Search

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.

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.

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 (

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.

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 &lt, 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.

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&ndash, 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.

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-&kappa, B) 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-&kappa, B 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-&kappa, B signaling pathway.

In this retrospective study, we identified risk factors for tooth loss in patients undergoing mid&ndash, long-term maintenance therapy. We surveyed 674 maintenance patients for &ge, 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 &plusmn, 8.0 years old) and 409 were women (mean age 54.0 &plusmn, 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).

Bone marrow-derived mesenchymal stem cells (BMSCs) have considerable potential for self-renewal and multi-differentiation. Tumor necrosis factor-α (TNF-α) is an inflammatory cytokine and is involved in tissue regeneration during wound healing. It was already reported that cultured human dental pulp cells acquire stem cell properties following short-term stimulation by TNF-α. However, it has not been clarified if BMSCs acquire stem cell properties after TNF-α treatment. The purpose of this study was to investigate the effect of short-term stimulation with TNF-α on BMSCs. Rat BMSCs were cultured up to 60% confluence and then incubated with 1–100 ng/ml of recombinant rat TNF-α (rTNF-α) for a further 2 days. After reaching subconfluence, cells were passaged once to remove rTNF-α completely before subsequent assays. Cells in the control group were passaged without stimulation. Expression levels of Nanog and Oct4 stem cell markers were significantly increased in the rTNF-α 10 ng/ml stimulation group. rTNF-α stimulation did not affect cell proliferation compared with the control group. However, rTNF-α stimulation led to a delay in cell differentiation. This study suggests that short-term TNF-α stimulation of BMSCs significantly increased their stem cell phenotype, but delayed their osteogenic cell differentiation.

Background: Tooth loss is generally considered the final outcome of oral disease. This retrospective study was performed to identify risk factors for tooth loss in patients undergoing long-term maintenance therapy. Methods: We surveyed 1145 adult patients who underwent maintenance therapy for ≥5 years after they had undergone active treatment from January 2015 to December 2016 and established a baseline status. The study variables were patient compliance, sex, number of teeth lost, cause of tooth loss, age at start of maintenance, number of remaining teeth at start of maintenance, duration of maintenance, smoking status, use of salivary secretion inhibitors, and diabetes mellitus. Additionally, 57 patients who did not undergo maintenance therapy were surveyed to examine and compare the effects of maintenance therapy. Statistical analyses were performed to assess the correlation of each variable with tooth loss. Results: The average number of teeth lost under maintenance therapy was 0.07/year. Significantly fewer teeth were lost in the maintenance than non-maintenance group. Most of the teeth lost were non-vital teeth, and the most common cause of tooth loss was tooth fracture. Patient age, number of remaining teeth at start of maintenance, use of salivary secretion inhibitors, and diabetes mellitus were related to tooth loss. Conclusions: To the best of our knowledge, this is first large-scale study of tooth loss in patients undergoing long-term maintenance therapy within a general dental clinic. Our findings demonstrate that starting maintenance therapy when patients are younger and possess more teeth may prevent future tooth loss.

Tumor necrosis factor-alpha (TNF-α) is an inflammatory cytokine known to cause bone resorption, swelling and edema during tissue organization. Conversely, TNF-α has also been shown to participate in tissue regeneration during the wound healing process. We have previously investigated the effects of TNF-α on human dental pulp cell differentiation. Dental pulp cells are composed of different cell types including primary odontoblasts and fibroblasts. We determined that the ratio of stem cells within the pulp cell population was increased following short-term stimulation with TNF-α. The aim of this study therefore was to investigate the effect of short-term stimulation with TNF-α on osteoblast-like MC3T3-E1 cell growth and differentiation. MC3T3-E1 cells were cultured in standard growth medium and on reaching sub-confluence were exposed to recombinant TNF-α (10 and 100 ng/ml) for 2 days prior to assessing their cell proliferation and differentiation properties in comparison to non-stimulated MC3T3-E1 cells (control). Although no significant differences in cell proliferation were observed between the TNF-α-stimulated and control groups, cell differentiation was delayed in the TNF-α-stimulated groups. In summary, short-term stimulation of cultured MC3T3-E1 cells with TNF-α had only minimal effect on their growth and differentiation.

The development of organoid techniques for regenerative therapy has progressed remarkably with the use of tissue-derived stem cells and pluripotent stem cells based on stem cell biology and tissue engineering technology. To realize whole-organ replacement therapy as next-generation regenerative medicine, it is expected that fully functional bioengineered organs can be reconstructed using an in vitro three-dimensional (3D) bioengineered organ germ and organoids by stem cell manipulation and self-organization. In this mini-review, we focused on substantial advances of 3D bioengineering technologies for the regeneration of complex oral organs with the reconstruction of 3D bioengineered organ germ using organ-inductive potential embryo-derived epithelial and mesenchymal cells. These bioengineering technologies have the potential for realization of future organ replacement therapy.

Background: Active learning is a concept that allows students to study and learn actively by themselves to get knowledge. There are several methods of active learning, including flipped classroom (FC) and team-based learning (TBL). In FC, students are required to study before classes. In TBL, students study before the class, take the Individual Readiness Assurance Test (IRAT) and Group Readiness Assurance Test (GRAT), and then discuss Group Assignment Projects (GAPs) during class. The purpose of this study was to compare the effectiveness between FC and TBL using longitudinal term-end examination data. Methods: Flipped classroom and TBL effectiveness was assessed from the results of the term-end examinations at the end of the semester from 2014 to 2017. The students were asked to answer questions on the favorable and unfavorable responses of FC and TBL. To check the difficulty of the term-end examinations, control dentists took the same examinations. The dentists were clinical prosthodontic residents who graduated within 3 years from undergraduate course. All prosthodontic residents attended the referential examinations. Results: The term-end examination score of FC and TBL did not show a statistical difference. Multi-way ANOVA showed that the referential examination scores by the dentists were significantly higher than that of the students (P < 0.0001). According to the students, the favorable responses of FC and TBL were on the study habit and the video contents, while the unfavorable responses were mainly on the study materials. Conclusions: There is no statistical difference between FC and TBL on term-end examination scores. There were no interactions between the test period and the participants (students or dentists), and the test period and class format.

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.

Background Active learning is a concept that allows students to study and learn actively by themselves to get knowledge. There are several methods of active learning, including flipped classroom (FC) and team-based learning (TBL). In FC, students are required to study before classes. In TBL, students also study before class. The students take individual readiness assurance test (IRAT) and group readiness assurance test (GRAT), then discuss group assignment projects (GAPs) during class. The purpose of this study was to compare the effectiveness between FC and TBL using longitudinal term-end examination data.Methods FC and TBL effectiveness was assessed from the results of the term-end examinations from 2014 to 2017. The students were asked to answer questions on the favorable and unfavorable responses of FC and TBL. To check the difficulty of the term-end examinations, control dentists took the same examinations.Results Multi-way ANOVA showed that the correct answer rate in term-end examinations was significantly different in comparison with the time of the trial, and for the participants (students and dentists). The term-end examination score of FC and TBL did not show a statistical difference. According to the students, the favorable responses of FC and TBL were on the study habit and the video contents, while the unfavorable responses were mainly on the study materials.Conclusions There is no statistical difference between FC and TBL on term-end examination scores. There were no interactions between the test period and the participants (students or dentists), and the test period and class format.

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.

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., Ishikawa, Takeo et al. present a 3D liver sinusoidal network analysis method and identify important parameters after partial hepatectomy. They find that in coordination with cytokine networks, mechanical homeostatic signalling, including shear stress and tension, plays critical roles in the initiation and termination of liver organ regeneration.

Oral function and associated health issues are impaired by irreversible dental problems, including dental caries and periodontal disease. Tooth regenerative therapy for tissue repair and whole tooth replacement are novel treatment approaches with the potential to fully recover tooth function. The dental tissue repair method uses stem cells and cytokines but not artificial materials to promote odontogenesis. Whole tooth replacement therapy as a form of bioengineered organ replacement is regarded as an important model system for the development of this concept. This article reviews recent findings and technologies underpinning tooth regenerative therapy.

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 &mu, L of 10&minus, 5 M), Minocycline (5 &mu, L containing 10 &mu, g) 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 &deg, C). Quantitative real-time PCR was performed to evaluate the mRNA expression of IL-1&beta, IL-6, TNF-&alpha, 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&ndash, 24 h (paired-t test, p &lt, 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-1&beta, and anti-inflammatory cytokine IL-1RA and NaV1.7 (one-way ANOVA followed by Dunnett&rsquo, s post hoc test, p &lt, 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-1&beta, and IL-6 cytokines significantly at 6 h (t-test, p &lt, 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.

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.

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.

© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd 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

Background A merit of subnormothermic perfusion has been reported to preserve grafts from ischemic injury in animal models. The split liver technique is commonly performed to solve the shortage of liver grafts. However, there has been no study showing the effect of a split liver graft on subnormothermic perfusion. We herein investigated the split liver protocol using a subnormothermic oxygenated circuit system (SOCS). Methods Auxiliary liver transplantation was performed in a porcine marginal donor model by using a SOCS. In the SOCS group, the portal vein and hepatic artery of the graft were cannulated, and the graft was perfused by SOCS. In the cold storage (CS) group, the graft was placed in cold preservation solution. In the preservation phase, the graft was split. Results There were no significant differences in the biochemical markers between the SOCS and CS groups. In terms of the histology, the sinusoidal spaces were widened in the CS group 12 hours after implantation. Conclusion We have demonstrated a possibility to use SOCS with the split liver protocol by using a porcine model. This split liver protocol using SOCS will extend the split liver criteria and rescue more patients from hepatic failure, including pediatric patients.

Three-dimensional organogenesis in vivo is principally regulated by the spatiotemporal developmental process that relies on the cellular behavior such as cell growth, migration, differentiation, and cell-to-cell interaction. Organ development and morphogenesis have been elucidated to be regulated by the proper transient expression of various signaling molecules including cytokines, extracellular matrix, and adhesion molecules based on the epithelial and mesenchymal interactions. Current bioengineering technology for regenerating three-dimensional organ has progressed to the replication of organogenesis, thereby enabling the development of fully functional bioengineered organs using bioengineered organ germs that are generated from immature stem cells via tissue engineering technology in vitro.To achieve precise replication of organogenesis, we have developed a novel three-dimensional cell manipulation method designated the organ germ method, and enabled the generation of a structurally correct and fully functional bioengineered tooth in vivo. This method is also expected to be utilized for analyzing gene and protein functions during organogenesis. Here, we describe protocols for the tooth germ reconstitution by using the organ germ method and for the functional analysis of tooth development in vitro and in vivo.

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.

Purpose The aim of this study was to compare saliva flow and protein composition induced using five basic taste stimulations between natural and bioengineered salivary glands. Materials and methods We developed a mouse saliva secretion model using taste stimulation and analyzed the saliva secretion from natural and bioengineered salivary glands using an assay. The protein components and alpha-amylase in the natural and bioengineered saliva were analyzed by gel electrophoresis and Western blotting. Results The salivary flow responses induced by sour (citric acid) and bitter (quinine-HCl) stimuli were significantly high in the natural and bioengineered salivary glands. Although the protein concentrations in the natural and bioengineered saliva induced using five basic taste stimulations were similar, the protein composition and the amylase concentration in the natural saliva after taste stimulation had different profiles. Sympathetic and non-sympathetic nerves were observed around the acini and ducts in the natural and bioengineered salivary glands. However, the frequency of neuropeptide Y-positive sympathetic nerves in the bioengineered gland was relatively high compared to that in the natural gland. Conclusions These results suggest that the signal balance between the sympathetic and parasympathetic components of the efferent nerves in an engrafted bioengineered salivary gland may differ from that in a natural salivary gland.

Dental disorders, including dental caries and periodontal disease, can cause fundamental problems for oral functions, such as enunciation, mastication and occlusion, as well as general health issues. Tooth regenerative therapies for tissue repair and whole tooth replacement are currently being developed as novel treatment approaches. As a form of bioengineered organ replacement, whole tooth replacement therapy is considered an important model system for next-generation regenerative therapy. We recently reported bioengineered tooth replacements after transplantation of a bioengineered tooth germ or mature tooth unit comprising the bioengineered tooth and periodontal tissues. Whole tooth regenerative therapy has the potential to fully restore tooth function, including masticatory potential in response to mechanical stress and perceptive potential for noxious stimulation. In this review, we describe recent findings and technologies underpinning tooth regenerative therapy.

Three-dimensional organogenesis in vivo is principally regulated by the spatiotemporal developmental process that relies on the cellular behavior such as cell growth, migration, differentiation, and cell-to-cell interaction. Organ development and morphogenesis have been elucidated to be regulated by the proper transient expression of various signaling molecules including cytokines, extracellular matrix, and adhesion molecules based on the epithelial and mesenchymal interactions. Current bioengineering technology for regenerating three-dimensional organ has progressed to the replication of organogenesis, thereby enabling the development of fully functional bioengineered organs using bioengineered organ germs that are generated from immature stem cells via tissue engineering technology in vitro.To achieve precise replication of organogenesis, we have developed a novel three-dimensional cell manipulation method designated the organ germ method, and enabled the generation of a structurally correct and fully functional bioengineered tooth in vivo. This method is also expected to be utilized for analyzing gene and protein functions during organogenesis. Here, we describe protocols for the tooth germ reconstitution by using the organ germ method and for the functional analysis of tooth development in vitro and in vivo.

in vivo in vivo in vivo in vivo. Whole-tooth replacement therapy is regarded as an important model system for the development of the concept of organ regeneration. A novel three-dimensional in vitro cell manipulation method, designated as an organ germ method, has been developed to recapitulate organogenesis. This method involves cell compartmentalization between epithelial and mesenchymal cells at a high cell density to mimic the multicellular assembly and epithelialmesenchymal interactions. The bioengineered tooth germ generates a structurally correct tooth in vitro, and erupted successfully with correct tooth structure when transplanted into a tooth socket in the oral cavity. We could also generate a size-controlled bioengineered mature tooth unit composed of periodontal ligament and alveolar bone. The bioengineered tooth unit was successfully engrafted into an adult jaw through bone integration. These bioengineered teeth were able to perform physiological tooth functions such as mastication, periodontal ligament function and response to noxious stimuli. Here, we review recent studies of tooth tissue-derived mesenchymal stem cells and the technologies underpinning tooth regenerative therapy.

Current osseointegrated dental implants have been widely used for the rehabilitation of tooth loss. Although dental implants are considered an available treatment in the paradigm shift from traditional dental therapies, such as fixed dental bridges and removable dentures, the fundamental problems must be overcome before their clinical use in young patients who are still undergoing jawbone growth. Here, we show a novel bioengineering method for a functional biohybrid implant that is combined with adult-derived periodontal tissue and attached with bone tissue as a substitute for cementum. This biohybrid implant was successfully engrafted using the bioengineered periodontal ligament, and it restored physiological function, such as orthodontic movement through bone remodeling and appropriate responsiveness to noxious stimuli. Thus, this study represents the functional biohybrid implant's potential for clinical use as a next-generation dental implant using adult-derived tissues.

Cell attachment is a crucial step in tissue regeneration. In this study, human bone marrow stromal cells (hBMSCs) were isolated, and the effects of CCN2 on their attachment were examined. CCN2 significantly enhanced the hBMSC attachment, and this enhanced cell attachment was mainly regulated by the C-terminal module of CCN2. This enhancement was negated by the anti-integrin alpha(v)beta(3) antibody and p38 MAPK inhibitor, and phosphorylation of p38 MAPK was detected upon the enhanced cell attachment mediated by CCN2. We thus conclude that CCN2 enhances hBMSC attachment via integrin-p38 MAPK signal pathway. Enhanced hBMSC attachment on hydroxyapatite plates by CCN2 further indicated the utility of CCN2 in bone regeneration.

Whole-organ regeneration has great potential for the replacement of dysfunctional organs through the reconstruction of a fully functional bioengineered organ using three-dimensional cell manipulation in vitro. Recently, many basic studies of whole-tooth replacement using three-dimensional cell manipulation have been conducted in a mouse model. Further evidence of the practical application to human medicine is required to demonstrate tooth restoration by reconstructing bioengineered tooth germ using a postnatal large-animal model. Herein, we demonstrate functional tooth restoration through the autologous transplantation of bioengineered tooth germ in a postnatal canine model. The bioengineered tooth, which was reconstructed using permanent tooth germ cells, erupted into the jawbone after autologous transplantation and achieved physiological function equivalent to that of a natural tooth. This study represents a substantial advancement in whole-organ replacement therapy through the transplantation of bioengineered organ germ as a practical model for future clinical regenerative medicine.

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.

Biological tissues have specific cell patterns and well-organized structures which result from various organogenesis processes. Among which is branching morphogenesis which is the key developmental processes for many glandular tissues including lungs, kidney, salivary gland and liver. Many approaches were suggested to regenerate salivary gland tissue in vitro. Here, we developed a 3D culture technique to study and recapitulate the dynamic forces generated during SMG organogenesis by utilizing time lapse live imaging to monitor the cellular migration and reorganization process of both epithelium and mesenchyme cells.

A novel method is used to develop a functional 3D integumentary organ system, including appendage organs from iPS cells., The integumentary organ system is a complex system that plays important roles in waterproofing, cushioning, protecting deeper tissues, excreting waste, and thermoregulation. We developed a novel in vivo transplantation model designated as a clustering-dependent embryoid body transplantation method and generated a bioengineered three-dimensional (3D) integumentary organ system, including appendage organs such as hair follicles and sebaceous glands, from induced pluripotent stem cells. This bioengineered 3D integumentary organ system was fully functional following transplantation into nude mice and could be properly connected to surrounding host tissues, such as the epidermis, arrector pili muscles, and nerve fibers, without tumorigenesis. The bioengineered hair follicles in the 3D integumentary organ system also showed proper hair eruption and hair cycles, including the rearrangement of follicular stem cells and their niches. Potential applications of the 3D integumentary organ system include an in vitro assay system, an animal model alternative, and a bioengineered organ replacement therapy.

A three-dimensional multicellular organism maintains the biological functions of life support by using the blood circulation to transport oxygen and nutrients and to regulate body temperature for intracellular enzymatic reactions. Donor organ transplantation using low-temperature storage is used as the fundamental treatment for dysfunctional organs. However, this approach has a serious problem in that donor organs maintain healthy conditions only during short-term storage. In this study, we developed a novel liver perfusion culture system based on biological metabolism that can maintain physiological functions, including albumin synthesis, bile secretion and urea production. This system also allows for the resurrection of a severely ischaemic liver. This study represents a significant advance for the development of an ex vivo organ perfusion system based on biological metabolism. It can be used not only to address donor organ shortages but also as the basis of future regenerative organ replacement therapy.