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Neuropathic pain is a debilitating chronic condition that lacks effective treatment. The role of cytokine- and chemokine-mediated neuroinflammation in its pathogenesis has been well documented. Follistatin (FST) is a secreted protein known to antagonize the biological activity of cytokines in the transforming growth factor–β (TGF-β) superfamily. The involvement of FST in neuropathic pain and the underlying mechanism remain largely unknown. Here, we report that FST was up-regulated in A-fiber sensory neurons after spinal nerve ligation (SNL) in mice. Inhibition or deletion of FST alleviated neuropathic pain and reduced the nociceptive neuron hyperexcitability induced by SNL. Conversely, intrathecal or intraplantar injection of recombinant FST, or overexpression of FST in the dorsal root ganglion (DRG) neurons, induced pain hypersensitivity. Furthermore, exogenous FST increased neuronal excitability in nociceptive neurons. The biolayer interferometry (BLI) assay and coimmunoprecipitation (co-IP) demonstrated direct binding of FST to the insulin-like growth factor–1 receptor (IGF1R), and IGF1R inhibition reduced FST-induced activation of extracellular signal–regulated kinase (ERK) and protein kinase B (AKT), as well as neuronal hyperexcitability. Further co-IP analysis revealed that the N-terminal domain of FST exhibits the highest affinity for IGF1R, and blocking this interaction with a peptide derived from FST attenuated Nav1.7-mediated neuronal hyperexcitability and neuropathic pain after SNL. In addition, FST enhanced neuronal excitability in human DRG neurons through IGF1R. Collectively, our findings suggest that FST, released from A-fiber neurons, enhances Nav1.7-mediated hyperexcitability of nociceptive neurons by binding to IGF1R, making it a potential target for neuropathic pain treatment. Editor's summary: Neuroinflammation contributes to the pathogenesis of neuropathic pain. The characterization of signaling pathways involved could help to identify new therapeutic targets. Here, Jiang et al. used a mouse model to show that the glycoprotein follistatin (FST) was up-regulated in sensory neurons in the dorsal root ganglia (DRG) after spinal nerve ligation. FST injections were sufficient to drive mechanical hypersensitivity and excitability of DRG neurons. FST interacted with insulin-like growth factor–1 receptor, and a peptide that blocked these interactions could alleviate neuropathic pain–related behaviors and FST-mediated voltage-gated sodium channel currents, highlighting a potential therapeutic strategy that warrants further examination. —Daniela Neuhofer [ABSTRACT FROM AUTHOR]

Phosphorylation of the serine 139 of the histone variant H2AX (γH2AX) is a DNA damage marker that regulates DNA damage response and various diseases. However, whether γH2AX is involved in neuropathic pain is still unclear. We found the expression of γH2AX and H2AX decreased in mice dorsal root ganglion (DRG) after spared nerve injury (SNI). Ataxia telangiectasia mutated (ATM), which promotes γH2AX, was also down-regulated in DRG after peripheral nerve injury. ATM inhibitor KU55933 decreased the level of γH2AX in ND7/23 cells. The intrathecal injection of KU55933 down-regulated DRG γH2AX expression and significantly induced mechanical allodynia and thermal hyperalgesia in a dose-dependent manner. The inhibition of ATM by siRNA could also decrease the pain threshold. The inhibition of dephosphorylation of γH2AX by protein phosphatase 2A (PP2A) siRNA partially suppressed the down-regulation of γH2AX after SNI and relieved pain behavior. Further exploration of the mechanism revealed that inhibiting ATM by KU55933 up-regulated extracellular-signal regulated kinase (ERK) phosphorylation and down-regulated potassium ion channel genes, such as potassium voltage-gated channel subfamily Q member 2 (Kcnq2) and potassium voltage-gated channel subfamily D member 2 (Kcnd2) in vivo, and KU559333 enhanced sensory neuron excitability in vitro. These preliminary findings imply that the down-regulation of γH2AX may contribute to neuropathic pain. [ABSTRACT FROM AUTHOR]

Recent studies have implicated chemokines in microglial activation and pathogenesis of neuropathic pain. C-X-C motif chemokine 13 (CXCL13) is a B lymphocyte chemoattractant that activates CXCR5. Using the spinal nerve ligation (SNL) model of neuropathic pain, we found that CXCL13 was persistently upregulated in spinal cord neurons after SNL, resulting in spinal astrocyte activation via CXCR5 in mice. shRNA-mediated inhibition of CXCL13 in the spinal cord persistently attenuated SNL-induced neuropathic pain. Interestingly, CXCL13 expression was suppressed by miR-186-5p, a microRNA that colocalized with CXCL13 and was downregulated after SNL. Spinal overexpression of miR-186-5p decreased CXCL13 expression, alleviating neuropathic pain. Furthermore, SNL induced CXCR5 expression in spinal astrocytes, and neuropathic pain was abrogated in [Cxcr5.sup.-/-] mice. CXCR5 expression induced by SNL was required for the SNL-induced activation of spinal astrocytes and microglia. Intrathecal injection of CXCL13 was sufficient to induce pain hypersensitivity and astrocyte activation via CXCR5 and ERK. Finally, intrathecal injection of CXCL13-activated astrocytes induced mechanical allodynia in naive mice. Collectively, our findings reveal a neuronal/astrocytic interaction in the spinal cord by which neuronally produced CXCL13 activates astrocytes via CXCR5 to facilitate neuropathic pain. Thus, miR-186-5p and CXCL13/CXCR5-mediated astrocyte signaling may be suitable therapeutic targets for neuropathic pain., Introduction Chemokines are a family of small secreted proteins that are well-known regulators of peripheral immune cell trafficking (1). It has become evident that chemokines are also expressed in the [...]

Background: Trigeminal nerve damage-induced neuropathic pain is a severely debilitating chronic orofacial pain syndrome. Spinal chemokine CXCL13 and its receptor CXCR5 were recently demonstrated to play a pivotal role in the pathogenesis of spinal nerve ligation-induced neuropathic pain. Whether and how CXCL13/CXCR5 in the trigeminal ganglion (TG) mediates orofacial pain are unknown.Methods: The partial infraorbital nerve ligation (pIONL) was used to induce trigeminal neuropathic pain in mice. The expression of ATF3, CXCL13, CXCR5, and phosphorylated extracellular signal-regulated kinase (pERK) in the TG was detected by immunofluorescence staining and western blot. The effect of shRNA targeting on CXCL13 or CXCR5 on pain hypersensitivity was checked by behavioral testing.Results: pIONL induced persistent mechanical allodynia and increased the expression of ATF3, CXCL13, and CXCR5 in the TG. Inhibition of CXCL13 or CXCR5 by shRNA lentivirus attenuated pIONL-induced mechanical allodynia. Additionally, pIONL-induced neuropathic pain and the activation of ERK in the TG were reduced in Cxcr5 (-/-) mice. Furthermore, MEK inhibitor (PD98059) attenuated mechanical allodynia and reduced TNF-α and IL-1β upregulation induced by pIONL. TNF-α inhibitor (Etanercept) and IL-1β inhibitor (Diacerein) attenuated pIONL-induced orofacial pain. Finally, intra-TG injection of CXCL13 induced mechanical allodynia, increased the activation of ERK and the production of TNF-α and IL-1β in the TG of WT mice, but not in Cxcr5 (-/-) mice. Pretreatment with PD98059, Etanercept, or Diacerein partially blocked CXCL13-induced mechanical allodynia, and PD98059 also reduced CXCL13-induced TNF-α and IL-1β upregulation.Conclusions: CXCL13 and CXCR5 contribute to orofacial pain via ERK-mediated proinflammatory cytokines production. Targeting CXCL13/CXCR5/ERK/TNF-α and IL-1β pathway in the trigeminal ganglion may offer effective treatment for orofacial neuropathic pain. [ABSTRACT FROM AUTHOR]

Recent studies have implicated chemokines in microglial activation and pathogenesis of neuropathic pain. C-X-C motif chemokine 13 (CXCL13) is a B lymphocyte chemoattractant that activates CXCR5. Using the spinal nerve ligation (SNL) model of neuropathic pain, we found that CXCL13 was persistently upregulated in spinal cord neurons after SNL, resulting in spinal astrocyte activation via CXCR5 in mice. shRNA-mediated inhibition of CXCL13 in the spinal cord persistently attenuated SNL-induced neuropathic pain. Interestingly, CXCL13 expression was suppressed by miR-186-5p, a microRNA that colocalized with CXCL13 and was downregulated after SNL. Spinal overexpression of miR-186-5p decreased CXCL13 expression, alleviating neuropathic pain. Furthermore, SNL induced CXCR5 expression in spinal astrocytes, and neuropathic pain was abrogated in Cxcr5-/- mice. CXCR5 expression induced by SNL was required for the SNL-induced activation of spinal astrocytes and microglia. Intrathecal injection of CXCL13 was sufficient to induce pain hypersensitivity and astrocyte activation via CXCR5 and ERK. Finally, intrathecal injection of CXCL13-activated astrocytes induced mechanical allodynia in naive mice. Collectively, our findings reveal a neuronal/astrocytic interaction in the spinal cord by which neuronally produced CXCL13 activates astrocytes via CXCR5 to facilitate neuropathic pain. Thus, miR-186-5p and CXCL13/CXCR5-mediated astrocyte signaling may be suitable therapeutic targets for neuropathic pain. [ABSTRACT FROM AUTHOR]

Abstract: Trigeminal nerve injury-induced neuropathic pain is a debilitating chronic orofacial pain syndrome but lacks effective treatment. G protein-coupled receptors (GPCRs), especially orphan GPCRs (oGPCRs) are important therapeutic targets in pain medicine. Here, we screened upregulated oGPCRs in the trigeminal ganglion (TG) after partial infraorbital nerve transection (pIONT) and found that Gpr151 was the most significantly upregulated oGPCRs. Gpr151 mRNA was increased from pIONT day 3 and maintained for more than 21 days. Furthermore, GPR151 was expressed in the neurons of the TG after pIONT. Global mutation or knockdown of Gpr151 in the TG attenuated pIONT-induced mechanical allodynia. In addition, the excitability of TG neurons was increased after pIONT in wild-type (WT) mice, but not in Gpr151-/- mice. Notably, GPR151 bound to Gαi protein, but not Gαq, Gα12, or Gα13, and activated the extracellular signal-regulated kinase (ERK) through Gβγ. Extracellular signal-regulated kinase was also activated by pIONT in the TG of WT mice, but not in Gpr151-/- mice. Gene microarray showed that Gpr151 mutation reduced the expression of a large number of neuroinflammation-related genes that were upregulated in WT mice after pIONT, including chemokines CCL5, CCL7, CXCL9, and CXCL10. The mitogen-activated protein kinase inhibitor (PD98059) attenuated mechanical allodynia and reduced the upregulation of these chemokines after pIONT. Collectively, this study not only revealed the involvement of GPR151 in the maintenance of trigeminal neuropathic pain but also identified GPR151 as a Gαi-coupled receptor to induce ERK-dependent neuroinflammation. Thus, GPR151 may be a potential drug target for the treatment of trigeminal neuropathic pain., (Copyright © 2020 International Association for the Study of Pain.)

Spinal cord plays an important role in the transmission and modulation of nociceptive information. Global changes in gene expression in the spinal cord contribute to the induction and maintenance of neuropathic pain. Transcribed Ultraconserved Regions (T-UCRs), a novel class of long noncoding RNAs, can regulate gene expression at both transcriptional and post-transcriptional levels and are related to many human diseases such as cancer, Alzheimer's disease, and heart diseases. In this study, we screened abnormal T-UCRs expression in the spinal cord under spinal nerve ligation (SNL)-induced neuropathic pain condition. Microarray data showed the alternation of T-UCRs at the transcriptional level in the spinal cord 10 days after SNL. Among 78 altered T-UCRs, 23 T-UCRs were upregulated by more than 1.5-fold and 55 ones downregulated by less than 0.5-fold after SNL. Hierarchical cluster analysis of T-UCRs expression profiles showed the opposite expression pattern between SNL and sham-operated mice. The quantitative real-time reverse transcription polymerase chain reaction analysis further confirmed the expression patterns of uc.305, uc.189, uc.46, and uc.217 after SNL. The gene ontology annotation and signaling pathway analysis for the T-UCRs host genes indicated that differentially expressed T-UCRs were involved in several intracellular activities and signaling pathways, including Ephrin receptor activity, soluble NSF attachment protein receptor (SNARE) interactions in vesicular transport pathway, and WNT signaling pathway. Collectively, the current data suggest the possible role of T-UCR in the pathogenesis of neuropathic pain. T-UCRs may serve as a new kind of target for the treatment of neuropathic pain.

Background: Neuropathic pain that caused by lesion or dysfunction of the nervous system is associated with gene expression changes in the sensory pathway. Long noncoding RNAs (lncRNAs) have been reported to be able to regulate gene expression. Identifying lncRNA expression patterns in the spinal cord under normal and neuropathic pain conditions is essential for understanding the genetic mechanisms behind the pathogenesis of neuropathic pain., Results: Spinal nerve ligation (SNL) induced rapid and persistent pain hypersensitivity, characterized by mechanical allodynia and heat hyperalgesia. Meanwhile, astrocytes and microglia were dramatically activated in the ipsilateral spinal cord dorsal horn at 10 days after SNL. Further lncRNA microarray and mRNA microarray analysis showed that the expression profiles of lncRNA and mRNA between SNL and sham-operated mice were greatly changed at 10 days. The 511 differentially expressed (>2 fold) lncRNAs (366 up-regulated, 145 down-regulated) and 493 mRNAs (363 up-regulated, 122 down-regulated) were finally identified. The expression patterns of several lncRNAs and mRNAs were further confirmed by qPCR. Functional analysis of differentially expressed (DE) mRNAs showed that the most significant enriched biological processes of up-regulated genes in SNL include immune response, defense response, and inflammation response, which are important pathogenic mechanisms underlying neuropathic pain. 35 DE lncRNAs have neighboring or overlapping DE mRNAs in genome, which is related to Toll-like receptor signaling, cytokine-cytokine receptor interaction, and peroxisome proliferator-activated receptor signaling pathway., Conclusion: Our findings uncovered the expression pattern of lncRNAs and mRNAs in the mice spinal cord under neuropathic pain condition. These lncRNAs and mRNAs may represent new therapeutic targets for the treatment of neuropathic pain.