Epigenetic combined with transcriptomic analysis of the m6A methylome after spared nerve injury-induced neuropathic pain in mice.

Epigenetic changes in the spinal cord play a key role in the initiation and maintenance of nerve injury-induced neuropathic pain. N6-methyladenosine (m6A) is one of the most abundant internal RNA modifications and plays an essential function in gene regulation in many diseases. However, the global m6A modification status of mRNA in the spinal cord at different stages after neuropathic pain is unknown.

Single-cell analyses reveal the dynamic functions of Itgb2 microglia subclusters at different stages of cerebral ischemia-reperfusion injury in transient middle cerebral occlusion mice model.

Introduction: The underlying pathophysiological mechanisms of cerebral ischemia reperfusion injury (CIRI) is intricate, and current studies suggest that neuron, astrocyte, microglia, endothelial cell, and pericyte all have different phenotypic changes of specific cell types after ischemic stroke. And microglia account for the largest proportion after CIRI. Previous transcriptomic studies of ischemic stroke have typically focused on the 24 hours after CIRI, obscuring the dynamics of cellular subclusters throughout the disease process.

Celastrol-encapsulated microspheres prepared by microfluidic electrospray for alleviating inflammatory pain.

Inflammatory pain is induced by trauma, infection, chemical stimulation, etc. It causes severe physical and psychological agony to patients. Celastrol has powerful anti-inflammatory property and has achieved good results in various inflammation-related diseases.

Celastrol targeting Nedd4 reduces Nrf2-mediated oxidative stress in astrocytes after ischemic stroke.

Stroke is the second leading cause of death worldwide, and oxidative stress plays a crucial role. Celastrol exhibits strong antioxidant properties in several diseases; however, whether it can affect oxidation in cerebral ischemic-reperfusion injury (CIRI) remains unclear. This study aimed to determine whether celastrol could reduce oxidative damage during CIRI and to elucidate the underlying mechanisms.

miR-26a-5p alleviates CFA-induced chronic inflammatory hyperalgesia through Wnt5a/CaMKII/NFAT signaling in mice.

Background: Inflammation often leads to the occurrence of chronic pain, and many miRNAs have been shown to play a key role in the development of inflammatory pain. However, whether miR-26a-5p relieves pain induced by inflammation and its possible mechanism are still unclear.

Methods: The complete Freund's adjuvant (CFA)-induced inflammatory pain mouse model was employed.

Human PMSCs-derived small extracellular vesicles alleviate neuropathic pain through miR-26a-5p/Wnt5a in SNI mice model.

Background: Mesenchymal stem cell (MSCs)-derived small Extracellular Vesicles (sEVs) are considered as a new cell-free therapy for pain caused by nerve injury, but whether human placental mesenchymal stem cell-derived sEVs relieve pain in sciatic nerve injury and its possible mechanism are still unclear. In this study, we investigated the roles of hPMSCs-derived sEVs and related mechanisms in neuropathic pain.

Methods: The spared nerve injury (SNI) mouse model was employed.

Effect of Celastrol on LncRNAs and mRNAs Profiles of Cerebral Ischemia-Reperfusion Injury in Transient Middle Cerebral Artery Occlusion Mice Model.

Celastrol plays a significant role in cerebral ischemia-reperfusion injury. Although previous studies have confirmed that celastrol post-treatment has a protective effect on ischemic stroke, the therapeutic effect of celastrol on ischemic stroke and the underlying molecular mechanism remain unclear. In the present study, focal transient cerebral ischemia was induced by transient middle cerebral artery occlusion (tMCAO) in mice and celastrol was administered immediately after reperfusion.

Celastrol Protects against Cerebral Ischemia/Reperfusion Injury in Mice by Inhibiting Glycolysis through Targeting HIF-1/PDK1 Axis.

Cerebral ischemia/reperfusion (I/R) injury is closely related to dysfunctional glucose metabolism. Celastrol is a bioactive compound that has been found to exhibit neuroprotective effects in cerebral ischemia, while whether it can protect against cerebral I/R injury by regulating glycolysis remains unclear. The goal of this study is to investigate the role of celastrol on cerebral I/R injury and its underlying mechanisms in transient middle cerebral artery occlusion (tMCAO) mice.

Supplemental N-3 Polyunsaturated Fatty Acids Limit A1-Specific Astrocyte Polarization via Attenuating Mitochondrial Dysfunction in Ischemic Stroke in Mice.

Ischemic stroke is one of the leading causes of death and disability for adults, which lacks effective treatments. Dietary intake of n-3 polyunsaturated fatty acids (n-3 PUFAs) exerts beneficial effects on ischemic stroke by attenuating neuron death and inflammation induced by microglial activation. However, the impact and mechanism of n-3 PUFAs on astrocyte function during stroke have not yet been well investigated.

A Simple and Efficient CRISPR Technique for Protein Tagging.

Genetic knock-in using homology-directed repair is an inefficient process, requiring the selection of few modified cells and hindering its application to primary cells. Here, we describe Homology independent gene Tagging (HiTag), a method to tag a protein of interest by CRISPR in up to 66% of transfected cells with one single electroporation. The technique has proven effective in various cell types and can be used to knock in a fluorescent protein for live cell imaging, to modify the cellular location of a target protein and to monitor the levels of a protein of interest by a luciferase assay in primary cells.

Propofol Exposure Disturbs the Differentiation of Rodent Neural Stem Cells via an miR-124-3p/Sp1/Cdkn1b Axis.

Accumulating studies have indicated that propofol may lead to neurotoxicity and its effect on neural stem cells (NSCs) may play pivotal role in propofol-related neurotoxicity. Previously, we found that propofol could promote NSCs proliferation and could regulate several microRNA expressions. However, the underlying mechanism between microRNAs and NSCs development after propofol exposure is still unclear.

GPR39 is coupled to TMEM16A in intestinal fibroblast-like cells.

GPR39 is a GPCR implicated as a regulator of gastrointestinal motility, although the mechanism remains elusive. Here, we report that GPR39 is expressed by a specific cell population cultured from mouse small intestine muscle layers, which was subsequently identified as fibroblast-like cells (FLCs) that have recently been shown to modulate gut motility. Application of the GPR39 agonist, Zn(2+), induced large currents and membrane depolarization in FLCs cultured from wild-type mice, but not Gpr39(-/-) mice.

TRPA1 agonist activity of probenecid desensitizes channel responses: consequences for screening.

The transient receptor potential channel subtype A member 1 (TRPA1) is a nonselective cation channel widely viewed as having therapeutic potential, particularly for pain-related indications. Realization of this potential will require potent, selective modulators; however, currently the pharmacology of TRPA1 is poorly defined. As TRPA1 is calcium permeable, calcium indicators offer a simple assay format for high-throughput screening.

Increased prokineticin 2 expression in gut inflammation: role in visceral pain and intestinal ion transport.

Background: Prokineticin 2 (PROK2) is an inflammatory cytokine-like molecule expressed predominantly by macrophages and neutrophils infiltrating sites of tissue damage. Given the established role of prokineticin signaling on gastrointestinal function, we have explored Prok2 gene expression in inflammatory conditions of the gastrointestinal tract and assessed the possible consequences on gut physiology.

Methods: Prokineticin expression was examined in normal and colitic tissues using qPCR and immunohistochemistry.

TRPC1 transcript variants, inefficient nonsense-mediated decay and low up-frameshift-1 in vascular smooth muscle cells.

Background: Transient Receptor Potential Canonical 1 (TRPC1) is a widely-expressed mammalian cationic channel with functional effects that include stimulation of cardiovascular remodelling. The initial aim of this study was to investigate variation in TRPC1-encoding gene transcripts.

Results: Extensive TRPC1 transcript alternative splicing was observed, with exons 2, 3 and 5-9 frequently omitted, leading to variants containing premature termination codons.

Potent suppression of vascular smooth muscle cell migration and human neointimal hyperplasia by KV1.3 channel blockers.

Aim: The aim of the study was to determine the potential for K(V)1 potassium channel blockers as inhibitors of human neoinitimal hyperplasia.

Methods And Results: Blood vessels were obtained from patients or mice and studied in culture. Reverse transcriptase-polymerase chain reaction and immunocytochemistry were used to detect gene expression.

Pregnenolone sulphate- and cholesterol-regulated TRPM3 channels coupled to vascular smooth muscle secretion and contraction.

Rationale: Transient receptor potential melastatin (TRPM)3 is a calcium-permeable ion channel activated by the neurosteroid pregnenolone sulfate and positively coupled to insulin secretion in beta cells. Although vascular TRPM3 mRNA has been reported, there is no knowledge of TRPM3 protein or its regulation and function in the cardiovascular system.

Objective: To determine the relevance and regulation of TRPM3 in vascular biology.

Glial cell-derived neurotrophic factor enhances synaptic communication and 5-hydroxytryptamine 3a receptor expression in enteric neurons.

Background & Aims: Glial cell-derived neurotrophic factor (GDNF) is essential for the development of the enteric nervous system during embryogenesis. We have observed the presence of Gdnf transcripts in the gastrointestinal tract of adult mice, and its early up-regulation after inflammation. We therefore investigated the effects of GDNF on enteric neuronal function in vitro.

Robotic multiwell planar patch-clamp for native and primary mammalian cells.

Robotic multiwell planar patch-clamp has become common in drug development and safety programs because it enables efficient and systematic testing of compounds against ion channels during voltage-clamp. It has not, however, been adopted significantly in other important areas of ion channel research, where conventional patch-clamp remains the favored method. Here, we show the wider potential of the multiwell approach with the ability for efficient intracellular solution exchange, describing protocols and success rates for recording from a range of native and primary mammalian cells derived from blood vessels, arthritic joints and the immune and central nervous systems.

TRPC channel activation by extracellular thioredoxin.

Mammalian homologues of Drosophila melanogaster transient receptor potential (TRP) are a large family of multimeric cation channels that act, or putatively act, as sensors of one or more chemical factor. Major research objectives are the identification of endogenous activators and the determination of cellular and tissue functions of these channels. Here we show the activation of TRPC5 (canonical TRP 5) homomultimeric and TRPC5-TRPC1 heteromultimeric channels by extracellular reduced thioredoxin, which acts by breaking a disulphide bridge in the predicted extracellular loop adjacent to the ion-selectivity filter of TRPC5.

K(v)1.5 potassium channel gene regulation by Sp1 transcription factor and oxidative stress.

K(V)1.5, a voltage-gated potassium channel, has functional importance in regulating blood vessel tone and cardiac action potentials and is a target for numerous therapeutic drug development programs. Despite the importance of K(V)1.

A sphingosine-1-phosphate-activated calcium channel controlling vascular smooth muscle cell motility.

In a screen of potential lipid regulators of transient receptor potential (TRP) channels, we identified sphingosine-1-phosphate (S1P) as an activator of TRPC5. We explored the relevance to vascular biology because S1P is a key cardiovascular signaling molecule. TRPC5 is expressed in smooth muscle cells of human vein along with TRPC1, which forms a complex with TRPC5.

Calcium-sensing mechanism in TRPC5 channels contributing to retardation of neurite outgrowth.

The calcium- and sodium-permeable transient receptor potential channel TRPC5 has an inhibitory role in neuronal outgrowth but the mechanisms governing its activity are poorly understood. Here we propose a mechanism involving the neuronal calcium sensor-1 (NCS-1) protein. Inhibitory mutants of TRPC5 and NCS-1 enhance neurite outgrowth similarly.

Sensing of lysophospholipids by TRPC5 calcium channel.

TRPC calcium channels are emerging as a ubiquitous feature of vertebrate cells, but understanding of them is hampered by limited knowledge of the mechanisms of activation and identity of endogenous regulators. We have revealed that one of the TRPC channels, TRPC5, is strongly activated by common endogenous lysophospholipids including lysophosphatidylcholine (LPC) but, by contrast, not arachidonic acid. Although TRPC5 was stimulated by agonists at G-protein-coupled receptors, TRPC5 activation by LPC occurred downstream and independently of G-protein signaling.

Generation of functional ion-channel tools by E3 targeting.

Here we describe a strategy for generating ion-channel inhibitors. It takes advantage of antibody specificity combined with a pattern recognition approach that targets the third extracellular region (E3) of a channel. To test the concept, we first focused on TRPC5, a member of the transient receptor potential (TRP) calcium channel family, the study of which has been hindered by poor pharmacological tools.