Physical exercise restores adult neurogenesis deficits induced by simulated microgravity.

Cognitive impairments have been reported in astronauts during spaceflights and documented in ground-based models of simulated microgravity (SMG) in animals. However, the neuronal causes of these behavioral effects remain largely unknown. We explored whether adult neurogenesis, known to be a crucial plasticity mechanism supporting memory processes, is altered by SMG. Adult male Long-Evans rats were submitted to the hindlimb unloading model of SMG. We studied the proliferation, survival and maturation of newborn cells in the following neurogenic niches: the subventricular zone (SVZ)/olfactory bulb (OB) and the dentate gyrus (DG) of the hippocampus, at different delays following various periods of SMG. SMG exposure for 7 days, but not shorter periods of 6 or 24 h, resulted in a decrease of newborn cell proliferation restricted to the DG. SMG also induced a decrease in short-term (7 days), but not long-term (21 days), survival of newborn cells in the SVZ/OB and DG. Physical exercise, used as a countermeasure, was able to reverse the decrease in newborn cell survival observed in the SVZ and DG. In addition, depending on the duration of SMG periods, transcriptomic analysis revealed modifications in gene expression involved in neurogenesis. These findings highlight the sensitivity of adult neurogenesis to gravitational environmental factors during a transient period, suggesting that there is a period of adaptation of physiological systems to this new environment.

Selection of Bis-Indolyl Pyridines and Triphenylamines as New Inhibitors of SARS-CoV-2 Cellular Entry by Modulating the Spike Protein/ACE2 Interfaces.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the infectious agent that has caused the current coronavirus disease (COVID) pandemic. Viral infection relies on the viral S (spike) protein/cellular receptor ACE2 interaction. Disrupting this interaction would lead to early blockage of viral replication. To identify chemical tools to further study these functional interfaces, 139,146 compounds from different chemical libraries were screened through an S/ACE2 in silico virtual molecular model. The best compounds were selected for further characterization using both cellular and biochemical approaches, reiterating SARS-CoV-2 entry and the S/ACE2 interaction. We report here two selected hits, bis-indolyl pyridine AB-00011778 and triphenylamine AB-00047476. Both of these compounds can block the infectivity of lentiviral vectors pseudotyped with the SARS-CoV-2 S protein as well as wild-type and circulating variant SARS-CoV-2 strains in various human cell lines, including pulmonary cells naturally susceptible to infection. AlphaLISA and biolayer interferometry confirmed a direct inhibitory effect of these drugs on the S/ACE2 association. A specific study of the AB-00011778 inhibitory properties showed that this drug inhibits viral replication with a 50% effective concentration (EC50) between 0.1 and 0.5 µM depending on the cell lines. Molecular docking calculations of the interaction parameters of the molecules within the S/ACE2 complex from both wild-type and circulating variants of the virus showed that the molecules may target multiple sites within the S/ACE2 interface. Our work indicates that AB-00011778 constitutes a good tool for modulating this interface and a strong lead compound for further therapeutic purposes.

miR-124-dependent tagging of synapses by synaptopodin enables input-specific homeostatic plasticity.

Homeostatic synaptic plasticity is a process by which neurons adjust their synaptic strength to compensate for perturbations in neuronal activity. Whether the highly diverse synapses on a neuron respond uniformly to the same perturbation remains unclear. Moreover, the molecular determinants that underlie synapse-specific homeostatic synaptic plasticity are unknown. Here, we report a synaptic tagging mechanism in which the ability of individual synapses to increase their strength in response to activity deprivation depends on the local expression of the spine-apparatus protein synaptopodin under the regulation of miR-124. Using genetic manipulations to alter synaptopodin expression or regulation by miR-124, we show that synaptopodin behaves as a "postsynaptic tag" whose translation is derepressed in a subpopulation of synapses and allows for nonuniform homeostatic strengthening and synaptic AMPA receptor stabilization. By genetically silencing individual connections in pairs of neurons, we demonstrate that this process operates in an input-specific manner. Overall, our study shifts the current view that homeostatic synaptic plasticity affects all synapses uniformly to a more complex paradigm where the ability of individual synapses to undergo homeostatic changes depends on their own functional and biochemical state.

MDGAs are fast-diffusing molecules that delay excitatory synapse development by altering neuroligin behavior.

MDGA molecules can bind neuroligins and interfere with trans-synaptic interactions to neurexins, thereby impairing synapse development. However, the subcellular localization and dynamics of MDGAs, or their specific action mode in neurons remain unclear. Here, surface immunostaining of endogenous MDGAs and single molecule tracking of recombinant MDGAs in dissociated hippocampal neurons reveal that MDGAs are homogeneously distributed and exhibit fast membrane diffusion, with a small reduction in mobility across neuronal maturation. Knocking-down/out MDGAs using shRNAs and CRISPR/Cas9 strategies increases the density of excitatory synapses, the membrane confinement of neuroligin-1, and the phosphotyrosine level of neuroligins associated with excitatory post-synaptic differentiation. Finally, MDGA silencing reduces the mobility of AMPA receptors, increases the frequency of miniature EPSCs (but not IPSCs), and selectively enhances evoked AMPA-receptor-mediated EPSCs in CA1 pyramidal neurons. Overall, our results support a mechanism by which interactions between MDGAs and neuroligin-1 delays the assembly of functional excitatory synapses containing AMPA receptors.

Emerging Roles of Extracellular Vesicles in the Central Nervous System: Physiology, Pathology, and Therapeutic Perspectives.

Extracellular vesicles or EVs are secreted by most, if not all, eukaryote cell types and recaptured by neighboring or distant cells. Their cargo, composed of a vast diversity of proteins, lipids, and nucleic acids, supports the EVs' inter-cellular communication. The role of EVs in many cellular processes is now well documented both in physiological and pathological conditions. In this review, we focus on the role of EVs in the central nervous system (CNS) in physiological as well as pathological conditions such as neurodegenerative diseases or brain cancers. We also discuss the future of EVs in clinical research, in particular, their value as biomarkers as well as innovative therapeutic agents. While an increasing number of studies reveal EV research as a promising field, progress in the standardization of protocols and innovation in analysis as well as in research tools is needed to make a breakthrough in our understanding of their impact in the pathophysiology of the brain.

In Silico, In Vitro and In Cellulo Models for Monitoring SARS-CoV-2 Spike/Human ACE2 Complex, Viral Entry and Cell Fusion.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the etiologic agent responsible for the recent coronavirus disease 2019 (COVID-19) pandemic. Productive SARS-CoV-2 infection relies on viral entry into cells expressing angiotensin-converting enzyme 2 (ACE2). Indeed, viral entry into cells is mostly mediated by the early interaction between the viral spike protein S and its ACE2 receptor. The S/ACE2 complex is, thus, the first contact point between the incoming virus and its cellular target; consequently, it has been considered an attractive therapeutic target. To further characterize this interaction and the cellular processes engaged in the entry step of the virus, we set up various in silico, in vitro and in cellulo approaches that allowed us to specifically monitor the S/ACE2 association. We report here a computational model of the SARS-CoV-2 S/ACE2 complex, as well as its biochemical and biophysical monitoring using pulldown, AlphaLISA and biolayer interferometry (BLI) binding assays. This led us to determine the kinetic parameters of the S/ACE2 association and dissociation steps. In parallel to these in vitro approaches, we developed in cellulo transduction assays using SARS-CoV-2 pseudotyped lentiviral vectors and HEK293T-ACE2 cell lines generated in-house. This allowed us to recapitulate the early replication stage of the infection mediated by the S/ACE2 interaction and to detect cell fusion induced by the interaction. Finally, a cell imaging system was set up to directly monitor the S/ACE2 interaction in a cellular context and a flow cytometry assay was developed to quantify this association at the cell surface. Together, these different approaches are available for both basic and clinical research, aiming to characterize the entry step of the original SARS-CoV-2 strain and its variants as well as to investigate the possible chemical modulation of this interaction. All these models will help in identifying new antiviral agents and new chemical tools for dissecting the virus entry step.

Spinal Inhibition of GABAB Receptors by the Extracellular Matrix Protein Fibulin-2 in Neuropathic Rats.

In the central nervous system, the inhibitory GABAB receptor is the archetype of heterodimeric G protein-coupled receptors (GPCRs). Receptor interaction with partner proteins has emerged as a novel mechanism to alter GPCR signaling in pathophysiological conditions. We propose here that GABAB activity is inhibited through the specific binding of fibulin-2, an extracellular matrix protein, to the B1a subunit in a rat model of neuropathic pain. We demonstrate that fibulin-2 hampers GABAB activation, presumably through decreasing agonist-induced conformational changes. Fibulin-2 regulates the GABAB-mediated presynaptic inhibition of neurotransmitter release and weakens the GABAB-mediated inhibitory effect in neuronal cell culture. In the dorsal spinal cord of neuropathic rats, fibulin-2 is overexpressed and colocalized with B1a. Fibulin-2 may thus interact with presynaptic GABAB receptors, including those on nociceptive afferents. By applying anti-fibulin-2 siRNA in vivo, we enhanced the antinociceptive effect of intrathecal baclofen in neuropathic rats, thus demonstrating that fibulin-2 limits the action of GABAB agonists in vivo. Taken together, our data provide an example of an endogenous regulation of GABAB receptor by extracellular matrix proteins and demonstrate its functional impact on pathophysiological processes of pain sensitization.

miRNA-Dependent Control of Homeostatic Plasticity in Neurons.

Homeostatic plasticity is a form of plasticity in which neurons compensate for changes in neuronal activity through the control of key physiological parameters such as the number and the strength of their synaptic inputs and intrinsic excitability. Recent studies revealed that miRNAs, which are small non-coding RNAs repressing mRNA translation, participate in this process by controlling the translation of multiple effectors such as glutamate transporters, receptors, signaling molecules and voltage-gated ion channels. In this review, we present and discuss the role of miRNAs in both cell-wide and compartmentalized forms of homeostatic plasticity as well as their implication in pathological processes associated with homeostatic failure.

Acquisition of analgesic properties by the cholecystokinin (CCK)/CCK2 receptor system within the amygdala in a persistent inflammatory pain condition.

Pain is associated with negative emotions such as anxiety, but the underlying neurocircuitry and modulators of the association of pain and anxiety remain unclear. The neuropeptide cholecystokinin (CCK) has both pronociceptive and anxiogenic properties, so we explored the role of CCK in anxiety and nociception in the central amygdala (CeA), a key area in control of emotions and descending pain pathways. Local infusion of CCK into the CeA of control rats increased anxiety, as measured in the light-dark box test, but had no effect on mechanical sensitivity. By contrast, intra-CeA CCK infusion 4 days after Complete Freund's Adjuvant (CFA) injection into the hindpaw resulted in analgesia, but also in loss of its anxiogenic capacity. Inflammatory conditions induced changes in the CeA CCK signaling system with an increase of CCK immunoreactivity and a decrease in CCK1, but not CCK2, receptor mRNA. In CFA rats, patch-clamp experiments revealed that CCK infusion increased CeA neuron excitability. It also partially blocked the discharge of wide dynamic range neurons in the dorsal spinal cord. These effects of CCK on CeA and spinal neurons in CFA rats were mimicked by the specific CCK2 receptor agonist, gastrin. This analgesic effect was likely mediated by identified CeA neurons projecting to the periaqueductal gray matter that express CCK receptors. Together, our data demonstrate that intra-CeA CCK infusion activated a descending CCK2 receptor-dependent pathway that inhibited spinal neuron discharge. Thus, persistent pain induces a functional switch to a newly identified analgesic capacity of CCK in the amygdala, indicating central emotion-related circuit controls pain transmission in spinal cord.

Small RNA-Seq reveals novel miRNAs shaping the transcriptomic identity of rat brain structures.

In the central nervous system (CNS), miRNAs are involved in key functions, such as neurogenesis and synaptic plasticity. Moreover, they are essential to define specific transcriptomes in tissues and cells. However, few studies were performed to determine the miRNome of the different structures of the rat CNS, although a major model in neuroscience. Here, we determined by small RNA-Seq, the miRNome of the olfactory bulb, the hippocampus, the cortex, the striatum, and the spinal cord and showed the expression of 365 known miRNAs and 90 novel miRNAs. Differential expression analysis showed that several miRNAs were specifically enriched/depleted in these CNS structures. Transcriptome analysis by mRNA-Seq and correlation based on miRNA target predictions suggest that the specifically enriched/depleted miRNAs have a strong impact on the transcriptomic identity of the CNS structures. Altogether, these results suggest the critical role played by these enriched/depleted miRNAs, in particular the novel miRNAs, in the functional identities of CNS structures.

Oxaliplatin treatment impairs extension of sensory neuron neurites in vitro through miR-204 overexpression.

Oxaliplatin is a platinum-based drug used in the treatment of gastric cancers. Oxaliplatin treatment induces sensory neuropathy characterized by cold hypersensibility in the acute phase and sensory impairment when the neuropathy becomes chronic. In order to determine the effect of oxaliplatin on sensory neurons, we used an in vitro model in which oxaliplatin treatment reduced arborization of dorsal root ganglia neurons in a dose-dependent manner. Moreover, we characterized the role of microRNAs in oxaliplatin induced-neuropathy. In particular, we focused on microRNAs that control the expression of axon guidance molecules, and therefore, regulate neurite arborization. As a result, we highlighted the upregulation of miR-204, a microRNA that controls the expression of PlexinA2, a semaphorin receptor involved in neurite guidance. Interaction of miR-204 and Plexin A2 was confirmed by luciferase assay. In addition, overexpression of miR-204 in dorsal root ganglia neuron cultures reduced length and extension of neurites and also reduced Plexin A2 labelling without increasing apoptosis rate. On the other hand, sequestration of miR-204 by a specific microRNA sponge increases neurite length and PlexinA2 expression. Taken together, our data indicate that oxaliplatin impairs sensory neurons arborization through up-regulation of miR-204 that decreases PlexinA2 expression and neurite length.

APE1/Ref-1 redox function contributes to inflammatory pain sensitization.

Inflammatory pain is a complex and multifactorial disorder. Apurinic/apyrimidinic endonuclease 1 (APE1), also called Redox Factor-1 (Ref-1), is constitutively expressed in the central nervous system and regulates various cellular functions including oxidative stress. In the present study, we investigated APE1 modulation and associated pain behavior changes in the complete Freund's adjuvant (CFA) model of inflammatory pain in rats. In addition we tested the anti-inflammatory effects of E3330, a selective inhibitor of APE1-redox activity, in CFA pain condition. We demonstrate that APE1 expression and subcellular distribution are significantly altered in rats at 4 days post CFA injection. We observed around 30% reduction in the overall APE1 mRNA and protein levels. Interestingly, our data point to an increased nuclear accumulation in the inflamed group as compared to the sham group. E3330 inhibitor injection in CFA rats normalized APE1 mRNA expression and changed its distribution toward cytosolic accumulation. Furthermore, intrathecal injection of E3330 decreased inflammation (i.e. reduced IL-6 expression) and alleviated pain, as assessed by measuring the paw withdrawal threshold with the von Frey test. In conclusion, our data indicate that changes in APE1 expression and sub-cellular distribution are implicated in inflammatory pain mechanisms mediated by APE1 redox functions. Further studies are required to elucidate the exact function of APE1 in inflammatory pain processes.

Group I metabotropic glutamate receptor plasticity after peripheral inflammation alters nociceptive transmission in the dorsal of the spinal cord in adult rats.

Abstract: The dorsal horn of the spinal cord is a crucial site for pain transmission and modulation. Dorsal horn neurons of the spinal cord express group I metabotropic glutamate receptors (group I mGluRs) that exert a complex role in nociceptive transmission. In particular, group I mGluRs promote the activation of L-type calcium channels, voltage-gated channels involved in short- and long-term sensitization to pain. In this study, we analyzed the role of group I mGluRs in spinal nociceptive transmission and the possible cooperation between these receptors and L-type calcium channels in the pathophysiology of pain transmission in the dorsal horn of the spinal cord. We demonstrate that the activation of group I mGluRs induces allodynia and L-type calcium channel-dependent increase in nociceptive field potentials following sciatic nerve stimulation. Surprisingly, in a model of persistent inflammation induced by complete Freund's adjuvant, the activation of group I mGluRs induced an analgesia and a decrease in nociceptive field potentials. Among the group I mGluRs, mGluR1 promotes the activation of L-type calcium channels and increased nociceptive transmission while mGluR5 induces the opposite through the inhibitory network. These results suggest a functional switch exists in pathological conditions that can change the action of group I mGluR agonists into possible analgesic molecules, thereby suggesting new therapeutic perspectives to treat persistent pain in inflammatory settings.

Spinal miRNA-124 regulates synaptopodin and nociception in an animal model of bone cancer pain.

Strong breakthrough pain is one of the most disabling symptoms of cancer since it affects up to 90% of cancer patients and is often refractory to treatments. Alteration in gene expression is a known mechanism of cancer pain in which microRNAs (miRNAs), a class of non-coding regulatory RNAs, play a crucial role. Here, in a mouse model of cancer pain, we show that miR-124 is down-regulated in the spinal cord, the first relay of the pain signal to the brain. Using in vitro and in vivo approaches, we demonstrate that miR-124 is an endogenous and specific inhibitor of synaptopodin (Synpo), a key protein for synaptic transmission. In addition, we demonstrate that Synpo is a key component of the nociceptive pathways. Interestingly, miR-124 was down-regulated in the spinal cord in cancer pain conditions, leading to an up-regulation of Synpo. Furthermore, intrathecal injections of miR-124 mimics in cancerous mice normalized Synpo expression and completely alleviated cancer pain in the early phase of the cancer. Finally, miR-124 was also down-regulated in the cerebrospinal fluid of cancer patients who developed pain, suggesting that miR-124 could be an efficient analgesic drug to treat cancer pain patients.

MicroRNA and chronic pain: From mechanisms to therapeutic potential.

Chronic pain is a major public health issue with an incidence of 20-25% worldwide that can take different forms like neuropathic, cancer-related or inflammatory pain. Chronic pain often limits patients in their daily activities leading to despair. Thus, the goal of treatments is to relieve pain sufficiently to enable patients to go back to a normal life. Unfortunately, few patients with chronic pain obtain complete relief from the analgesics that are currently available. It is thus of prime importance to get a better understanding of chronic pain mechanisms to design new therapeutic strategies and pain-killers. In this sense, the study of microRNA (miRNAs) in chronic pain conditions could lead to a breakthrough in pain management. miRNAs have emerged as master regulators of gene expression in the nervous system where they contribute to neuronal network plasticity. The involvement of miRNAs in the maladaptive plasticity mechanisms of chronic pain is now well documented. Here, we review studies conducted in different animal models and in patients that screened chronic pain-related miRNAs and their targets. Clinical studies suggest that miRNAs expression could reflect the high variability among pain patients that could help to categorize patients and finally lead to personalized therapies. We also point out the different strategies investigated to design miRNA-based analgesics. Finally, we highlight the current miRNA-based clinical trials to hypothesize their potential as therapeutic tool for chronic pain.

Glucocerebrosidase deficiency in dopaminergic neurons induces microglial activation without neurodegeneration.

Mutations in the GBA1 gene encoding the lysosomal enzyme glucocerebrosidase (GBA1) are important risk factors for Parkinson's disease (PD). In vitro, altered GBA1 activity promotes alpha-synuclein accumulation whereas elevated levels of alpha-synuclein compromise GBA1 function, thus supporting a pathogenic mechanism in PD. However, the mechanisms by which GBA1 deficiency is linked to increased risk of PD remain elusive, partially because of lack of aged models of GBA1 deficiency. As knocking-out GBA1 in the entire brain induces massive neurodegeneration and early death, we generated a mouse model of GBA1 deficiency amenable to investigate the long-term consequences of compromised GBA1 function in dopaminergic neurons. DAT-Cre and GBA1-floxed mice were bred to obtain selective homozygous disruption of GBA1 in midbrain dopamine neurons (DAT-GBA1-KO). Mice were followed for motor function, neuronal survival, alpha-synuclein phosphorylation and glial activation. Susceptibility to nigral viral vector-mediated overexpression of mutated (A53T) alpha-synuclein was assessed. Despite loss of GBA1 and substrate accumulation, DAT-GBA1-KO mice displayed normal motor performances and preserved dopaminergic neurons despite robust microglial activation in the substantia nigra, without accumulation of endogenous alpha-synuclein with respect to wild-type mice. Lysosomal function was only marginally affected. Screening of micro-RNAs linked to the regulation of GBA1, alpha-synuclein or neuroinflammation did not reveal significant alterations. Viral-mediated overexpression of A53T-alpha-synuclein yielded similar neurodegeneration in DAT-GBA1-KO mice and wild-type mice. These results indicate that loss of GBA1 function in mouse dopaminergic neurons is not critical for alpha-synuclein accumulation or neurodegeneration and suggest the involvement of GBA1 deficiency in other cell types as a potential mechanism.

Cav1.2 and Cav1.3 L-type calcium channels independently control short- and long-term sensitization to pain.

KEY POINTS: L-type calcium channels in the CNS exist as two subunit forming channels, Cav1.2 and Cav1.3, which are involved in short- and long-term plasticity. We demonstrate that Cav1.3 but not Cav1.2 is essential for wind-up. These results identify Cav1.3 as a key conductance responsible for short-term sensitization in physiological pain transmission. We confirm the role of Cav1.2 in a model of long-term plasticity associated with neuropathic pain. Up-regulation of Cav1.2 and down-regultation of Cav1.3 in neuropathic pain underlies the switch from physiology to pathology. Finally, the results of the present study reveal that therapeutic targeting molecular pathways involved in wind-up may be not relevant in the treatment of neuropathy. ABSTRACT: Short-term central sensitization to pain temporarily increases the responsiveness of nociceptive pathways after peripheral injury. In dorsal horn neurons (DHNs), short-term sensitization can be monitored through the study of wind-up. Wind-up, a progressive increase in DHNs response following repetitive peripheral stimulations, depends on the post-synaptic L-type calcium channels. In the dorsal horn of the spinal cord, two L-type calcium channels are present, Cav1.2 and Cav1.3, each displaying specific kinetics and spatial distribution. In the present study, we used a mathematical model of DHNs in which we integrated the specific patterns of expression of each Cav subunits. This mathematical approach reveals that Cav1.3 is necessary for the onset of wind-up, whereas Cav1.2 is not and that synaptically triggered wind-up requires NMDA receptor activation. We then switched to a biological preparation in which we knocked down Cav subunits and confirmed the prominent role of Cav1.3 in both naive and spinal nerve ligation model of neuropathy (SNL). Interestingly, although a clear mechanical allodynia dependent on Cav1.2 expression was observed after SNL, the amplitude of wind-up was decreased. These results were confirmed with our model when adapting Cav1.3 conductance to the changes observed after SNL. Finally, our mathematical approach predicts that, although wind-up amplitude is decreased in SNL, plateau potentials are not altered, suggesting that plateau and wind-up are not fully equivalent. Wind-up and long-term hyperexcitability of DHNs are differentially controlled by Cav1.2 and Cav1.3, therefore confirming that short- and long-term sensitization are two different phenomena triggered by distinct mechanisms.

miR-92a regulates expression of synaptic GluA1-containing AMPA receptors during homeostatic scaling.

We investigated whether microRNAs could regulate AMPA receptor expression during activity blockade. miR-92a strongly repressed the translation of GluA1 receptors by binding the 3' untranslated region of rat GluA1 (also known as Gria1) mRNA and was downregulated in rat hippocampal neurons after treatment with tetrodotoxin and AP5. Deleting the seed region in GluA1 or overexpressing miR-92a blocked homeostatic scaling, indicating that miR-92a regulates the translation and synaptic incorporation of new GluA1-containing AMPA receptors.

MicroRNAs regulate neuronal plasticity and are involved in pain mechanisms.

MicroRNAs (miRNAs) are emerging as master regulators of gene expression in the nervous system where they contribute not only to brain development but also to neuronal network homeostasis and plasticity. Their function is the result of a cascade of events including miRNA biogenesis, target recognition, and translation inhibition. It has been suggested that miRNAs are major switches of the genome owing to their ability to regulate multiple genes at the same time. This regulation is essential for normal neuronal activity and, when affected, can lead to drastic pathological conditions. As an example, we illustrate how deregulation of miRNAs can affect neuronal plasticity leading to chronic pain. The origin of pain and its dual role as a key physiological function and a debilitating disease has been highly debated until now. The incidence of chronic pain is estimated to be 20-25% worldwide, thus making it a public health problem. Chronic pain can be considered as a form of maladaptive plasticity. Long-lasting modifications develop as a result of global changes in gene expression, and are thus likely to be controlled by miRNAs. Here, we review the literature on miRNAs and their targets responsible for maladaptive plasticity in chronic pain conditions. In addition, we conduct a retrospective analysis of miRNA expression data published for different pain models, taking into account recent progress in our understanding of the role of miRNAs in neuronal plasticity.

High epiregulin expression in human U87 glioma cells relies on IRE1α and promotes autocrine growth through EGF receptor.

BACKGROUND: Epidermal growth factor (EGF) receptors contribute to the development of malignant glioma. Here we considered the possible implication of the EGFR ligand epiregulin (EREG) in glioma development in relation to the activity of the unfolded protein response (UPR) sensor IRE1α. We also examined EREG status in several glioblastoma cell lines and in malignant glioma. METHODS: Expression and biological properties of EREG were analyzed in human glioma cells in vitro and in human tumor xenografts with regard to the presence of ErbB proteins and to the blockade of IRE1α. Inactivation of IRE1α was achieved by using either the dominant-negative strategy or siRNA-mediated knockdown. RESULTS: EREG was secreted in high amounts by U87 cells, which also expressed its cognate EGF receptor (ErbB1). A stimulatory autocrine loop mediated by EREG was evidenced by the decrease in cell proliferation using specific blocking antibodies directed against either ErbB1 (cetuximab) or EREG itself. In comparison, anti-ErbB2 antibodies (trastuzumab) had no significant effect. Inhibition of IRE1α dramatically reduced EREG expression both in cell culture and in human xenograft tumor models. The high-expression rate of EREG in U87 cells was therefore linked to IRE1α, although being modestly affected by chemical inducers of the endoplasmic reticulum stress. In addition, IRE1-mediated production of EREG did not depend on IRE1 RNase domain, as neither the selective dominant-negative invalidation of the RNase activity (IRE1 kinase active) nor the siRNA-mediated knockdown of XBP1 had significant effect on EREG expression. Finally, chemical inhibition of c-Jun N-terminal kinases (JNK) using the SP600125 compound reduced the ability of cells to express EREG, demonstrating a link between the growth factor production and JNK activation under the dependence of IRE1α. CONCLUSION: EREG may contribute to glioma progression under the control of IRE1α, as exemplified here by the autocrine proliferation loop mediated in U87 cells by the growth factor through ErbB1.