Recent advances in the treatment of oral ulcerative mucositis from clinical and basic perspectives.

BACKGROUND: Oral ulcerative mucositis (OUM) is common in patients with cancer, particularly in those undergoing chemoradiation therapy. The effective management of OUM is crucial for continuous cancer care and patient well-being. Recent studies have advanced our understanding of the causes, leading to clinical trials toward novel treatments. This review focuses on the contemporary therapeutic landscape, and provides the latest insights into the mechanisms of mucosal healing and pain. HIGHLIGHTS: Management strategies for oral ulcerative mucositis in patients with cancer include maintaining good oral hygiene, reducing mucosal irritation against radiation, and using various topical analgesic treatments, including herbal medicines. However, the current management practices have limitations that necessitate the development of more efficacious and novel treatments. Molecular research on transient receptor potential (TRP) channels in the oral mucosa is crucial for understanding the mechanisms of wound healing and pain in patients with OUM. Targeting TRP subfamily V member 3 (V3) and TRPV4 can enhance wound healing through re-epithelialization. The suppression of TRPV1, TRPA1, and TRPV4 may be effective in alleviating OUM-induced pain. CONCLUSION: Research advancements have improved our understanding and potentially led to novel treatments that offer symptomatic relief. This progress highlights the importance of collaborations between clinical researchers and scientists in the development of innovative therapies.

Analgesic effect of linalool odor on oral ulcerative mucositis-induced pain in rats.

Oral ulcerative mucositis (OUM) induces severe pain, leading to a low quality of life. Linalool odor exposure has recently been reported to suppress inflammatory pain in the hind paws. However, the analgesic effect of linalool odor on orofacial pain remains unclear. In this study, we examined the mechanism underlying the analgesic effect of linalool odor on oral pain caused by OUM using nocifensive behavioral and immunohistochemical analyses in rats. OUM was developed by treating the labial fornix region of the inferior incisors with acetic acid. Linalool at 1% was exposed for 5 min at 30 min before nocifensive behavioral measurements. OUM induced spontaneous pain and mechanical allodynia, which were suppressed by the linalool odor. Mechanical allodynia in the hind paw following the injection of complete Freund's adjuvant was also suppressed by linalool odor. Application of lidocaine to the olfactory bulb attenuated the inhibition of spontaneous pain and hyperactivation of trigeminal spinal nucleus caudalis neurons in OUM model rats. Linalool odor exposure-induced neuronal activation in the locus coeruleus (LC) of OUM model rats was decreased by lidocaine application to the olfactory bulb. The decrease in neuronal activation in the LC was attenuated by the administration of orexin 1 receptor (OX-1) antagonist to the LC. These results suggest that linalool odor stimulation through the olfactory pathway activates LC neurons via OX-1 signaling, leading to the suppression of OUM-induced oral pain.

Involvement of interferon gamma signaling in spinal trigeminal caudal subnucleus astrocyte in orofacial neuropathic pain in rats with infraorbital nerve injury.

Background: Trigeminal nerve injury causes orofacial pain that can interfere with activities of daily life. However, the underlying mechanism remains unknown, and the appropriate treatment has not been established yet. This study aimed to examine the involvement of interferon gamma (IFN-γ) signaling in the spinal trigeminal caudal subnucleus (Vc) in orofacial neuropathic pain. Methods: Infraorbital nerve (ION) injury (IONI) was performed in rats by partial ION ligation. The head-withdrawal reflex threshold (HWT) to mechanical stimulation of the whisker pad skin was measured in IONI or sham rats, as well as following a continuous intracisterna magna administration of IFN-γ and a mixture of IFN-γ and fluorocitrate (inhibitor of astrocytes activation) in naïve rats, or an IFN-γ antagonist in IONI rats. The IFN-γ receptor immunohistochemistry and IFN-γ Western blotting were analyzed in the Vc after IONI or sham treatment. The glial fibrillary acid protein (GFAP) immunohistochemistry and Western blotting were also analyzed after administration of IFN-γ and the mixture of IFN-γ and fluorocitrate. Moreover, the change in single neuronal activity in the Vc was examined in the IONI, sham, and IONI group administered IFN-γ antagonist. Results: The HWT decreased after IONI. The IFN-γ and IFN-γ receptor were upregulated after IONI, and the IFN-γ receptor was expressed in Vc astrocytes. IFN-γ administration decreased the HWT, whereas the mixture of IFN-γ and fluorocitrate recovered the decrement of HWT. IFN-γ administration upregulated GFAP expression, while the mixture of IFN-γ and fluorocitrate recovered the upregulation of GFAP expression. IONI significantly enhanced the neuronal activity of the mechanical-evoked responses, and administration of an IFN-γ antagonist significantly inhibited these enhancements. Conclusions: IFN-γ signaling through the receptor in astrocytes is a key mechanism underlying orofacial neuropathic pain associated with trigeminal nerve injury. These findings will aid in the development of therapeutics for orofacial neuropathic pain.

Involvement of oxidative stress in orofacial mechanical pain hypersensitivity following neonatal maternal separation in rats.

Patients with persistent pain have sometimes history of physical abuse or neglect during infancy. However, the pathogenic mechanisms underlying orofacial pain hypersensitivity associated with early-life stress remain unclear. The present study focused on oxidative stress and investigated its role in pain hypersensitivity in adulthood following early-life stress. To establish an early-life stress model, neonatal pups were separated with their mother in isolated cages for 2 weeks. The mechanical head-withdrawal threshold (MHWT) in the whisker pad skin of rats received maternal separation (MS) was lower than that of non-MS rats at postnatal week 7. In MS rats, the expression of 8-hydroxy-deoxyguanosine, a marker of DNA oxidative damage, was enhanced, and plasma antioxidant capacity, but not mitochondrial complex I activity, decreased compared with that in non-MS rats. Reactive oxygen species (ROS) inactivation and ROS-sensitive transient receptor potential ankyrin 1 (TRPA1) antagonism in the whisker pad skin at week 7 suppressed the decrease of MHWT. Corticosterone levels on day 14 increased in MS rats. Corticosterone receptor antagonism during MS periods suppressed the reduction in antioxidant capacity and MHWT. The findings suggest that early-life stress potentially induces orofacial mechanical pain hypersensitivity via peripheral nociceptor TRPA1 hyperactivation induced by oxidative stress in the orofacial region.

M2 macrophage-derived cathepsin S promotes peripheral nerve regeneration via fibroblast-Schwann cell-signaling relay.

BACKGROUND: Although peripheral nerves have an intrinsic self-repair capacity following damage, functional recovery is limited in patients. It is a well-established fact that macrophages accumulate at the site of injury. Numerous studies indicate that the phenotypic shift from M1 macrophage to M2 macrophage plays a crucial role in the process of axon regeneration. This polarity change is observed exclusively in peripheral macrophages but not in microglia and CNS macrophages. However, the molecular basis of axonal regeneration by M2 macrophage is not yet fully understood. Herein, we aimed to identify the M2 macrophage-derived axon regeneration factor. METHODS: We established a peripheral nerve injury model by transection of the inferior alveolar nerve (IANX) in Sprague-Dawley rats. Transcriptome analysis was performed on the injured nerve. Recovery from sensory deficits in the mandibular region and histological reconnection of IAN after IANX were assessed in rats with macrophage depletion by clodronate. We investigated the effects of adoptive transfer of M2 macrophages or M2-derived cathepsin S (CTSS) on the sensory deficit. CTSS initiating signaling was explored by western blot analysis in IANX rats and immunohistochemistry in co-culture of primary fibroblasts and Schwann cells (SCs). RESULTS: Transcriptome analysis revealed that CTSS, a macrophage-selective lysosomal protease, was upregulated in the IAN after its injury. Spontaneous but partial recovery from a sensory deficit in the mandibular region after IANX was abrogated by macrophage ablation at the injured site. In addition, a robust induction of c-Jun, a marker of the repair-supportive phenotype of SCs, after IANX was abolished by macrophage ablation. As in transcriptome analysis, CTSS was upregulated at the injured IAN than in the intact IAN. Endogenous recovery from hypoesthesia was facilitated by supplementation of CTSS but delayed by pharmacological inhibition or genetic silencing of CTSS at the injured site. Adoptive transfer of M2-polarized macrophages at this site facilitated sensory recovery dependent on CTSS in macrophages. Post-IANX, CTSS caused the cleavage of Ephrin-B2 in fibroblasts, which, in turn, bound EphB2 in SCs. CTSS-induced Ephrin-B2 cleavage was also observed in human sensory nerves. Inhibition of CTSS-induced Ephrin-B2 signaling suppressed c-Jun induction in SCs and sensory recovery. CONCLUSIONS: These results suggest that M2 macrophage-derived CTSS contributes to axon regeneration by activating SCs via Ephrin-B2 shedding from fibroblasts.

PAR2-dependent phosphorylation of TRPV4 at the trigeminal nerve terminals contributes to tongue cancer pain.

OBJECTIVE: This study aimed to clarify the interactions between the tongue and primary afferent fibers in tongue cancer pain. METHODS: A pharmacological analysis was conducted to evaluate mechanical hypersensitivity of the tongues of rats with squamous cell carcinoma (SCC). Changes in trigeminal ganglion (TG) neurons projecting to the tongue were analyzed using immunohistochemistry and western blotting. RESULTS: SCC inoculation of the tongue caused persistent mechanical sensitization and tumor formation. Trypsin expression was significantly upregulated in cancer lesions. Continuous trypsin inhibition or protease-activated receptor 2 (PAR2) antagonism in the tongue significantly inhibited SCC-induced mechanical sensitization. No changes were observed in PAR2 and transient receptor potential vanilloid 4 (TRPV4) levels in the TG or the number of PAR2-and TRPV4-expressing TG neurons after SCC inoculation. In contrast, the relative amount of phosphorylated TRPV4 in the TG was significantly increased after SCC inoculation and abrogated by PAR2 antagonism in the tongue. TRPV4 antagonism in the tongue significantly ameliorated the mechanical sensitization caused by SCC inoculation. CONCLUSIONS: Our findings indicate that tumor-derived trypsin sensitizes primary afferent fibers by PAR2 stimulation and subsequent TRPV4 phosphorylation, resulting in severe tongue pain.

Microglia cause structural remodeling of noradrenergic axon in the trigeminal spinal subnucleus caudalis after infraorbital nerve injury in rats.

The dysfunction of descending noradrenergic (NAergic) modulation in second-order neurons has long been observed in neuropathic pain. In clinical practice, antidepressants that increase noradrenaline levels in the synaptic cleft are used as first-line agents, although adequate analgesia has not been occasionally achieved. One of the hallmarks of neuropathic pain in the orofacial regions is microglial abnormalities in the trigeminal spinal subnucleus caudalis (Vc). However, until now, the direct interaction between descending NAergic system and Vc microglia in orofacial neuropathic pain has not been explored. We found that reactive microglia ingested the dopamine-ß-hydroxylase (DßH)-positive fraction, NAergic fibers, in the Vc after infraorbital nerve injury (IONI). Major histocompatibility complex class I (MHC-I) was upregulated in Vc microglia after IONI. Interferon-γ (IFNγ) was de novo induced in trigeminal ganglion (TG) neurons following IONI, especially in C-fiber neurons, which conveyed to the central terminal of TG neurons. Gene silencing of IFNγ in the TG reduced MHC-I expression in the Vc after IONI. Intracisternal administration of exosomes from IFNγ-stimulated microglia elicited mechanical allodynia and a decrease in DßH in the Vc, which did not occur when exosomal MHC-I was knocked down. Similarly, in vivo MHC-I knockdown in Vc microglia attenuated the development of mechanical allodynia and a decrease in DßH in the Vc after IONI. These results show that microglia-derived MHC-I causes a decrease in NAergic fibers, culminating in orofacial neuropathic pain.

Hepcidin expression in the trigeminal ganglion and the oral mucosa in an oral ulcerative mucositis rat model.

Severe intraoral pain induces difficulty in eating and speaking, leading to a decline in the quality of life. However, the molecular mechanisms underlying intraoral pain remain unclear. Here, we investigated gene modulation in the trigeminal ganglion and intraoral pain-related behavior in a rat model of acetic acid-induced oral ulcerative mucositis. Oral ulceration was observed on day 2 after acetic acid treatment to the oral mucosa of male Wistar rats, causing spontaneous pain and mechanical allodynia. Deoxyribonucleic acid microarray analysis of trigeminal ganglion tissue indicated that Hamp (a hepcidin gene that regulates cellular iron transport) was the most upregulated gene. In the oral ulcerative mucositis model, the upregulation of Hamp was also induced in the ulcer region but not in the liver, with no increase in hepcidin levels in the plasma and saliva, indicating that hepcidin was produced locally in the ulcer region in the model. Systemic antibiotic pretreatment did not increase the mRNA levels of Hamp in the trigeminal ganglion and ulcer regions. Hepcidin injection into the oral mucosa enhanced neuronal excitability in response to noxious mechanical stimulation of the oral mucosa in trigeminal spinal subnucleus interpolaris/caudalis neurons. These results imply that oral ulcerative mucositis induces oral mucosal pain because of infectious inflammation of the ulcerative area and potentiates Hamp, which represents anti-bacterial and anti-peptidase gene expression in the ulcer region and trigeminal ganglion. The regulation of cellular iron transport by hepcidin is likely involved in oral ulcerative mucositis-induced pain.

Neonatal Injury Modulates Incisional Pain Sensitivity in Adulthood: An Animal Study.

Neonatal pain experiences including traumatic injury influence negatively on development of nociceptive circuits, resulting in persistent pain hypersensitivity in adults. However, the detailed mechanism is not yet well understood. In the present study, to clarify the pathogenesis of orofacial pain hypersensitivity associated with neonatal injury, the involvement of the voltage-gated sodium channel (Nav) 1.8 and the C-C chemokine ligand 2 (CCL2)/C-C chemokine receptor 2 (CCR2) signaling in the trigeminal ganglion (TG) in facial skin incisional pain hypersensitivity was examined in 190 neonatal facial-injured and sham male rats. The whisker pad skin was incised on postnatal day 4 and week 7 (Incision-Incision group). Compared to the group without neonatal incision (Sham-Incision group), mechanical hypersensitivity in the whisker pad skin was enhanced in Incision-Incision group. The number of Nav1.8-immunoreactive TG neurons and the amount of CCL2 expressed in the macrophages and satellite glial cells in the TG were increased on day 14 after re-incision in the Incision-Incision group, compared with Sham-Incision group. Blockages of Nav1.8 in the incised region and CCR2 in the TG suppressed the enhancement of mechanical hypersensitivity in the Incision-Incision group. Administration of CCL2 into the TG enhanced mechanical hypersensitivity in the Sham-Sham, Incision-Sham and Sham-Incision group. Our results suggest that neonatal facial injury accelerates the TG neuronal hyperexcitability following orofacial skin injury in adult in association with Nav1.8 overexpression via CCL2 signaling, resulting in the enhancement of orofacial incisional pain hypersensitivity in the adulthood.

Age-related Changes in Trigeminal Ganglion Macrophages Enhance Orofacial Ectopic Pain After Inferior Alveolar Nerve Injury.

BACKGROUND/AIM: The ectopic pain associated with inferior alveolar nerve (IAN) injury has been reported to involve macrophage expression in the trigeminal ganglion (TG). However, the effect of age-related changes on this abnormal pain conditions are still unknown. This study sought to clarify the involvement of age-related changes in macrophage expression and phenotypic conversion in the TG and how these changes enhance ectopic mechanical allodynia after IAN transection (IANX). MATERIALS AND METHODS: We used senescence-accelerated mouse (SAM)-prone 8 (SAMP8) and SAM-resistance 1 (SAMR1) mice, which are commonly used to study ageing-related changes. Mechanical stimulation was applied to the whisker pad skin under light anaesthesia; the mechanical head withdrawal threshold (MHWT) was measured for 21 d post-IANX. We subsequently counted the numbers of Iba1 (macrophage marker)-immunoreactive (IR) cells, Iba1/CD11c (M1-like inflammatory macrophage marker)-co-IR cells, and Iba1/CD206 (M2-like anti-inflammatory macrophage marker)-co-IR cells in the TG innervating the whisker pad skin. After continuous intra-TG administration of liposomal clodronate Clophosome®-A (LCCA) to IANX-treated SAMP8-mice, the MHWT values of the whisker pad skin were examined. RESULTS: Five days post-IANX, the MHWT had significantly decreased in SAMP8 mice compared to SAMR1-mice. Iba1-IR and Iba1/CD11c-co-IR cell counts were significantly increased in SAMP8 mice compared to SAMR1 mice 5 d post-IANX. LCCA administration significantly restored MHWT compared to control-LCCA administration. CONCLUSION: Ectopic mechanical allodynia of whisker pad skin after IANX is exacerbated by ageing, which involves increases in M1-like inflammatory macrophages in the TG.

Pannexin 1 role in the trigeminal ganglion in infraorbital nerve injury-induced mechanical allodynia.

OBJECTIVES: The detailed pathological mechanism of orofacial neuropathic pain remains unknown. We aimed to examine the pannexin 1 (Panx1) signaling in the trigeminal ganglion (TG) involvement in infraorbital nerve injury (IONI)-induced orofacial neuropathic pain. MATERIALS AND METHODS: Mechanical head-withdrawal threshold (MHWT) was measured in IONI-treated rats receiving intra-TG Panx1 inhibitor or metabotropic glutamate receptor 5 (mGluR5) antagonist administration and MHWTs in naive rats receiving intra-TG mGluR5 agonist administration post-IONI. Glutamate and Panx1 in the TG were measured post-IONI. Panx1, mGluR5, and glutamine synthetase expression in TG were immunohistochemically identified, and changes in the number of mGluR5-P2X3 -expressed TG neurons were examined. RESULTS: MHWT was significantly decreased post-IONI, and this decrease was reversed by Panx1 inhibition or mGluR5 antagonism. mGluR5 agonism induced a decrease in the MHWT. IONI increased extracellular glutamate in TG. Panx1 was expressed in satellite glial cells and TG neurons, and intra-TG mGluR5 antagonism decreased the number of mGluR5 and P2X3 positive TG neurons post-IONI. CONCLUSIONS: IONI facilitates glutamate release via Panx1 that activates mGluR5 which was expressed in the nociceptive TG neurons innervating the orofacial region. In turn, P2X3 receptor-expressed TG neurons are enhanced via mGluR5 signaling, resulting in orofacial neuropathic pain.

Astrocytic and microglial interleukin-1ß mediates complement C1q-triggered orofacial mechanical allodynia.

Glial cells, such as microglia and astrocytes, in the trigeminal spinal subnucleus caudalis (Vc) are activated after trigeminal nerve injury and interact with Vc neurons to contribute to orofacial neuropathic pain. Complement C1q released from microglia has been reported to activate astrocytes and causes orofacial mechanical allodynia. However, how C1q-induced phenotypic alterations in Vc astrocytes are involved in orofacial pain remains to be elucidated. Intracisternal administration of C1q caused mechanical allodynia in the whisker pad skin and concurrent significant upregulation of glial fibrillary acidic protein and ionized calcium-binding adapter molecule 1 in the Vc. Immunohistochemical analyses clarified that C1q induces a significant increase in the cytokine interleukin (IL)-1ß, predominantly in Vc astrocytes and partially in Vc microglia. The number of c-Fos-positive neurons in the Vc increased significantly in response to C1q. IL-1 receptor antagonist (IL-1Ra) was used to analyze the involvement of IL-1ß in C1q-induced mechanical allodynia. Intracisternal administration of IL-1Ra ameliorated C1q-induced orofacial mechanical allodynia. The present findings suggest that IL-1ß released from activated astrocytes and microglia in the Vc mediates C1q-induced orofacial pain.

Decreased PPARgamma in the trigeminal spinal subnucleus caudalis due to neonatal injury contributes to incision-induced mechanical allodynia in female rats.

Whisker pad skin incision in infancy causes the prolongation of mechanical allodynia after re-incision in adulthood. A recent study also proposed the importance of sex differences in pain signaling in the spinal cord. However, the sex difference in re-incision-induced mechanical allodynia in the orofacial region is not fully understood. In the rats that experienced neonatal injury in the whisker pad skin, the mechanical allodynia in the whisker pad was significantly prolonged after re-incision in adulthood compared to sham injury in infancy. No significant sex differences were observed in the duration of mechanical allodynia. The duration of mechanical allodynia in male rats was shortened by intracisternal administration of minocycline. However, minocycline had no effects on the duration of mechanical allodynia in female rats. In contrast, intracisternal administration of pioglitazone markedly suppressed mechanical allodynia in female rats after re-incision. Following re-incision, the number of peroxisome proliferator-activated receptor gamma (PPARgamma)-positive cells were reduced in the trigeminal spinal subnucleus caudalis (Vc) in female rats that experienced neonatal injury. Immunohistochemical analyses revealed that PPARgamma was predominantly expressed in Vc neurons. Pioglitazone increased the number of PPARgamma-positive Vc neurons in female rats whose whisker pad skin was incised in both infancy and adulthood stages. Pioglitazone also upregulated heme oxygenase 1 and downregulated NR1 subunit in the Vc in female rats after re-incision. Together, PPARgamma signaling in Vc neurons is a female-specific pathway for whisker pad skin incision-induced mechanical allodynia.

Effect of cisplatin on oral ulcer-induced nociception in rats.

OBJECTIVE: The aim of this study is to investigate effects of cisplatin preadministration on oral ulcerative mucositis-induced nociception by using an experimental model of rats. DESIGN: After two rounds of cisplatin administration, oral ulcers developed with topical acetic acid treatment in rats. Spontaneous mouth rubbing behavior was observed as spontaneous nociceptive behavior in a plastic cage. Head-withdrawal behavior was observed as mechanical allodynia by using von Frey test in the oral mucosa of conscious rats. Bacterial invasion and inflammatory cell infiltration into oral ulcerative region and systemic leukocyte phagocytic activity were assessed. RESULTS: Following cisplatin preadministration, oral ulcerative mucositis-induced spontaneous nociceptive behavior was not observed in the model. The preadministration enhanced leukocyte phagocytic activity, leading to reduce bacterial invasion and inflammatory cell infiltration in the oral ulcerative region. In contrast, oral ulcerative mucositis-induced mechanical allodynia was induced. The exaggerated mechanical allodynia in the oral ulcerative region was largely inhibited by topical treatment with the antioxidative drug, ɑ-lipoic acid, or the blocker of N-formyl peptide receptor 1, N-t-butoxycarbonyl-methionyl-leucyl-phenylalanine. CONCLUSIONS: These results suggest that cisplatin preadministration suppresses spontaneous nociception in oral ulcerative region, due to antiinflammatory effects by enhancement of leukocyte phagocytic activity, but exaggerates mechanical allodynia due to oxidative stress with N-formyl peptide receptor 1 activation. The suppression of spontaneous nociception is one of the advantages of cisplatin treatment for head and neck cancer patients although the exaggerated allodynia is a serious symptom.

Plastic changes in nociceptive pathways contributing to persistent orofacial pain.

BACKGROUND: Pain is a warning signal for the body defense mechanisms and is a critical sensation for supporting life. However, there are still many unclear points about the pathophysiological mechanism of orofacial pain. This situation makes it difficult for many clinicians to treat orofacial pain hypersensitivity. HIGHLIGHT: Noxious information on the orofacial region received by trigeminal ganglion neurons is recognized as "orofacial pain" by being transmitted to the somatosensory cortex and limbic system via the spinal trigeminal nucleus and the thalamic sensory nuclei. Orofacial inflammation or trigeminal nerve injury causes neuropathic changes in various nociceptive signaling pathways, resulting in persistent orofacial pain. It is also considered that persistent orofacial pain is triggered by plastic changes in nociceptive signaling pathways involving various cells such as satellite glial cells, astrocytes, microglia, and macrophages, as well as nociceptive neurons. CONCLUSION: Recent studies have shown that hyperexcitability of nociceptive neurons in the nociceptive signaling pathways of the orofacial region caused by a variety of factors causes persistent orofacial pain. This review outlines the pathophysiology of orofacial pain along with the results of our study.

Age-Related Differences in Transient Receptor Potential Vanilloid 1 and 2 Expression Patterns in the Trigeminal Ganglion Neurons Contribute to Changes in the Palatal Mucosal Heat Pain Sensitivity.

Aging affects various sensory functions of the body. However, the effect on the oral mucosal nociception has remain unclear, so this elucidation is very important. Therefore, this study aimed to evaluate the effect of age-related changes in transient receptor potential vanilloid 1 (TRPV1) and TRPV2 expression in the trigeminal ganglion (TG) neurons on intraoral mucosal heat sensitivity in the senescence-accelerated mouse prone 8 (SAMP8) model. We used 23-week-old (aged) and 7-week-old (young) SAMP8 mice. Heat stimulation was applied to the palatal mucosa under light anesthesia; moreover, the heat head withdrawal threshold (HHWT) was measured. We counted the number of TRPV1-immunoreactive (IR) and TRPV2-IR TG neurons innervating the palatal mucosa. Additionally, we investigated changes in HHWT when TRPV1 or TRPV2 antagonists (SB366791 or Tranilast) were administered to the palatal mucosa. Aged SAMP8 mice showed a higher HHWT than young SAMP8 mice. Compared with the aged SAMP8 mice, young SAMP8 mice showed a larger number of TRPV1-IR small-diameter neurons and a smaller number of TRPV2-IR medium-sized neurons innervating the palatal mucosa. SB366791 administration increased the HHWT in young, but not aged SAMP8 mice. Contrastingly, Tranilast administration increased the HHWT in aged, but not young SAMP8 mice. These results suggest that the modulation of heat pain sensitivity in the oral mucosa due to aging is dependent on changes in the TRPV1 and TRPV2 expression patterns in the TG neurons innervating the palatal mucosa.

Butyric acid modulates periodontal nociception in Porphyromonas gingivalis-induced periodontitis.

PURPOSE: Periodontitis progresses with chronic inflammation, without periodontal pain. However, the underlying mechanisms are not well known. Here, the involvement of butyric acid (BA) in periodontal pain sensitivity in Porphyromonas gingivalis (P. gingivalis)-induced periodontitis was examined. METHODS: P. gingivalis was inoculated into the ligature which was tied around the molar (P. gingivalis-L) and the gingival mechanical head withdrawal threshold (MHWT) was measured. Following P. gingivalis-L, the expressions of orphan G protein-coupled receptor 41 (GPR41) in trigeminal ganglion (TG) neurons were examined. The amount of gingival BA was analyzed following the P. gingivalis-L and the changes in the MHWT in complete Freund's adjuvant (CFA)-injected gingival tissue by gingival BA were examined. The changes in the MHWT following P. gingivalis-L by gingival GPR41 antagonist (HA) were examined. RESULTS: No change in the MHWT was observed, GPR41-immunoreactive TG neurons were increased following P. gingivalis-L. The gingival BA amount increased following P. gingivalis-L, and the gingival BA suppressed the decrease in MHWT following CFA. HA decreased MHWT following P. gingivalis-L. CONCLUSION: Gingival BA modulates periodontal mechanical nociception via GPR41 signaling in P. gingivalis-L-induced periodontitis.

IL-33 induces orofacial neuropathic pain through Fyn-dependent phosphorylation of GluN2B in the trigeminal spinal subnucleus caudalis.

Orofacial neuropathic pain can cause considerable disruptions in patients' daily lives, especially because of a lack of effective medications as its underlying causative mechanisms are not fully understood. Here, we found neuron-specific expression of the interleukin (IL)-33 receptor in the trigeminal spinal subnucleus caudalis (Vc), distinct from the spinal dorsal horn. Reduction in head withdrawal threshold in response to von Frey filament stimulation of the whisker pad skin was inversely correlated with the upregulation of IL-33 in the Vc after infraorbital nerve injury (IONI). Neutralization of IL-33 in the Vc alleviated mechanical allodynia in the whisker pad skin after IONI; conversely, intracisternal administration of IL-33 elicited mechanical allodynia in the whisker pad skin, which was relieved by GluN2B antagonism. Moreover, IL-33 triggered the potentiation of GluN2B-containing N-methyl-D-aspartate receptor-mediated synaptic currents and phosphorylation of synaptosomal GluN2B in the Vc, whereas IONI-induced GluN2B phosphorylation was inhibited by neutralization of IL-33 in the Vc. IL-33-induced GluN2B phosphorylation was mediated by phosphorylation of Fyn kinase, and inhibition of the Fyn kinase pathway prevented the development of IL-33-induced mechanical allodynia. Our findings provide insights into a new mechanism by which IL-33 directly regulates synaptic transmission and suggest that IL-33 signaling could be a candidate target for therapeutic interventions for orofacial neuropathic pain.

Periodontal acidification contributes to tooth pain hypersensitivity during orthodontic tooth movement.

Tooth movements associated with orthodontic treatment often cause tooth pain. However, the detailed mechanism remains unclear. Here, we examined the involvement of periodontal acidification caused by tooth movement in mechanical tooth pain hypersensitivity. Elastics were inserted between the first and second molars to move the teeth in Sprague-Dawley rats. Mechanical head-withdrawal reflex threshold to first molar stimulation and the pH of the gingival sulcus around the tooth were measured. The expression of acid-sensing ion channel 3 (ASIC3) in trigeminal ganglion neurons and phosphorylation of ASIC3 in the periodontal tissue were analyzed. The mechanical head-withdrawal reflex threshold to first molar stimulation and pH in the gingival sulcus decreased on day 1 after the elastic insertion. These decreases recovered to the sham level by buffering periodontal acidification. Periodontal inhibition of ASIC3 channel activity reversed the decreased mechanical head-withdrawal reflex threshold to first molar stimulation. On day 1 after elastic insertion, the tooth movement did not change the number of ASIC3 immunoreactive trigeminal ganglion neurons innervating the periodontal tissue but increased phosphorylated-ASIC3 levels in the periodontal tissue. Periodontal acidification induced by tooth movement causes phosphorylation of ASIC3, resulting in mechanical pain hypersensitivity in mechanically forced tooth.

Cisplatin induces TRPA1-mediated mechanical allodynia in the oral mucosa.

OBJECTIVE: Cisplatin, a platinum-based anticancer drug, produces reactive oxygen species (ROS) in many cell types and induces mechanical allodynia in the hands and/or feet (chemotherapy-induced painful neuropathy: CIPN). In this study, we examined the possibility of inducing neuropathy in the oral region using oral keratinocytes and rats. METHODS: Human oral keratinocytes (HOKs) were used to evaluate ROS generation after cisplatin application by a ROS-reactive fluorescent assay. In rats, after cisplatin administrations (two times), the trigeminal ganglion (TG) was investigated by electron microscopy and quantitative RT-PCR. Using our proprietary assay system, oral pain-related behaviors were observed in cisplatin-treated rats. RESULTS: In rats, cisplatin administration reduced food intake and body weight. In electron microscopic analysis, glycogen granules in the TG were depleted following administration, although organelles were intact. In HOK cells, cisplatin significantly increased ROS generation with cell death, similar to glycolysis inhibitors. Cisplatin administration did not show any effects on Trpa1 mRNA levels in the TG. However, the same procedure induced hypersensitivity to mechanical stimulation and the TRPA1 agonist allyl isothiocyanate in the oral mucosa. Mechanical hypersensitivity was inhibited by the antioxidative drug α-lipoic acid and the TRPA1 antagonist HC-030031, similar to that of the hind paw. CONCLUSION: The present findings suggest that cisplatin induces TRPA1-mediated CIPN due to ROS generation in the oral region. This study will provide a better understanding of persistent oral pain in cancer patients.