation into the brain parenchyma

Small extracellular vesicles released from germinated kiwi pollen (pollensomes) present characteristics similar to mammalian exosomes and carry a plant homolog of ALIX

Introduction: In the last decade, it has been discovered that allergen-bearing extracellular nanovesicles, termed “pollensomes”, are released by pollen during germination. These extracellular vesicles (EVs) may play an important role in pollen-pistil interaction during fertilization, stabilizing the secreted bioactive molecules and allowing long-distance signaling. However, the molecular composition and the biological role of these EVs are still unclear. The present study had two main aims: (I) to clarify whether pollen germination is needed to release pollensomes, or if they can be secreted also in high humidity conditions; and (II) to investigate the molecular features of pollensomes following the most recent guidelines for EVs isolation and identification. Methods: To do so, pollensomes were isolated from hydrated and germinated kiwi (Actinidia chinensis Planch.) pollen, and characterized using imaging techniques, immunoblotting, and proteomics. Results: These analyses revealed that only germinated kiwi pollen released detectable concentrations of nanoparticles compatible with small EVs for shape and protein content. Moreover, a plant homolog of ALIX, which is a well-recognized and accepted marker of small EVs and exosomes in mammals, was found in pollensomes. Discussion: The presence of this protein, along with other proteins involved in endocytosis, is consistent with the hypothesis that pollensomes could comprehend a prominent subpopulation of plant exosome-like vesicles