Lanatoside C inhibits herpes simplex virus 1 replication by regulating NRF2 distribution within cells.

BACKGROUND: In the past decades, extensive research has been conducted to identify new drug targets for the treatment of Herpes simplex virus type 1 (HSV-1) infections. However, the emergence of drug-resistant HSV-1 strains remains a major challenge. This necessitates the identification of new drugs with novel mechanisms of action. Lanatoside C (LanC), a cardiac glycoside (CG) approved by the US Food and Drug Administration (FDA), has demonstrated anticancer and antiviral properties. Nevertheless, its potential as an agent against HSV-1 infections and the underlying mechanism of action are currently unknown. PURPOSE: This study aimed to investigate the antiviral activity of LanC against HSV-1 and elucidate its molecular mechanisms. METHODS: The in vitro antiviral activity of LanC was assessed by examining the levels of viral genes, proteins, and virus titers in HSV-1-infected ARPE-19 and Vero cells. Immunofluorescence (IF) analysis was performed to determine the intracellular distribution of NRF2. Additionally, an in vivo mouse model of HSV-1 infection was developed to evaluate the antiviral activity of LanC, using indicators such as intraepidermal nerve fibers (IENFs) loss and viral gene inhibition. RESULTS: Our findings demonstrate that LanC significantly inhibits HSV-1 replication both in vitro and in vivo. The antiviral effect of LanC is mediated by the perinuclear translocation of NRF2. CONCLUSIONS: LanC exhibits anti-HSV-1 effects in viral infections, which are associated with the intracellular translocation of NRF2. These findings suggest that LanC has the potential to serve as a novel NRF2 modulator in the treatment of viral diseases.

HSV-1 infection-induced herpetic neuralgia involves a CCL5/CCR5-mediated inflammation mechanism.

Herpetic-related neuralgia (HN) caused by varicella-zoster virus (VZV) infection is one of the most typical and common neuropathic pain in the clinic. However, the potential mechanisms and therapeutic approaches for the prevention and treatment of HN are still unclear. This study aims to provide a comprehensive understanding of the molecular mechanisms and potential therapeutic targets of HN. We used an HSV-1 infection-induced HN mouse model and screened the differentially expressed genes (DEGs) in the DRG and spinal cord using an RNAseq technique. Moreover, bioinformatics methods were used to figure out the signaling pathways and expression regulation patterns of the DEGs enriched. In addition, quantitative real-time RT-PCR and western blot were carried out to further confirm the expression of DEGs. HSV-1 inoculation in mice resulted in mechanical allodynia, thermal hyperalgesia, and cold allodynia, following the infection of HSV-1 in both DRG and spinal cord. Besides, HSV-1 inoculation induced an up-regulation of ATF3, CGRP, and GAL in DRG and activation of astrocytes and microglia in the spinal cord. Moreover, 639 genes were upregulated, 249 genes were downregulated in DRG, whereas 534 genes were upregulated and 12 genes were downregulated in the spinal cord of mice 7 days after HSV-1 inoculation. GO and KEGG enrichment analysis suggested that immune responses and cytokine-cytokine receptor interaction are involved in DRG and spinal cord neurons in mice after HSV-1 infection. In addition, CCL5 and its receptor CCR5 were significantly upregulated in DRG and spinal cord upon HSV-1 infection in mice. And blockade of CCR5 exhibited a significant analgesic effect and suppressed the upregulation of inflammatory cytokines in DRG and spinal cord induced by HSV-1 infection in mice. HSV-1 infection-induced allodynia and hyperalgesia in mice through dysregulation of immune response and cytokine-cytokine receptor interaction mechanism. Blockade of CCR5 alleviated allodynia and hyperalgesia probably through the suppression of inflammatory cytokines. Therefore, CCR5 could be a therapeutic target for the alleviation of HSV-1 infection-induced HN.

Comparative Transcriptome of Dorsal Root Ganglia Reveals Distinct Etiologies of Paclitaxel- and Oxaliplatin-induced Peripheral Neuropathy in Rats.

Chemotherapy-induced peripheral neuropathy is one of the most common side effects of anticancer therapy. It is anticipated that chemotherapies with different mechanisms of action may affect somatosensory neurons differently. This study aimed to explore similar and differential etiologies of oxaliplatin- and paclitaxel-induced neuropathy by comparing the transcriptomes of dorsal root ganglia (DRGs). We retrieved our previously published transcriptome data of DRGs extracted from vehicle-, oxaliplatin- and paclitaxel-treated rats (GSE160543), to analyze in parallel the differentially expressed genes (DEGs) and Gene ontology (GO) terms enrichment. We found that both oxaliplatin and paclitaxel treatments consistently produced mechanical allodynia, thermal hyperalgesia, and cold hyperalgesia in rats. Compared to vehicle, 320 and 150 DEGs were identified after oxaliplatin and paclitaxel treatment, respectively. Only 17 DEGs were commonly dysregulated by the two reagents. Activating transcription factor 3 (Atf3), a marker of nerve injury, was elevated only after paclitaxel treatment. GO analysis suggested that paclitaxel treatment was associated with neuronal changes characterized by numerous terms that are related to synaptic transmission, while oxaliplatin was more likely to affect dividing cells (e.g., the glia) and neuroinflammation. Notably, 29 biological processes GO terms were commonly enriched in response to both drugs. However, 28 out of 29 terms were oppositely modulated. This study suggests that distinct mechanisms underly paclitaxel- and oxaliplatin-induced neuropathy. Paclitaxel might directly affect somatosensory neurons while oxaliplatin primarily targets dividing cells and immune cells.

Transcriptome analysis reveals dysregulation of inflammatory and neuronal function in dorsal root ganglion of paclitaxel-induced peripheral neuropathy rats.

Chemotherapy-induced peripheral neuropathy (CIPN) is the most common side-effect of anti-cancer therapy. To date, there are no clinically effective analgesics that could prevent and treat CIPN. However, the exact pathogenesis of CIPN is still unclear. In the present study, we use the paclitaxel-induced peripheral neuropathy (PIPN) model, aiming to better understand the transcriptomic level of the Dorsal root ganglia (DRG) neurons in rats with PIPN. mRNA from each DRG sample was reverse transcribed to cDNA and sequenced using next-generation high throughput sequencing technology. Quantitative RT-PCR verification was used to confirm the identified Differentially expressed genes (DEGs) in the DRG of PIPN rats. RNAseq results have identified 384 DEGs (adjusted P-value < 0.05; fold change ≥ 2) in the DRG of rats 14 days after paclitaxel injection in total, including 97 up-regulated genes, and 287 down-regulated genes. GO analysis revealed that these DEGs were majorly involved in neuropeptide activity, chemokine receptor activity, defense response, and inflammatory response. Kyoto Encyclopedia of Gene and Genomes analysis showed that neuroactive ligand-receptor interaction and cytokine-cytokine receptor interaction were involved in sensory neurons of rats with PIPN. Besides, comparison analysis identified that 11 DEGs in the PIPN model are shared with either inflammatory pain (Ces1d, Cfd, Retn, and Fam150b) or neuropathic pain (Atf3, Csrp3, Ecel1, Gal, Sprr1a, Tgm1, and Vip). Quantitative RT-PCR results also confirmed the validation of the RNAseq data. These results suggested that neuroactive ligand-receptor interaction and cytokine-cytokine receptor interaction are majorly involved in sensory neurons of rats with PIPN. Immune, inflammatory responses and neuron functional changes are the major pathogenesis of PIPN. Paclitaxel-induced peripheral neuropathy has shared characteristics with both inflammatory pain and neuropathic pain.

Three-Day Continuous Oxytocin Infusion Attenuates Thermal and Mechanical Nociception by Rescuing Neuronal Chloride Homeostasis via Upregulation KCC2 Expression and Function.

Oxytocin (OT) and its receptor are promising targets for the treatment and prevention of the neuropathic pain. In the present study, we compared the effects of a single and continuous intrathecal infusion of OT on nerve injury-induced neuropathic pain behaviours in mice and further explore the mechanisms underlying their analgesic properties. We found that three days of continuous intrathecal OT infusion alleviated subsequent pain behaviours for 14 days, whereas a single OT injection induced a transient analgesia for 30 min, suggesting that only continuous intrathecal OT attenuated the establishment and development of neuropathic pain behaviours. Supporting this behavioural finding, continuous intrathecal infusion, but not short-term incubation of OT, reversed the nerve injury-induced depolarizing shift in Cl- reversal potential via restoring the function and expression of spinal K+-Cl- cotransporter 2 (KCC2), which may be caused by OT-induced enhancement of GABA inhibitory transmission. This result suggests that only continuous use of OT may reverse the pathological changes caused by nerve injury, thereby mechanistically blocking the establishment and development of pain. These findings provide novel evidence relevant for advancing understanding of the effects of continuous OT administration on the pathophysiology of pain.

Selective activation of metabotropic glutamate receptor 7 blocks paclitaxel-induced acute neuropathic pain and suppresses spinal glial reactivity in rats.

RATIONALE: Paclitaxel-induced acute pain syndrome (P-APS), characterized by deep muscle aches and arthralgia, occurs in more than 70% of patients who receive paclitaxel. P-APS can be debilitating for patients and lead to reductions and discontinuation of potentially curable therapy. Despite being relatively common in clinical practice, no clear treatment exists for P-APS and the underlying mechanisms remain poorly defined. Regulation of glutamatergic transmission by metabotropic glutamate receptors (mGluRs) has received growing attention with respect to its role in neuropathic pain. To our knowledge, no study has been conducted on alterations and functions of group III mGluR7 signaling in P-APS. OBJECTIVES: In the present study, we determined whether a single administration of paclitaxel induces glutamatergic alterations and whether mGluR7 activation blocks paclitaxel-induced neuropathic pain by suppressing glial reactivity in the spinal cord. RESULTS: A single paclitaxel injection dose-dependently induced acute mechanical and thermal hypersensitivity, and was associated with increased glutamate level accompanied by reduction in mGluR7 expression in the spinal cord. Selective activation of mGluR7 by its positive allosteric modulator, AMN082, blocked the development of paclitaxel-induced acute mechanical and thermal hypersensitivity, without affecting the normal pain behavior of control rats. Moreover, activation of mGluR7 by AMN082 inhibited glial reactivity and decreased pro-inflammatory cytokine release during P-APS. Abortion of spinal glial reaction to paclitaxel alleviated paclitaxel-induced acute mechanical and thermal hypersensitivity. CONCLUSIONS: There results support the hypothesis that spinal mGluR7 signaling plays an important role in P-APS; Selective activation of mGluR7 by its positive allosteric modulator, AMN082, blocks P-APS in part by reducing spinal glial reactivity and neuroinflammatory process.

Abnormal Intrinsic Brain Activity and Neuroimaging-Based fMRI Classification in Patients With Herpes Zoster and Postherpetic Neuralgia.

Objective: Neuroimaging studies on neuropathic pain have discovered abnormalities in brain structure and function. However, the brain pattern changes from herpes zoster (HZ) to postherpetic neuralgia (PHN) remain unclear. The present study aimed to compare the brain activity between HZ and PHN patients and explore the potential neural mechanisms underlying cognitive impairment in neuropathic pain patients. Methods: Resting-state functional magnetic resonance imaging (MRI) was carried out among 28 right-handed HZ patients, 24 right-handed PHN patients, and 20 healthy controls (HC), using a 3T MRI system. The amplitude of low-frequency fluctuation (ALFF) was analyzed to detect the brain activity of the patients. Correlations between ALFF and clinical pain scales were assessed in two groups of patients. Differences in brain activity between groups were examined and used in a support vector machine (SVM) algorithm for the subjects' classification. Results: Spontaneous brain activity was reduced in both patient groups. Compared with HC, patients from both groups had decreased ALFF in the precuneus, posterior cingulate cortex, and middle temporal gyrus. Meanwhile, the neural activities of angular gyrus and middle frontal gyrus were lowered in HZ and PHN patients, respectively. Reduced ALFF in these regions was associated with clinical pain scales in PHN patients only. Using SVM algorithm, the decreased brain activity in these regions allowed for the classification of neuropathic pain patients (HZ and PHN) and HC. Moreover, HZ and PHN patients are also roughly classified by the same model. Conclusion: Our study indicated that mean ALFF values in these pain-related regions can be used as a functional MRI-based biomarker for the classification of subjects with different pain conditions. Altered brain activity might contribute to PHN-induced pain.

Oxytocin Elicits Itch Scratching Behavior via Spinal GRP/GRPR System.

Oxytocin (OT), a neuropeptide involved in the regulation of complex social and sexual behavior in mammals, has been proposed as a treatment for a number of psychiatric disorders including pain. It has been well documented that central administration of OT elicits strong scratching and grooming behaviors in rodents. However, these behaviors were only described as symptoms, few studies have investigated their underlying neural mechanisms. Thus, we readdressed this question and undertook an analysis of spinal circuits underlying OT-induced scratching behavior in the present study. We demonstrated that intrathecal OT induced robust but transient hindpaw scratching behaviors by activating spinal OT receptors (OTRs). Combining the pre-clinical and clinical evidence, we speculated that OT-induced scratching may be an itch symptom. Further RNAscope studies revealed that near 80% spinal GRP neurons expressed OTRs. OT activated the expression of c-fos mRNA in spinal GRP neurons. Chemical ablation of GRPR neurons significantly reduced intrathecal OT-induced scratching behaviors. Given GRP/GRPR pathway plays an important role in spinal itch transmission, we proposed that OT binds to the OTRs expressed on the GRP neurons, and activates GRP/GRPR pathway to trigger itch-scratching behaviors in mice. These findings provide novel evidence relevant for advancing understanding of OT-induced behavioral changes, which will be important for the development of OT-based drugs to treat a variety of psychiatric disorders.

The antiviral alkaloid berberine ameliorates neuropathic pain in rats with peripheral nerve injury.

Neuropathic pain is a major public health problem. There is a need to develop safer and more effective analgesia compounds with less side effects. Berberine has been used to treat diarrhea and gastroenteritis due to its anti-microbial, anti-motility and anti-secretory properties. Berberine has also been reported to play an analgesic role in some pathological conditions of pain. However, the analgesic roles of berberine in neuropathic pain are still unclear. Therefore, this study aims to explore the analgesic effects of berberine in neuropathic pain. Partial sciatic nerve ligation (pSNL) was performed to create neuropathic pain model. Paw withdrawal responses to mechanical and thermal stimuli were measured using a set of electronic von Frey apparatus and hot plate, respectively. The time that rats spent licking, flinching and lifting its paw during 5 min following capsaicin application was recorded. mRNA and protein expression levels were examined by quantitative RT-PCR and western blot, respectively. Berberine administration (i.p.) increased both mechanical and thermal pain thresholds in a dose-dependent manner. Moreover, berberine administration reversed the mRNA and protein expression of TRPV1 in dorsal root ganglion neurons after peripheral nerve injury. In addition, berberine significantly inhibited capsaicin-induced pain behaviors. The amelioration of neuropathic pain by berberine may be associated with the down-regulation of TRPV1 in DRG of neuropathic pain rats. This study highlighted the potential of berberine in the treatment of neuropathic pain originated in the peripheral nervous system.

(-)-menthol increases excitatory transmission by activating both TRPM8 and TRPA1 channels in mouse spinal lamina II layer.

(-)-menthol, a major form of menthol, is one of the most commonly used chemicals. Many studies have demonstrated that (-)-menthol produces analgesic action through peripheral mechanisms which are mainly mediated by activation of TRPM8. Moreover, intrathecal injection of menthol induces analgesia as well. However, the central actions and mechanisms of (-)-menthol remain unclear. Here, we have investigated the action of (-)-menthol on excitatory synaptic transmission in spinal lamina II layer which plays a pivotal role in modulating nociceptive transmission from the periphery by using patch-clamp technique in mice spinal cord. We found that (-)-menthol increased miniature excitatory postsynaptic current frequency. The frequency increases which (-)-menthol induced were in a dose-dependent manner (EC50: 0.1079 mM). However, neither genetic knockout nor pharmacological inhibition of TRPM8 could block (-)-menthol-induced effects entirely. Furthermore, this increase was also impaired by TRPA1 antagonist HC030031, but abolished utterly by co-application of TRPM8 and TRPA1 antagonist. Our results indicate that (-)-menthol increases the excitatory synaptic transmission by activating either TRPA1 or TRPM8 channels in spinal lamina II layer.

Huachansu suppresses TRPV1 up-regulation and spinal astrocyte activation to prevent oxaliplatin-induced peripheral neuropathic pain in rats.

Chemotherapy-induced peripheral neuropathic pain (CIPNP) is a major dose- and therapy-limiting side effect that is particularly difficult to treat. Huachansu, an aqueous extract from toad skin, is a widely used anti-cancer natural product in China. Clinical findings have established the safety and effectiveness of Huachansu in combination with chemotherapy to promote the therapeutic efficacy while alleviate the side effects, especially cancer-related pain symptoms. Unfortunately, experimental data on the effects and mechanisms of Huachansu in combination with chemotherapy is not available. In this study, the effects of Huachansu were tested in vivo on a rat model of oxaliplatin-induced CIPNP. The results show, a single injection of Huachansu 2.5 g/kg produced a short-term analgesic effect on pre-established oxaliplatin-induced CIPNP after 60 min, as indicated by decreased mechanical and thermal hypersensitivity in comparison to oxaliplatin-treated rats. Repeated doses of Huachansu, given during CIPNP induction, prevented the development of oxaliplatin-induced CIPNP. This prophylactic effect of Huachansu was associated with suppressed oxaliplatin-induced TRPV1 up-regulation in the dorsal root ganglia and spinal astrocyte activation. These findings reveal Huachansu therapeutic potential in treating and preventing CIPNP.

Cinobufacini protects against paclitaxel-induced peripheral neuropathic pain and suppresses TRPV1 up-regulation and spinal astrocyte activation in rats.

Chemotherapy-induced peripheral neuropathic pain is a major limiting factor affecting cancer patients. No effective treatment is currently available. Cinobufacini, an aqueous extract from toad skin, is a widely used anti-cancer drug in China. Clinical evidence has demonstrated the safety and effectiveness of cinobufacini in combination with chemotherapy to promote the therapeutic efficacy while alleviating side effects, especially cancer-related pain symptoms. In this study, the effects of cinobufacini were investigated in a rat model of paclitaxel-induced peripheral neuropathic pain (PIPNP) to better understand and expand its clinical application. A single injection of cinobufacini (2.5 g/kg, i.p.) alleviated pre-established PIPNP, as indicated by decreased mechanical and thermal hypersensitivity compared with paclitaxel-treated rats. Repeated cinobufacini (1.25 and 2.5 g/kg, i.p.), given during the induction of PIPNP, prevented the establishment of paclitaxel-induced mechanical and thermal hypersensitivity. This preventative effect was associated with suppressed paclitaxel-induced TRPV1 up-regulation and spinal astrocyte activation, as well as decreased production of spinal TNF-α and IL-1ß. These findings reveal cinobufacini as a therapeutic potential to treat and prevent paclitaxel-induced peripheral neuropathic pain.

Oxytocin Relieves Neuropathic Pain Through GABA Release and Presynaptic TRPV1 Inhibition in Spinal Cord.

Objective: Oxytocin (OT) is synthesized within the paraventricular nucleus and supraoptic nucleus of the hypothalamus. In addition to its role in uterine contraction, OT plays an important antinociceptive role; however, the underlying molecular mechanisms of antinociceptive role of OT remain elusive. We hypothesized that the antinociceptive effect of OT on neuropathic pain may occur via inhibition of TRPV1 activation in the spinal cord. The present study explores the antinociceptive role of OT and its mechanisms in neuropathic pain. Methods: Partial sciatic nerve ligation (pSNL) was performed to induce neuropathic pain. Animal behaviors were measured using a set of electronic von Frey apparatus and hot plate. Electrophysiological recordings and molecular biological experiments were performed. Results: Intrathecal administration of OT alleviated both mechanical allodynia and thermal hyperalgesia in pSNL rats (n = 6, per group, P < 0.0001, saline vs. OT group). Electrophysiological data revealed that OT significantly inhibited the enhancement of frequency and amplitude of spontaneous excitatory post-synaptic currents induced presynaptically by TRPV1 activation in the spinal cord. Moreover, the inhibitory effect of OT on capsaicin-induced facilitation of excitatory transmission was blocked by co-treatment with saclofen, while intrathecal administration of OT dramatically inhibited capsaicin-induced ongoing pain in rats, (n = 6, per group, P < 0.0001, saline vs. OT group). The paw withdrawal latency in response to heat stimulation was significantly impaired in TRPV1KO mice 3 days after pSNL upon OT (i.t.) treatment, compared with wild type mice (n = 6, P < 0.05). Finally, OT prevented TRPV1 up-regulation in spinal cords of pSNL model rats. Conclusion: OT relieves neuropathic pain through GABA release and presynaptic TRPV1 inhibition in the spinal cord. OT and its receptor system might be an intriguing target for the treatment and prevention of neuropathic pain.