Animal Studies

Necrostatin-1 is an inhibitor of necroptosis, a form of programmed cell death that has been reported to be involved in various neurological diseases. Presently, the role of necroptosis in neuropathic pain induced by peripheral nerve injury is still unclear. This study was focused on investigating the potential effects of necroptosis in the development and progression of neuropathic pain in a rat model and the possible neuroprotective effects of necrostatin-1 in neuropathic pain. The results indicated that the necroptosis-related proteins RIP1 and RIP3 significantly increased postoperation in the spinal cord in a neuropathic pain model and peaked 7 days postoperation, which was consistent with the time-dependent changes of hyperalgesia. Additionally, we found that peripheral nerve injury-related behavioral and biochemical changes were significantly reduced by necrostatin-1. In particular, hyperalgesia was attenuated, and the levels of RIP1 and RIP3 were decreased. Furthermore, the ultrastructure of necrotic cell death and neuroinflammation were alleviated by necrostatin-1. Collectively, these results suggest that necroptosis is an important mechanism of cell death in neuropathic pain induced by peripheral nerve injury and that necrostatin-1 may be a promising neuroprotective treatment for neuropathic pain.

Complex regional pain syndrome type-I (CRPS-I) is a progressive and devastating pain condition, which remains clinically challenging. The mechanisms of CRPS-I still remain largely unknown. We aim to identify transcriptome profiles of genes relevant to pain mechanisms and major pathways involved in CRPS-I. A rat model of chronic post-ischemia pain (CPIP) was established to mimic CRPS-I. RNA-sequencing (RNA-Seq) was used to profile transcriptome of L4-6 dorsal root ganglia (DRGs) of a rat model of CRPS-I. CPIP model rats developed persistent mechanical/thermal hyperalgesia in ipsilateral hind paw. RNA-Seq identified a total of 295 differentially expressed genes (DEGs), including 195 up- and 100 downregulated, in ipsilateral DRGs of CPIP rats compared with sham rats. The expression of several representative genes was confirmed by qPCR. Functional analysis of DEGs revealed that the most significant enriched biological processes of upregulated genes include response to lipopolysaccharide, inflammatory response and cytokine activity, which are all important mechanisms mediating pain. We further screened DEGs implicated in pain progress, genes enriched in small- to medium-sized sensory neurons and enriched in TRPV1-lineage nociceptors. By comparing our dataset with other published datasets of neuropathic or inflammatory pain models, we identified a core set of genes and pathways that extensively participate in CPIP and other neuropathic pain states. Our study identified transcriptome gene changes in DRGs of an animal model of CRPS-I and could provide insights into identifying promising genes or pathways that can be potentially targeted to ameliorate CRPS-I.

The present study aimed to identify microRNAs (miRNAs) that are involved in neuropathic pain and predict their corresponding roles in the pathogenesis and development process of neuropathic pain. The rat model of neuropathic pain caused by spared nerve injury (SNI) was established in Sprague-Dawley male rats, followed by small RNA sequencing of the L3-L6 dorsal root ganglion. Real-time PCR was performed to validate the differently expressed miRNAs. Functional verification was performed by intrathecally injecting the animals with miRNA agomir. A total of 72 differentially expressed miRNAs were identified in the SNI rats, including 33 upregulated and 39 downregulated miRNAs. The results of qPCR further verified the expression levels of rno-miR-6215 (P=0.015), rno-miR-1224 (P=0.030), rno-miR-1249 (P=0.038), and rno-miR-488-3p (P=0.048), which were all significantly downregulated in the SNI rats compared to the control ones. The majority of differentially expressed miRNAs were associated with phosphorylation, intracellular signal transduction, and cell death. Target prediction, Gene Ontology, and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses suggested that these differentially expressed miRNAs targeted genes that are related to axon guidance, focal adhesion, and Ras and Wnt signaling pathways. Moreover, miR-1224 agomir significantly alleviated SNI-induced neuropathic pain. The current findings provide new insights into the role of miRNAs in the pathogenesis of neuropathic pain.

Chronic pain is a major comorbidity of sickle cell disease (SCD). Acupuncture, a non-opioid and non-addictive therapy to treat pain, was found to reduce pain in the majority (80%) of SCD patients in an earlier retrospective review. We observed that electroacupuncture (EA) decreased hyperalgesia in transgenic mice with SCD with varied analgesia from high to moderate to no response. Interestingly, poor responders exhibited high levels of substance P (SP), a mediator of chronic pain, as well as active p38 MAPK in spinal cords. The present study aimed to investigate the roles of inhibition of SP and SP-activated p38 MAPK in chronic pain in sickle mice that are poorly responsive to EA intervention (moderate/non-responders). Humanized mouse model with SCD defined as moderate- and non-responders to EA were intraperitoneally administered with antagonist of SP receptor NK1R (netupitant, 10 mg/kg/day, i.p.) or p38 MAPK inhibitor (SB203580, 10 mg/kg/day, i.p.) alone or in combination with EA (acupoint GB30, every 3rd day until day 12). Hyperalgesia to mechanical, thermal and cold stimuli, as well as deep tissue were measured. Phosphorylated p38 MAPK (phospho-p38 MAPK) in the lumbar spinal cord was quantified using western blotting. Phospho-p38 MAPK nuclear translocation in spinal dorsal horn was examined using immunohistochemical staining and confocal microscopy. In EA poor-responders, combined treatment with EA and netupitant significantly enhanced the analgesic effects of EA in poor-responders on mechanical, heat, cold, and deep tissue pain, and decreased phosphorylation of p38 MAPK in lumbar spinal cords and its nuclear translocation in the spinal dorsal horn. Furthermore, combined treatment with EA and SB203580 significantly improved analgesic effects of EA on mechanical and heat hyperalgesia, but not cold or deep tissue hyperalgesia. However, additional EA treatment only, or administration of either netupitant or SB203580 alone did not lead to analgesic effects. These results suggest a pivotal role of SP in maintaining the chronic pain in SCD via spinal phospho-p38 MAPK signaling, which may hinder the effect of EA in poor responders. Inhibition of SP signaling pathway or activity of p38 MAPK significantly improved the EA analgesia In EA poor-responders with SCD, which provides a promising way to treat the chronic pain in patients with SCD.

This study determined the role of miR-1906 in neuropathic pain and proliferation in neuronal cells using a chronic constriction injury (CCI)-induced neuropathic pain (NP) rat model.

Using a high resolution hybridization technique we have measured , , and transcripts in mouse dorsal root ganglion (DRG) neurons. Consistent with previous studies, transcripts were highly expressed in DRG neurons of all sizes, including most notably the largest diameter neurons implicated in mediating touch and proprioception. In contrast, transcripts were selectively expressed in smaller DRG neurons, which are implicated in mediating nociception. Moreover, the small neurons expressing were mostly distinct from those neurons that strongly expressed , one of the channels implicated in heat-nociception. Interestingly, while and expressing neurons form essentially non-overlapping populations, showed co-expression in both populations. Using an functional test for the selective expression, we found that Yoda1, a PIEZO1-specific agonist, induced a mechanical hyperalgesia that displayed a significantly prolonged time course compared with that induced by capsaicin, a TRPV1-specific agonist. Taken together, our results indicate that PIEZO1 should be considered a potential candidate in forming the long sought channel mediating mechano-nociception.

Complex regional pain syndrome (CRPS) is a rare but refractory pain disorder. Recent advanced information retrieval studies using text-mining and network analysis have suggested nuclear factor kappa B (NFκB) as a possible central mediator of CRPS. The brain is also known to play important roles in CRPS. The aim of this study was to evaluate changes in cerebral NFκB in rats with CRPS.