Spinal high-mobility group box-1 induces long-lasting mechanical hypersensitivity through the toll-like receptor 4 and upregulation of interleukin-1ß in activated astrocytes.

Intrathecal treatment with recombinant high-mobility group box-1 (rHMGB1) in naïve mice leads to a persistent and significantly decreased hind paw withdrawal threshold to mechanical stimuli, suggesting that spinal HMGB1 evokes abnormal pain processing. By contrast, repeated intrathecal treatment with anti-HMGB1 antibody significantly reverses hind paw mechano-hypersensitivity in mice with a partial sciatic nerve ligation (PSNL). By contrast, the cellular mechanism by which spinal HMGB1 induces neuropathic pain has yet to be fully elaborated. The current study tested the hypothesis that spinal HMGB1 could induce mechanical hypersensitivity through the activation of specific receptor in glial cells. Intrathecal pretreatment with toll-like receptor (TLR) 4 inhibitors, but not TLR5, receptor for advanced glycation end-products and C-X-C chemokine receptor type 4 inhibitors, prevented rHMGB1-evoked mechanical hypersensitivity. Activation of spinal astrocytes appears to be crucial for the mechanism of action of rHMGB1 in naïve mice, as intrathecal pretreatment with astrocytic inhibitors prevented the rHMGB1-induced mechanical hypersensitivity. Interleukin-1ß (IL-1ß) was up-regulated within activated astrocytes and block of TLR4 prevented the upregulation of IL-1ß. Interleukin-1ß appears to be secreted by activated astrocytes, as IL-1ß neutralizing antibody prevented rHMGB1-induced mechanical hypersensitivity. Furthermore, intrathecal pretreatment with either MK801 or gabapentin prevented the rHMGB1-induced mechanical hypersensitivity, suggesting roles for spinal glutamate and the N-methyl-d-aspartate receptor in the mediation of rHMGB1-induced mechanical hypersensitivity. Thus, the current findings suggest that spinal HMGB1 upregulates IL-1ß in spinal astrocytes through a TLR4-dependent pathway and increases glutamatergic nociceptive transduction. These spinal mechanisms could be key steps that maintain neuropathic pain.

Downregulation of spinal astrocytic connexin43 leads to upregulation of interleukin-6 and cyclooxygenase-2 and mechanical hypersensitivity in mice.

Connexin43 (Cx43), involved in intercellular signaling, is expressed in spinal dorsal horn astrocytes and crucial in the maintenance of neuropathic pain. Downregulation of spinal astrocytic Cx43 in mice enhances glutamatergic neurotransmission by decreasing glutamate transporter GLT-1 expression, resulting in cutaneous hypersensitivity. Decreased expression of astrocytic Cx43 could lead to altered expression of other nociceptive molecules. Transfection of Cx43-targeting siRNA in cultured spinal astrocytes increased expression of the pronociceptive cytokine interleukin-6 (IL-6) and the prostaglandin synthesizing enzyme cyclooxygenase-2 (COX-2). Increased expression of IL-6 and COX-2 was due to decreased Cx43 expression rather than due to diminished Cx43 channel function. In mice, downregulation of spinal Cx43 expression by intrathecal treatment with Cx43-targeting siRNA increased IL-6 and COX-2 expression and induced hind paw mechanical hypersensitivity. Cx43 siRNA-induced mechanical hypersensitivity was attenuated by intrathecal treatment with anti-IL-6 neutralizing antibody and intraperitoneal treatment of selective COX-2 inhibitor celecoxib, demonstrating that these molecules play a role in nociceptive processing following Cx43 downregulation. Restoring spinal Cx43 by intrathecal injection of an adenovirus vector expressing Cx43 in mice with a partial sciatic nerve ligation reduced spinal IL-6 and COX-2 expression. Suppression of glycogen synthase kinase-3ß (GSK-3ß), a serine/threonine protein kinase, prevented upregulation of IL-6 and COX-2 expression induced by Cx43 downregulation in both cultured astrocytes and in mouse spinal dorsal horn. Inhibition of spinal GSK-3ß also ameliorated Cx43 siRNA-induced mechanical hypersensitivity. The current findings indicate that downregulation of spinal astrocytic Cx43 leads to changes in spinal expression of pronociceptive molecules underlying the maintenance of pain following nerve injury.

Effect of VEGF (vascular endothelial growth factor) on expression of IL-8 (interleukin-8), IL-1beta and their receptors in bovine theca cells.

Cytokines such as VEGF (vascular endothelial growth factor) and ILs (interleukins) are involved in follicular development in the mammalian ovary. The aim of the present study is to examine the transcripts of IL-8 and -1 that differ during follicular development; the relationships between IL-8, IL-1 and VEGF in theca cells is still unknown. We first examined the gene expression of IL-8, IL-1beta and their respective receptors, CXCR1 and IL-1R1 in the theca cells of PRF (preselection) and POF (postselection follicles) from the bovine ovary. Expression of IL-8 and CXCR1 genes were observed in POF, whereas expression of IL-1beta and IL-1R1 genes was observed in both follicles. Secondly, we examined the effects of VEGF on the expression of IL-8, IL-1beta and their receptors genes in cultured bovine theca cells. mRNA expression was quantified by using real-time PCR methods. VEGF stimulates the expression of IL-8 and CXCR1 mRNA. However, VEGF down-regulates the expression of CXCR2 mRNA during the culture period. Expression of IL-1beta and -1R1 mRNA was induced in the cultured theca cells at 48 h. Our data demonstrate that VEGF stimulated the expression of the IL-8 and CXCR1 genes and that CXCR2 expression was suppressed by VEGF, suggesting a follicle stage-dependent expression pattern for the IL-8 system. Furthermore, our results suggest that the transcription system for CXCR genes may have different pathways of VEGF stimulation in bovine theca cells. Taken together, our data suggested that VEGF is associated with the IL system in theca cells in bovine ovary.

Stimulation of spinal dorsal horn ß2-adrenergic receptor ameliorates neuropathic mechanical hypersensitivity through a reduction of phosphorylation of microglial p38 MAP kinase and astrocytic c-jun N-terminal kinase.

The noradrenaline-adrenergic system has a crucial role in controlling nociceptive transduction at the spinal level. While α-adrenergic receptors are known to regulate nociceptive neurotransmitter release at the spinal presynaptic level, it is not entirely clear whether ß-adrenergic receptors are involved in controlling pain transduction at the spinal level as well. The current study elucidated a role of ß-adrenergic receptors in neuropathic pain in mice following a partial sciatic nerve ligation (PSNL). In addition, the cellular and intracellular signaling cascade induced by ß-adrenergic receptors in neuropathic mice was elaborated. Intrathecal injection of isoproterenol (1 nmol), a nonselective ß-adrenergic receptor agonist, briefly ameliorated hind paw mechanical hypersensitivity of PSNL mice. Isoproterenol's antinociceptive effect was mediated through ß2-adrenergic receptors since pretreatment with ICI118551, a selective ß2-adrenergic receptor antagonist, but not with CGP20712A, a selective ß1-adrenergic receptor antagonist, significantly attenuated isoproterenol's effect. Furthermore, intrathecal treatment with a selective ß2-adrenergic receptor agonist, terbutaline, but not a selective ß1-adrenergic receptor agonist, dobutamine, also significantly ameliorated neuropathic pain. Fourteen days after PSNL, increased phosphorylation of both p38 Mitogen-activated protein kinase (MAPK) in microglia and c-jun N-terminal kinase (JNK) in astrocytes of ipsilateral spinal dorsal horn were observed. Phosphorylation of both microglial p38 MAPK and astrocytic JNK were downregulated by stimulation of the ß2-adrenergic receptor. Together, these results suggest that spinal ß2-adrenergic receptor have an inhibitory role in neuropathic nociceptive transduction at the spinal level through a downregulation of glial activity, perhaps through modulation of MAP kinases phosphorylation. Thus, targeting of ß2-adrenergic receptors could be an effective therapeutic strategy in treating neuropathic pain.

Lycopene ameliorates neuropathic pain by upregulating spinal astrocytic connexin 43 expression.

AIM: Peripheral nerve injury upregulates tumor necrosis factor (TNF) expression. In turn, connexin 43 (Cx43) expression in spinal astrocytes is downregulated by TNF. Therefore, restoration of spinal astrocyte Cx43 expression to normal level could lead to the reduction of nerve injury-induced pain. While the non-provitaminic carotenoid lycopene reverses thermal hyperalgesia in mice with painful diabetic neuropathy, the antinociceptive mechanism is not entirely clear. The current study evaluated whether the antinociceptive effect of lycopene is mediated through the modulation of Cx43 expression in spinal astrocytes. MAIN METHODS: The effect of lycopene on Cx43 expression was examined in cultured rat spinal astrocytes. The effect of intrathecal lycopene on Cx43 expression and neuropathic pain were evaluated in mice with partial sciatic nerve ligation (PSNL). KEY FINDINGS: Treatment of cultured rat spinal astrocytes with lycopene reversed TNF-induced downregulation of Cx43 protein expression through a transcription-independent mechanism. By contrast, treatment of cultured spinal astrocytes with either pro-vitamin A carotenoid ß-carotene or antioxidant N-acetyl cysteine had no effect on TNF-induced downregulation of Cx43 protein expression. In addition, repeated, but not single, intrathecal treatment with lycopene of mice with a partial sciatic nerve ligation significantly prevented not only the downregulation of Cx43 expression in spinal dorsal horn but mechanical hypersensitivity as well. SIGNIFICANCE: The current findings suggest a significant spinal mechanism that mediates the analgesic effect of lycopene, through the restoration of normal spinal Cx43 expression.

Preoperative assessment of the relationship between the mandibular third molar and the mandibular canal by axial computed tomography with coronal and sagittal reconstruction.

OBJECTIVE: We sought to evaluate the relationship between the mandibular third molar and the mandibular canal by using axial computed tomography with coronal and sagittal reconstruction for third molar surgery. STUDY DESIGN: Forty-seven impacted third molars in 41 patients were found in close association with the mandibular canal during a panoramic radiographic assessment. The relationship between the mandibular third molar and the mandibular canal was evaluated by using computed tomography and compared in terms of operative exposure of the inferior alveolar nerve and postoperative labial dysesthesia. RESULTS: Twenty-four (51%) mandibular canals were buccal relative to the third molar, 12 were lingual, 9 were inferior, and 2 were between roots. At the time of the surgical procedure, the inferior alveolar nerve was visible in 7 patients. Postoperative lower lip dysesthesia occurred in 1 patient whose mandibular canal was in the lingual position. CONCLUSIONS: Axial computed tomography with coronal and sagittal reconstruction provides useful information to surgeons regarding the relationship between the mandibular third molar and the mandibular canal.

Occult fractures of articular eminence and glenoid fossa presenting as temporomandibular disorder: a case report.

We report an unusual case of occult fractures of the articular eminence and glenoid fossa due to a previous traffic accident presenting as a temporomandibular disorder. A 24-year-old Japanese man was referred for trismus and pain in the right temporomandibular joint, and was suspected of having temporomandibular disorder. Although the magnetic resonance image did not show displacement of the articular disk, T2-weighted images revealed disruption of the cortical low-intensity line of the right articular eminence. On a computerized tomography (CT) scan, an isolated bone fragment of the right articular eminence was clearly seen, and fractures of the right glenoid fossa and articular eminence were noted. The patient was treated by instructing him not to open his mouth widely and to remain on a soft diet for 4 weeks. A CT examination performed 1 year after the treatment showed complete healing of the fracture in the right articular eminence and glenoid fossa.