Analgesic Effect of Tranilast in an Animal Model of Neuropathic Pain and Its Role in the Regulation of Tetrahydrobiopterin Synthesis.

Trigeminal neuralgia is unilateral, lancinating, episodic pain that can be provoked by routine activities. Anticonvulsants, such as carbamazepine, are the drugs of choice; however, these possess side-effects. Microvascular decompression is the most effective surgical technique with a higher success rate, although occasionally causes adverse effects. The potential treatment for this type of pain remains unmet. Increased tetrahydrobiopterin (BH4) levels have been reported in association with axonal injury. This study aimed to evaluate the effect of tranilast on relieving neuropathic pain in animal models and analyze the changes in BH4 synthesis. Neuropathic pain was induced via infraorbital nerve constriction. Tranilast, carbamazepine, or saline was injected intraperitoneally to assess the rat's post-intervention pain response. In the von Frey's test, the tranilast and carbamazepine groups showed significant changes in the head withdrawal threshold in the ipsilateral whisker pad area. The motor coordination test showed no changes in the tranilast group, whereas the carbamazepine group showed decreased performance, indicating impaired motor coordination. Trigeminal ganglion tissues were used for the PCR array analysis of genes that regulate the BH4 pathway. Downregulation of the sepiapterin reductase (Spr) and aldoketo reductase (Akr) genes after tranilast injection was observed compared to the pain model. These findings suggest that tranilast effectively treats neuropathic pain.

Role of CGRP in Neuroimmune Interaction via NF-κB Signaling Genes in Glial Cells of Trigeminal Ganglia.

Activation of the trigeminal system causes the release of various neuropeptides, cytokines, and other immune mediators. Calcitonin gene-related peptide (CGRP), which is a potent algogenic mediator, is expressed in the peripheral sensory neurons of trigeminal ganglion (TG). It affects the inflammatory responses and pain sensitivity by modulating the activity of glial cells. The primary aim of this study was to use array analysis to investigate the effect of CGRP on the glial cells of TG in regulating nuclear factor kappa B (NF-κB) signaling genes and to further check if CGRP in the TG can affect neuron-glia activation in the spinal trigeminal nucleus caudalis. The glial cells of TG were stimulated with CGRP or Minocycline (Min) + CGRP. The effect on various genes involved in NF-κB signaling pathway was analyzed compared to no treatment control condition using a PCR array analysis. CGRP, Min + CGRP or saline was directly injected inside the TG and the effect on gene expression of Egr1, Myd88 and Akt1 and protein expression of cleaved Caspase3 (cleav Casp3) in the TG, and c-Fos and glial fibrillary acidic protein (GFAP) in the spinal section containing trigeminal nucleus caudalis was analyzed. Results showed that CGRP stimulation resulted in the modulation of several genes involved in the interleukin 1 signaling pathway and some genes of the tumor necrosis factor pathway. Minocycline pre-treatment resulted in the modulation of several genes in the glial cells, including anti-inflammatory genes, and neuronal activation markers. A mild increase in cleav Casp3 expression in TG and c-Fos and GFAP in the spinal trigeminal nucleus of CGRP injected animals was observed. These data provide evidence that glial cells can participate in neuroimmune interaction due to CGRP in the TG via NF-κB signaling pathway.

Fibronectin-induced ductal formation in salivary gland self-organization model.

BACKGROUND: Recent advances in tissue regeneration approaches including 3D organoids, were based on various 3D organogenesis models. However, 3D models are generally technique-sensitive and time-consuming. Thus, we utilized an existing model of submandibular salivary gland (SMG) to modify a simple and highly reproducible in vitro 3D culture model of primary SMG cells self-organization into a well-developed cell spheroid inside Matrigel substrate. We used this model to observe the collective multicellular behavior during spheroid formation. Further, we applied various quantitative approaches including real-time live imaging and immune histochemical image analysis to dissect the cellular dynamics during tissue patterning. RESULTS: On a time-scale of hours, we observed marked size and shape transformations in the developed 3D spheroid which resulted in a spatially-controlled growth differential from the canter to the periphery of the formed aggregates. Moreover, we investigated the effect of fibronectin (FN) on SMG cells self-organization using our simplified culture model. Interestingly, we discovered a novel role of FN in inducing duct-like elongation during initial stages of SMG bud formation. CONCLUSION: This in vitro model provides an excellent tool for analyzing the intercellular dynamics during early SMG tissue development as well as revealing a novel role of FN in SMG ductal expansion.

CGRP Induces Differential Regulation of Cytokines from Satellite Glial Cells in Trigeminal Ganglia and Orofacial Nociception.

Neuron-glia interactions contribute to pain initiation and sustainment. Intra-ganglionic (IG) secretion of calcitonin gene-related peptide (CGRP) in the trigeminal ganglion (TG) modulates pain transmission through neuron-glia signaling, contributing to various orofacial pain conditions. The present study aimed to investigate the role of satellite glial cells (SGC) in TG in causing cytokine-related orofacial nociception in response to IG administration of CGRP. For that purpose, CGRP alone (10 µL of 10-5 M), Minocycline (5 µL containing 10 µg) followed by CGRP with one hour gap (Min + CGRP) were administered directly inside the TG in independent experiments. Rats were evaluated for thermal hyperalgesia at 6 and 24 h post-injection using an operant orofacial pain assessment device (OPAD) at three temperatures (37, 45 and 10 °C). Quantitative real-time PCR was performed to evaluate the mRNA expression of IL-1ß, IL-6, TNF-α, IL-1 receptor antagonist (IL-1RA), sodium channel 1.7 (NaV 1.7, for assessment of neuronal activation) and glial fibrillary acidic protein (GFAP, a marker of glial activation). The cytokines released in culture media from purified glial cells were evaluated using antibody cytokine array. IG CGRP caused heat hyperalgesia between 6⁻24 h (paired-t test, p < 0.05). Between 1 to 6 h the mRNA and protein expressions of GFAP was increased in parallel with an increase in the mRNA expression of pro-inflammatory cytokines IL-1ß and anti-inflammatory cytokine IL-1RA and NaV1.7 (one-way ANOVA followed by Dunnett's post hoc test, p < 0.05). To investigate whether glial inhibition is useful to prevent nociception symptoms, Minocycline (glial inhibitor) was administered IG 1 h before CGRP injection. Minocycline reversed CGRP-induced thermal nociception, glial activity, and down-regulated IL-1ß and IL-6 cytokines significantly at 6 h (t-test, p < 0.05). Purified glial cells in culture showed an increase in release of 20 cytokines after stimulation with CGRP. Our findings demonstrate that SGCs in the sensory ganglia contribute to the occurrence of pain via cytokine expression and that glial inhibition can effectively control the development of nociception.

Ligature induced peri-implantitis: tissue destruction and inflammatory progression in a murine model.

OBJECTIVES: The aim of this study was to investigate tissue destruction and inflammatory progression of ligature-induced peri-implantitis in mice and to establish an alternative murine model of peri-implantitis. MATERIAL AND METHODS: Sixty male C57BL/6NCrSlc mice (4-week-old) were used and the maxillary right first molars were extracted. Eight weeks after extraction, custom-made pure titanium machined screw type implants (0.8 × 1.5 mm) were placed, one implant per animal. Four weeks later, 5-0 silk ligatures were applied around implant necks to induce peri-implantitis. Animals were sacrificed at 0 (before ligature), 7, 14, 21 and 28 days after ligature. Half of the samples were analyzed radiologically and histologically to measure bone level change, osteoclast number, density, and distribution. The rest of the samples was used to determine the relative mRNA expression levels of IL-1 and TNF-α with RT-PCR analysis. RESULTS: Bone levels at all sites (buccal, palatal, mesial, distal) decreased 40-50% significantly 28 days after ligature (P < 0.01). Osteoclast number at all post-ligature time points increased significantly (P < 0.05). However, their density at day 28 decreased significantly compared to that of day 21 (P < 0.05). Accordingly, IL-1 and TNF-α mRNA expression increased significantly at the early time points but decreased significantly at day 28 after ligature (P < 0.05). CONCLUSIONS: Inflammatory response followed by significant peri-implant bone resorption suggested 28 days ligation is sufficient to successfully induce peri-implantitis in the current mice model. This model might open a new avenue to study the pathogenesis and mechanism of peri-implantitis.

Whole Tooth Regeneration as a Future Dental Treatment.

Dental problems caused by dental caries, periodontal disease and tooth injury compromise the oral and general health issues. Current advances for the development of regenerative therapy have been influenced by our understanding of embryonic development, stem cell biology, and tissue engineering technology. Tooth regenerative therapy for tooth tissue repair and whole tooth replacement is currently expected a novel therapeutic concept with the full recovery of tooth physiological functions. Dental stem cells and cell-activating cytokines are thought to be candidate approach for tooth tissue regeneration because they have the potential to differentiate into tooth tissues in vitro and in vivo. Whole tooth replacement therapy is considered to be an attractive concept for next generation regenerative therapy as a form of bioengineered organ replacement. For realization of whole tooth regeneration, we have developed a novel three-dimensional cell manipulation method designated the "organ germ method". This method involves compartmentalisation of epithelial and mesenchymal cells at a high cell density to mimic multicellular assembly conditions and epithelial-mesenchymal interactions in organogenesis. The bioengineered tooth germ generates a structurally correct tooth in vitro, and erupted successfully with correct tooth structure when transplanted into the oral cavity. We have ectopically generated a bioengineered tooth unit composed of a mature tooth, periodontal ligament and alveolar bone, and that tooth unit was engrafted into an adult jawbone through bone integration. Bioengineered teeth were also able to perform physiological tooth functions such as mastication, periodontal ligament function and response to noxious stimuli. In this review, we describe recent findings and technologies underpinning whole tooth regenerative therapy.

Functional tooth regenerative therapy: tooth tissue regeneration and whole-tooth replacement.

Oral and general health is compromised by irreversible dental problems, including dental caries, periodontal disease and tooth injury. Regenerative therapy for tooth tissue repair and whole-tooth replacement is currently considered a novel therapeutic concept with the potential for the full recovery of tooth function. Several types of stem cells and cell-activating cytokines have been identified in oral tissues. These cells are thought to be candidate cell sources for tooth tissue regenerative therapies because they have the ability to differentiate into tooth tissues in vitro and in vivo. Whole-tooth replacement therapy is regarded as an important model for the development of an organ regenerative concept. A novel three-dimensional cell-manipulation method, designated the organ germ method, has been developed to recapitulate organogenesis. This method involves compartmentalisation of epithelial and mesenchymal cells at a high cell density to mimic multicellular assembly conditions and epithelial-mesenchymal interactions. A bioengineered tooth germ can generate a structurally correct tooth in vitro and erupt successfully with the correct tooth structure when transplanted into the oral cavity. We have ectopically generated a bioengineered tooth unit composed of a mature tooth, periodontal ligament and alveolar bone, and that tooth unit was successfully engrafted into an adult jawbone through bone integration. Such bioengineered teeth were able to perform normal physiological tooth functions, such as developing a masticatory potential in response to mechanical stress and a perceptive potential for noxious stimuli. In this review, we describe recent findings and technologies underpinning tooth regenerative therapy.

Identification of microRNA Signatures in Peripheral Blood of Young Women as Potential Biomarkers for Metal Allergy.

MicroRNA (miRNA) is a short (19-24 nucleotide) endogenous non-protein RNA that exists in the body and controls the translation process from genes to proteins. It has become useful as a diagnostic tool and a potential treatment target in cancer research. To explore the function of miRNA in contact dermatitis, female participants with a positive metal allergy diagnosis (n = 3) were enrolled along with additional female participants with no medical history of metal allergy (n = 3). A patch test was performed on each participant. Peripheral blood was collected from all the participants before the patch test and at days 3 and 7 after starting the patch test. After total RNA was obtained from peripheral blood leukocytes and cDNA was generated, microarray analysis was performed to analyze the large-scale circulating miRNA profile. Real-time polymerase chain reaction (RT-PCR) was then used to clarify the overall target miRNA expression. Downregulation of hsa-let-7d-5p, hsa-miR-24-3p, hsa-miR-23b-3p, hsa-miR-26b-5p, and hsa-miR-150-5p was found on day 7. Certain miRNAs were confirmed using RT-PCR. These peripheral blood miRNAs could be diagnostic biomarkers for metal allergies.

ADAMTSL6ß protein rescues fibrillin-1 microfibril disorder in a Marfan syndrome mouse model through the promotion of fibrillin-1 assembly.

Marfan syndrome (MFS) is a systemic disorder of the connective tissues caused by insufficient fibrillin-1 microfibril formation and can cause cardiac complications, emphysema, ocular lens dislocation, and severe periodontal disease. ADAMTSL6ß (A disintegrin-like metalloprotease domain with thrombospondin type I motifs-like 6ß) is a microfibril-associated extracellular matrix protein expressed in various connective tissues that has been implicated in fibrillin-1 microfibril assembly. We here report that ADAMTSL6ß plays an essential role in the development and regeneration of connective tissues. ADAMTSL6ß expression rescues microfibril disorder after periodontal ligament injury in an MFS mouse model through the promotion of fibrillin-1 microfibril assembly. In addition, improved fibrillin-1 assembly in MFS mice following the administration of ADAMTSL6ß attenuates the overactivation of TGF-ß signals associated with the increased release of active TGF-ß from disrupted fibrillin-1 microfibrils within periodontal ligaments. Our current data thus demonstrate the essential contribution of ADAMTSL6ß to fibrillin-1 microfibril formation. These findings also suggest a new therapeutic strategy for the treatment of MFS through ADAMTSL6ß-mediated fibrillin-1 microfibril assembly.

Risk Factors for Tooth Loss in Patients with ≥25 Remaining Teeth Undergoing Mid-Long-Term Maintenance: A Retrospective Study.

Tooth loss represents a diffused pathologic condition affecting the worldwide population. Risk factors have been identified in both general features (smoking, diabetes, economic status) and local tooth-related factors (caries, periodontitis). In this retrospective study, we examined the data of 366 patients with a large number of remaining teeth (≥25) undergoing maintenance therapy in order to identify specific risk factors for tooth loss. The number of remaining teeth, number of non-vital teeth, and number of occlusal units were investigated for their correlation with tooth loss. The mean follow-up of patients was 9.2 years (range 5 to 14). Statistically significant risk factors for tooth loss were identified as number of remaining teeth at baseline (p = 0.05), number of occlusal units (p = 0.03), and number of non-vital teeth in posterior regions (p < 0.001). Multiple logistic regression showed that the number of occlusal units and number of non-vital teeth in the posterior regions were significantly associated with a greater risk of tooth loss (odds ratio 1.88 and 3.17, respectively). These results confirm that not only the number of remaining teeth, but also their vital or non-vital status and the distribution between the anterior and posterior regions influence the long-term survival.

Mechanical homeostasis of liver sinusoid is involved in the initiation and termination of liver regeneration.

Organogenesis and regeneration are fundamental for developmental progress and are associated with morphogenesis, size control and functional properties for whole-body homeostasis. The liver plays an essential role in maintaining homeostasis of the entire body through various functions, including metabolic functions, detoxification, and production of bile, via the three-dimensional spatial arrangement of hepatic lobules and has high regenerative capacity. The regeneration occurs as hypertrophy, which strictly controls the size and lobule structure. In this study, we established a three-dimensional sinusoidal network analysis method and determined valuable parameters after partial hepatectomy by comparison to the static phase of the liver. We found that mechanical homeostasis, which is crucial for organ morphogenesis and functions in various phenomena, plays essential roles in liver regeneration for both initiation and termination of liver regeneration, which is regulated by cytokine networks. Mechanical homeostasis plays critical roles in the initiation and termination of organogenesis, tissue repair and organ regeneration in coordination with cytokine networks.

Peripherally Administered Botulinum Toxin Type A Localizes Bilaterally in Trigeminal Ganglia of Animal Model.

Peripheral nerve injury leads to sensory ganglion hyperexcitation, which increases neurotransmitter release and neuropathic pain. Botulinum toxin type A (BoNT/A) regulates pain transmission by reducing neurotransmitter release, thereby attenuating neuropathic pain. Despite multiple studies on the use of BoNT/A for managing neuropathic pain in the orofacial region, its exact mechanism of transport remains unclear. In this study, we investigated the effects of BoNT/A in managing neuropathic pain in two different animal models and its transport mechanism in the trigeminal nerve. Intraperitoneal administration of cisplatin induced bilateral neuropathic pain in the orofacial region, reducing the head withdrawal threshold to mechanical stimulation. Unilateral infraorbital nerve constriction (IONC) also reduced the ipsilateral head withdrawal threshold to mechanical stimulation. Unilateral peripheral administration of BoNT/A to the rat whisker pad attenuated cisplatin-induced pain behavior bilaterally. Furthermore, contralateral peripheral administration of BoNT/A attenuated neuropathy-induced behavior caused by IONC. We also noted the presence of BoNT/A in the blood using the mouse bioassay. In addition, the Alexa Fluor-488-labeled C-terminal half of the heavy chain of BoNT/A (BoNT/A-Hc) was localized in the neurons of the bilateral trigeminal ganglia following its unilateral administration. These findings suggest that axonal and hematogenous transport are involved in the therapeutic effects of peripherally administered BoNT/A in the orofacial region.

Dentin-Pulp Complex Tissue Regeneration via Three-Dimensional Cell Sheet Layering.

The dentin-pulp complex is a unique structure in teeth that contains both hard and soft tissues. Generally, deep caries and trauma cause damage to the dentin-pulp complex, and if left untreated, this damage will progress to irreversible pulpitis. The aim of this study was to fabricate a layered cell sheet composed of rat dental pulp (DP) cells and odontogenic differentiation of pulp (OD) cells and to investigate the ability to regenerate the dentin-pulp complex in a scaffold tooth. We fabricated two single cell sheets composed of DP cells (DP cell sheet) or OD cells (OD cell sheet) and a layered cell sheet made by layering both cells. The characteristics of the fabricated cell sheets were analyzed using light microscopy, scanning electron microscope (SEM), hematoxylin-eosin (HE) staining, and immunohistochemistry (IHC). Furthermore, the cell sheets were transplanted into the subrenal capsule of immunocompromised mice for 8 weeks. After this, the regenerative capacity to form dentin-like tissue was evaluated using micro-computed tomography (micro-CT), HE staining, and IHC. The findings of SEM and IHC confirmed that layered cell sheets fabricated by stacking OD cells and DP cells maintained their cytological characteristics. Micro-CT of layered cell sheet transplants revealed a mineralized capping of the access cavity in the crown area, similar to that of natural dentin. In contrast, the OD cell sheet group demonstrated the formation of irregular fragments of mineralized tissue in the pulp cavity, and the DP cell sheet did not develop any hard tissue. Moreover, bone volume/tissue volume (BV/TV) showed a significant increase in hard tissue formation in the layered cell sheet group compared with that in the single cell sheet group (p < 0.05). HE staining also showed a combination of soft and hard tissue formation in the layered cell sheet group. Furthermore, IHC confirmed that the dentin-like tissue generated from the layered cell sheet expressed characteristic markers of dentin but not bone equivalent to that of a natural tooth. In conclusion, this study demonstrates the feasibility of regenerating dentin-pulp complex using a bioengineered tissue designed to simulate the anatomical structure. Impact statement The dentin-pulp complex can be destroyed by deep caries and trauma, which may cause pulpitis and progress to irreversible pulpitis, apical periodontitis, and even tooth loss. Current treatments cannot maintain pulp health, and teeth can become brittle. We developed a three-dimensional (3D) layered cell sheet using dental pulp cells and odontogenic differentiation of pulp cells for dentin-pulp complex regeneration. Our layered cell sheet enables the regeneration of an organized 3D dentin-pulp-like structure comparable with that of natural teeth. This layered cell sheet technology may contribute to dentin-pulp complex regeneration and provide a novel method for complex tissue engineering.

Risk Factors for Tooth Loss in Patients Undergoing Mid-Long-Term Maintenance: A Retrospective Study.

In this retrospective study, we identified risk factors for tooth loss in patients undergoing mid-long-term maintenance therapy. We surveyed 674 maintenance patients for ≥5 years after active treatment who visited a dental clinic between January 2015 and December 2016. Of these, 265 were men (mean age 54.6 ± 8.0 years old) and 409 were women (mean age 54.0 ± 7.9 years old). Study variables included patient compliance, sex, number of teeth lost, cause of tooth loss (dental caries, periodontal disease, root fracture, others, vital or non-vital teeth), age at start of maintenance, number of remaining teeth at start of maintenance, smoking, use of salivary secretion inhibitors, presence of diabetes mellitus, condition of periodontal bone loss, and use of a removable denture. Most lost teeth were non-vital teeth (91.7% of all cases) and the most common cause of tooth loss was tooth fracture (62.1% of all cases). A statistically significant risk factors for tooth loss was number of remaining teeth at the start of maintenance (p = 0.003).

The role of chemical transmitters in neuron-glia interaction and pain in sensory ganglion.

Neuropathic pain (NP) develops because of damage to the peripheral or central nervous system. It results in the hyperalgesia and allodynia. In the recent years, various researchers have studied the involvement of neuro-immune system in causing persistence of pain. The absence of synaptic contacts in the sensory ganglion makes them distinctive in terms of pain related signalling. In sensory ganglia, the neurotransmitters or the other modulators such as inflammatory substances produced by the ganglion cells, because of an injury, are responsible for the cross-excitation between neurons and neuron-glial interaction, thus affecting chemical transmission. This chemical transmission is considered mainly responsible for the chronicity and the persistent nature of neuropathic pain. This review examines the pain signalling due to neurotransmitter or cytokine release within the sensory ganglia. The specific areas focused on include: 1) the role of neurotransmitters released from the somata of sensory neurons in pain, 2) neuron-glia interaction and 3) role of cytokines in neuromodulation and pain.

Immediate Effect of Masticatory Muscle Activity with Transcutaneous Electrical Nerve Stimulation in Muscle Pain of Temporomandibular Disorders Patients.

Transcutaneous electrical nerve stimulation (TENS) is a non-invasive treatment modality for acute and chronic pain. However, little information for muscle activity is available on the immediate effects of TENS in masticatory muscle pain related to temporomandibular disorders (TMDs). The present study aimed to evaluate the immediate effects of TENS treatment on TMD-related muscle pain. Thirty-six patients with TMD-related muscle pain and 39 healthy subjects served as TMD and control groups, respectively. For objective evaluations, maximum mouth opening, and maximum bite force were measured before and after TENS. The pain intensity was assessed according to a 100-mm visual analog scale (VAS). TENS was applied to painful muscles for 20 min with frequencies of 100-200 Hz. The treatment outcome was evaluated using Global Rating of Change (GRC) scales. In the TMD group, VAS values significantly decreased after TENS. Although there was significant increase in the maximum mouth opening after TENS for only TMD group, the maximum bite force of both groups was significantly greater after TENS. According to GRC scales, one patient with TMD-related muscle pain expressed negative feelings after TENS. Conclusively, TENS treatment might quickly relieve pain in masticatory muscles and improve masticatory functions in patients with TMD-related muscle pain.

Three-dimensional periodontal tissue regeneration using a bone-ligament complex cell sheet.

Periodontal tissue is a distinctive tissue structure composed three-dimensionally of cementum, periodontal ligament (PDL) and alveolar bone. Severe periodontal diseases cause fundamental problems for oral function and general health, and conventional dental treatments are insufficient for healing to healthy periodontal tissue. Cell sheet technology has been used in many tissue regenerations, including periodontal tissue, to transplant appropriate stem/progenitor cells for tissue regeneration of a target site as a uniform tissue. However, it is still difficult to construct a three-dimensional structure of complex tissue composed of multiple types of cells, and the transplantation of a single cell sheet cannot sufficiently regenerate a large-scale tissue injury. Here, we fabricated a three-dimensional complex cell sheet composed of a bone-ligament structure by layering PDL cells and osteoblast-like cells on a temperature responsive culture dish. Following ectopic and orthotopic transplantation, only the complex cell sheet group was demonstrated to anatomically regenerate the bone-ligament structure along with the functional connection of PDL-like fibers to the tooth root and alveolar bone. This study represents successful three-dimensional tissue regeneration of a large-scale tissue injury using a bioengineered tissue designed to simulate the anatomical structure.

IL-10 and CXCL2 in trigeminal ganglia in neuropathic pain.

Many trigeminal neuropathic pain patients suffer severe chronic pain. The neuropathic pain might be related with cross-excitation of the neighboring neurons and satellite glial cells (SGCs) in the sensory ganglia and increasing the pain signals from the peripheral tissue to the central nervous system. We induced trigeminal neuropathic pain by infraorbital nerve constriction injury (IONC) in Sprague-Dawley rats. We tested cytokine (CXCL2 and IL-10) levels in trigeminal ganglia (TGs) after trigeminal neuropathic pain induction, and the effect of direct injection of the anti-CXCL2 and recombinant IL-10 into TG. We found that IONC induced pain behavior. Additionally, IONC induced satellite glial cell activation in TG and cytokine levels of TGs were changed after IONC. CXCL2 levels increased on day 1 of neuropathic pain induction and decreased gradually, with IL-10 levels showing the opposite trend. Recombinant IL-10 or anti-CXCL2 injection into TG decreased pain behavior. Our results show that IL-10 or anti-CXCL2 are therapy options for neuropathic pain.

Insulin-like growth factor 1 modulates bioengineered tooth morphogenesis.

Regenerative therapy to replace missing teeth is a critical area of research. Functional bioengineered teeth have been produced by the organ germ method using mouse tooth germ cells. However, these bioengineered teeth are significantly smaller in size and exhibit an abnormal crown shape when compared with natural teeth. The proper sizes and shapes of teeth contribute to their normal function. Therefore, a method is needed to control the morphology of bioengineered teeth. Here, we investigated whether insulin-like growth factor 1 (IGF1) can regulate the sizes and shapes of bioengineered teeth, and assessed underlying mechanisms of such regulation. IGF1 treatment significantly increased the size of bioengineered tooth germs, while preserving normal tooth histology. IGF1-treated bioengineered teeth, which were developed from bioengineered tooth germs in subrenal capsules and jawbones, showed increased sizes and cusp numbers. IGF1 increased the number of fibroblast growth factor (Fgf4)-expressing enamel knots in bioengineered tooth germs and enhanced the proliferation and differentiation of dental epithelial and mesenchymal cells. This study is the first to reveal that IGF1 increases the sizes and cusp numbers of bioengineered teeth via the induction of enamel knot formation, as well as the proliferation and differentiation of dental epithelial and mesenchymal cells.

Promotion of hydroxyapatite-associated, stem cell-based bone regeneration by CCN2.

Multiple roles have been already recognized for CCN2 in cartilage development and regeneration. However, the effects of CCN2 on bone regeneration remain to be elucidated. In this study, the utility of CCN2 on bone regeneration was examined in vitro and in vivo in combination with hydroxyapatite (HAp) as a scaffold. Human bone marrow stromal cells (hBMSCs) were isolated from human iliac bone marrow aspirates of healthy donors and expanded, and the effects of CCN2 on their proliferation and migration were examined in vitro. The proliferation of hBMSCs on a plastic or HAp plate was significantly enhanced by CCN2. Moreover, the migration of hBMSCs also dramatically increased by CCN2. Interestingly, a C-terminal signal modular fragment of CCN2 (CT-module) also enhanced the cell proliferation and migration as efficiently as the full-length CCN2. Next, in order to estimate the effect of CCN2 on the migration and survival of hBMSCs and bone formation inside the HAp scaffold in vivo, two experiments were performed. First, the porous HAp carrier was cultured with hBMSCs for a week, and the cell-scaffold hybrid was transplanted with or without CCN2 subcutaneously into immunocompromised mice. CCN2 accelerated the hBMSC-like cell migration and survival inside the porous HAp within 4 weeks after transplantation. Second, the porous HAp carrier with or without CCN2 was directly implanted into bone defects within a rabbit mandible, and bone regeneration inside was evaluated. As a result, CCN2 efficiently induced the cell invasion and bone formation inside the porous HAp scaffold. These findings suggest that CCN2 and its CT-module fragment could be useful for regeneration and reconstruction of large-scale bone defects.