Iroquois homeobox 3 regulates odontoblast proliferation and differentiation mediated by Wnt5a expression.

Iroquois homeobox (Irx) genes are TALE-class homeobox genes that are evolutionarily conserved across species and have multiple critical cellular functions in fundamental tissue development processes. Previous studies have shown that Irxs genes are expressed during tooth development. However, the precise roles of genes in teeth remain unclear. Here, we demonstrated for the first time that Irx3 is an essential molecule for the proliferation and differentiation of odontoblasts. Using cDNA synthesized from postnatal day 1 (P1) tooth germs, we examined the expression of all Irx genes (Irx1-Irx6) by RT-PCR and found that all genes except Irx4 were expressed in the tooth tissue. Irx1-Irx3 a were expressed in the dental epithelial cell line M3H1 cells, while Irx3 and Irx5 were expressed in the dental mesenchymal cell line mDP cells. Only Irx3 was expressed in both undifferentiated cell lines. Immunostaining also revealed the presence of IRX3 in the dental epithelial cells and mesenchymal condensation. Inhibition of endogenous Irx3 by siRNA blocks the proliferation and differentiation of mDP cells. Wnt3a, Wnt5a, and Bmp4 are factors involved in odontoblast differentiation and were highly expressed in mDP cells by quantitative PCR analysis. Interestingly, the expression of Wnt5a (but not Wnt3a or Bmp4) was suppressed by Irx3 siRNA. These results suggest that Irx3 plays an essential role in part through the regulation of Wnt5a expression during odontoblast proliferation and differentiation.

Detergent exposure induces epithelial barrier dysfunction and eosinophilic inflammation in the esophagus.

BACKGROUND: Eosinophilic esophagitis (EoE) is a chronic allergic disease associated with type 2 inflammation and epithelial barrier dysfunction. The etiology is unknown, however, genetic heritability studies suggest environmental factors play a key role in pathogenesis. Detergents, such as sodium dodecyl sulfate (SDS), are common ingredients in household products such as dish soap and toothpaste. We hypothesized detergent exposure decreases epithelial barrier function and induces esophageal inflammation. METHODS: Immortalized esophageal epithelial cells (EPC2) were cultured in air-liquid interface (ALI) and exposed to SDS. Barrier function/activity was assessed by transepithelial electrical resistance (TEER), FITC-dextran flux, and RT-PCR. Additionally, SDS-treated mouse esophageal organoids were evaluated for morphology. To investigate the effects of SDS in vivo, mice were treated with 0.5% SDS in drinking water for 14 days. Esophagi were assessed by gross morphology, histopathology, protein expression, and bulk RNA sequencing. RESULTS: When EPC2 cells were exposed to SDS (5 µg/ml) for 96 h, TEER decreased (p = 0.03), and FITC-dextran flux increased (p = 0.0002). mRNA expression of IL-33 increased 4.5-fold (p = 0.02) at 6 h and DSG1 decreased (p < 0.0001) by 72 h. Disrupted epithelial integrity was noted in SDS-treated esophageal organoids. When mice were exposed to SDS, they showed increased esophageal width, chemokine, and metalloprotease levels. Mice treated with SDS also showed increased IL-33 protein expression, basal zone hyperplasia, CD4+ cell infiltration, and esophageal eosinophilia. RNA sequencing revealed upregulation of immune response pathway genes. CONCLUSION: Exposure to SDS decreases esophageal barrier integrity, stimulates IL-33 production, and promotes epithelial hyperplasia and tissue eosinophilia. Detergents may be a key environmental trigger in EoE pathogenesis.

Oral-maxillofacial trauma of a geriatric population in a super-ageing country.

BACKGROUND/AIM: World population has been ageing, and oral-maxillofacial trauma of geriatric population is expected to increase. The aim of this study was to analyse the characteristic features of oral-maxillofacial trauma in the geriatric population. MATERIALS AND METHODS: Data from 127 patients aged 65 years old or older, who were treated for oral-maxillofacial trauma at the Department of Oral and Maxillofacial Surgery, Hirosaki University, from 2000 to 2014, were retrospectively analysed. The data from 292 patients aged 20-64 years were used as a comparison. RESULTS: Oral-maxillofacial trauma in the geriatric population had been increasing over 15-year period. The male to female ratio was 1.05:1 in the older group and 2.3:1 in the younger group. In the older group, 117 patients (92.1%) had one or more underlying systemic diseases, and 16 (12.6%) had suffered injuries in association with acute medical disorders. The most common injuries in the older group were bone fractures (46.5%). The ratio of fractures in the older group was lower than in the younger group (69.2%). Trauma in the older group most frequently occurred because of falls from a standing height or lower (52.0%), and the mandible was the most common site of fracture (74.6%). A conservative form of treatment for maxillofacial fractures was most commonly (86.4%) chosen for the older group, whilst surgical treatment was most commonly in the younger group (55.0%). CONCLUSION: Oral-maxillofacial trauma in the geriatric population shows characteristic features in terms of aetiology, patterns and treatment modalities.

Impalement of an unusual foreign body on the temporomandibular joint causing severe trismus.

A penetrating injury by a foreign body is comparatively common in the oral and maxillofacial region. On the other hand, injury to the temporomandibular joint (TMJ) by a foreign object is very rare. The TMJ is an anatomically narrow space surrounded by hard bony processes. An unusual case of trauma with severe trismus caused by a foreign body that impaled the TMJ is reported. A 55-year-old man presented with a 5 × 1-cm laceration to the right cheek caused by a flying object propelled during the use of a lawn mower. The edge of the foreign body had a metallic wire, which became imbedded in the wound. His jaw opening was severely limited. Computed tomography revealed that the foreign body was 3 mm in diameter and was impaled on the articular capsule. The object was successfully removed, and the wound and interior of the TMJ were irrigated. Rehabilitation of mouth opening was started on postoperative day 3. On day 9, mouth opening had improved to 35 mm, and he was discharged. After 1 year, mouth opening was 45 mm with no sign of any TMJ disorders.

An ATF4-Signal-Modulating Machine Other Than GADD34 Acts in ATF4-to-CHOP Signaling to Block CHOP Expression in ER-Stress-Related Autophagy.

Cells respond to ER-stress via ER-stress sensors, leading to the UPR and subsequent apoptosis; however, occasionally, they activate autophagy without subsequent apoptosis in response to ER-stress. We previously showed that the induction of apoptosis by ER-stress was related to the presence or absence of CHOP expression; nevertheless, how ATF4 expression is elicited without downstream CHOP expression is unknown. We studied the role of GADD34 on the induction of autophagy and/or apoptosis by NaF- or tunicamycin-induced ER-stress in HepG2 cells transfected with GADD34 siRNA. Although NaF and tunicamycin both induced PERK activation followed by eIF2α phosphorylation and ATF4 expression, CHOP expression was only induced by tunicamycin. Concomitant with the signaling change, autophagy was activated both by NaF and tunicamycin, and apoptosis was induced only by tunicamycin. After 4 h, GADD34 mRNA expression was also increased by NaF and tunicamycin. Suppression of GADD34 by GADD34 siRNA increased ATF4 expression in both NaF- and tunicamycin-treated cells. The GADD34 siRNA increased CHOP expression, which corresponded to increased ATF4 in tunicamycin-treated cells; however, the increased ATF4 did not induce CHOP expression in NaF-treated cells. In concert with signal changes, siRNA treatment additively increased the autophagic activity of both NaF- and tunicamycin-treated cells; however, apoptosis was produced and accelerated only for tunicamycin-treated cells. These findings indicate that GADD34 expression induced by ER-stress delays CHOP expression and retards apoptotic cell death, and that an ATF4-signal-modulating machine other than GADD34 acts on ATF4-to-CHOP signaling to block ATF4-induced CHOP expression in ER-stress related autophagy.

Professional oral health care reduces oral mucositis pain in patients treated by superselective intra-arterial chemotherapy concurrent with radiotherapy for oral cancer.

PURPOSE: Oral mucositis (OM) is a painful complication of radiation therapy (RT) for head and neck cancer. OM can compromise nutrition, require opioid analgesics and hospitalization for pain control, and lead to interruption of treatment. Severe oral mucositis appears inevitable in superselective intra-arterial chemotherapy concurrent with radiotherapy (SSIACRT), requiring management of OM for the patient. The objective of this study was to assess the utility of professional oral health care (POHC) for the management of OM in patients undergoing SSIACRT. METHODS: Thirty-three patients were enrolled in this study. The first 17 patients underwent SSIACRT before we created an oral management team, and thus did not receive POHC. The remaining 16 patients received POHC. Fever duration, duration of oral feeding difficulty, opioid usage, duration of opioid administration, duration of hospitalization, and number of hospital days from the end of irradiation to discharge were compared between these two groups. RESULTS: Median total dose of morphine during SSIACRT, median number of hospital days from end of irradiation to discharge, and duration of hospitalization all differed significantly between groups (P < 0.05). Duration of opioid administration, fever duration, and duration of oral feeding difficulty did not differ significantly between groups. CONCLUSIONS: These findings indicate that POHC may reduce opioid use and shorten the hospital stay. Such results might be obtained through infection control by POHC. This report appears to be the first study to evaluate the efficiency of POHC in SSIACRT for oral cancer from the perspective of mucositis pain and opioid use.

Polyelectrolyte-protein synergism: pH-responsive polyelectrolyte/insulin complexes as versatile carriers for targeted protein and drug delivery.

The co-assembly of polyelectrolytes (PE) with proteins offers a promising approach for designing complex structures with customizable morphologies, charge distribution, and stability for targeted cargo delivery. However, the complexity of protein structure limits our ability to predict the properties of the formed nanoparticles, and our goal is to identify the key triggers of the morphological transition in protein/PE complexes and evaluate their ability to encapsulate multivalent ionic drugs. A positively charged PE can assemble with a protein at pH above isoelectric point due to the electrostatic attraction and disassemble at pH below isoelectric point due to the repulsion. The additional hydrophilic block of the polymer should stabilize the particles in solution and enable them to encapsulate a negatively charged drug in the presence of PE excess. We demonstrated that diblock copolymers, poly(ethylene oxide)-block-poly(N,N-dimethylaminoethyl methacrylate) and poly(ethylene oxide)-block-poly(N,N,N-trimethylammonioethyl methacrylate), consisting of a polycation block and a neutral hydrophilic block, reversibly co-assemble with insulin in pH range between 5 and 8. Using small-angle neutron and X-ray scattering (SANS, SAXS), we showed that insulin arrangement within formed particles is controlled by intermolecular electrostatic forces between protein molecules, and can be tuned by varying ionic strength. For the first time, we observed by fluorescence that formed protein/PE complexes with excess of positive charges exhibited potential for encapsulating and controlled release of negatively charged bivalent drugs, protoporphyrin-IX and zinc(II) protoporphyrin-IX, enabling the development of nanocarriers for combination therapies with adjustable charge, stability, internal structure, and size.

Fracture of the oxidized zirconium femoral component after total knee arthroplasty.

BACKGROUND: Femoral component fracture is a rare complication of total knee arthroplasty (TKA). CASE: We report a case of oxidized zirconium (Oxinium) femoral component fracture after total knee arthroplasty. The fracture site was the junction of the central and medial flanges. The patellar component and polyethylene insert had delamination at the contact point of the fracture line, and the tibial tray had loosening at the medial side. There was no cement adherence at the component fracture site, suggesting that debonding had occurred at the cement-implant interface in this area. Examination with a scanning electron microscope revealed beach marks, which are characteristic findings of metal fatigue. CONCLUSION: We considered that the cause of femoral component fracture was a fatigue fracture due to poor fixation of the component to the bone caused by poor osteotomy technique or poor cementing technique. To our knowledge, this is the first case of Oxinium femoral component fracture.

Fibroblast growth factor 2 suppresses the expression of C-C motif chemokine 11 through the c-Jun N-terminal kinase pathway in human dental pulp-derived mesenchymal stem cells.

The regulation of the mesenchymal stem cell (MSC) programming mechanism promises great success in regenerative medicine. Tissue regeneration has been associated not only with the differentiation of MSCs, but also with the microenvironment of the stem cell niche that involves various cytokines and immune cells in the tissue regeneration site. In the present study, fibroblast growth factor 2 (FGF2), the principal growth factor for tooth development, dental pulp homeostasis and dentin repair, was reported to affect the expression of cytokines in human dental pulp-derived MSCs. FGF2 significantly inhibited the expression of chemokine C-C motif ligand 11 (CCL11) in a time- and dose-dependent manner in the SDP11 human dental pulp-derived MSC line. This inhibition was diminished following treatment with the AZD4547 FGF receptor (FGFR) inhibitor, indicating that FGF2 negatively regulated the expression of CCL11 in SDP11 cells. Furthermore, FGF2 activated the phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK), extracellular signal-regulated kinase 1/2 (ERK1/2) and c-Jun N-terminal kinases (JNK) in SDP11 cells. The mechanism of the FGFR-downstream signaling pathway was then studied using the SB203580, U0126 and SP600125 inhibitors for p38 MAPK, ERK1/2, and JNK, respectively. Interestingly, only treatment with SP600125 blocked the FGF2-mediated suppression of CCL11. The present results suggested that FGF2 regulated the expression of cytokines and suppressed the expression of CCL11 via the JNK signaling pathway in human dental pulp-derived MSCs. The present findings could provide important insights into the association of FGF2 and CCL11 in dental tissue regeneration therapy.

Combination of ions promotes cell migration via extracellular signal­regulated kinase 1/2 signaling pathway in human gingival fibroblasts.

Wound healing is a dynamic process that involves highly coordinated cellular events, including proliferation and migration. Oral gingival fibroblasts serve a central role in maintaining oral mucosa homeostasis, and their functions include the coordination of physiological tissue repair. Recently, surface pre­reacted glass­ionomer (S­PRG) fillers have been widely applied in the field of dental materials for the prevention of dental caries, due to an excellent ability to release fluoride (F). In addition to F, S­PRG fillers are known to release several types of ions, including aluminum (Al), boron (B), sodium (Na), silicon (Si) and strontium (Sr). However, the influence of these ions on gingival fibroblasts remains unknown. The aim of the present study was to examine the effect of various concentrations of an S­PRG filler eluate on the growth and migration of gingival fibroblasts. The human gingival fibroblast cell line HGF­1 was treated with various dilutions of an eluent solution of S­PRG, which contained 32.0 ppm Al, 1,488.6 ppm B, 505.0 ppm Na, 12.9 ppm Si, 156.5 ppm Sr and 136.5 ppm F. Treatment with eluate at a dilution of 1:10,000 was observed to significantly promote the migration of HGF­1 cells. In addition, the current study evaluated the mechanism underlying the mediated cell migration by the S­PRG solution and revealed that it activated the phosphorylation of extracellular signal­regulated kinase 1/2 (ERK1/2), but not of p38. Furthermore, treatment with a MEK inhibitor blocked the cell migration induced by the solution. Taken together, these results suggest that S­PRG fillers can stimulate HGF­1 cell migration via the ERK1/2 signaling pathway, indicating that a dental material containing this type of filler is useful for oral mucosa homeostasis and wound healing.

Recruitment of mesenchymal stem cells by stromal cell-derived factor 1α in pulp cells from deciduous teeth.

Dental pulp cells (DPCs), including dental pulp (DP) stem cells, play a role in dentine repair under certain conditions caused by bacterial infections associated with caries, tooth fracture and injury. Mesenchymal stem cells (MSCs) have also been shown to be involved in this process of repair. However, the mechanisms through which MSCs are recruited to the DP have not yet been elucidated. Therefore, the aim of the present in vitro study was to investigate whether stromal cell-derived factor 1α (SDF1)-C-X-C chemokine receptor type 4 (CXCR4) signaling is involved in tissue repair in the DP of deciduous teeth. A single-cell clone from DPCs (SDP11) and UE7T-13 cells were used as pulp cells and MSCs, respectively. The MG-63 and HuO9 cells, two osteosarcoma cell lines, were used as positive control cells. Reverse transcription polymerase chain reaction (RT-PCR) revealed that all cell lines (SDP11, UE7T-13 MG-63 and HuO9) were positive for both SDF1 and CXCR4 mRNA expression. Moreover, immunocytochemical analysis indicated that SDF1 and CXCR4 proteins were expressed in the SDP11 and UE7T-13 cells. SDF1 was also detected in the cell lysates (CLs) and conditioned medium (CM) collected from the SDP11 and UE7T-13 cells, and AMD3100, a specific antagonist of CXCR4, inhibited the migration of the UE7T-13 cells; this migration was induced by treatment with CM, which was collected from the SDP11 cells. In addition, real-time PCR showed that the expression of SDF1 in the SDP11 cells was inhibited by treatment with 20 ng/ml fibroblast growth factor (FGF)-2, and exposure to AZD4547, an inhibitor of the FGF receptor, blocked this inhibition. Collectively, these data suggest that SDF1 produced by DP plays an important role in homeostasis, repair and regeneration via the recruitment of MSCs.

Quercetin suppresses bone resorption by inhibiting the differentiation and activation of osteoclasts.

Although quercetin has suppressed bone resorption in several animal studies, its target cells and the mechanism of its action related to bone resorption has not been fully elucidated. We investigated the effect of quercetin on the differentiation and activation of osteoclasts. We used cocultures of mouse spleen cells and ST2 cells, and cultures of osteoclast progenitor cells [M-CSF-dependent (MD) cells from mouse bone marrow and murine monocytic RAW 264 (RAW) cells]. Quercetin dose-dependently inhibited osteoclast-like (OCL) cell formation at 2-5 microM concentration in both the coculture and MD cell culture. Quercetin inhibited the increase of tartrate-resistant acid phosphatase (TRAP) activity of mononuclear preosteoclasts (pOCs) induced by receptor activator of nuclear factor-kappaB (NF-kappaB) ligand (RANKL) in both MD and RAW cell cultures. Quercetin reversely induced the disruption of actin rings in OCLs. Quercetin also suppressed both pit formation induced by osteoclasts on dentine slices and PTH-stimulated (45)Ca release in mouse long bone cultures. These results suggest that osteoclast progenitors as well as mature osteoclasts, are quercetin's target cells in relation to bone resorption, and that quercetin's suppressive effect on bone resorption results from both its inhibitory effect on the differentiation of osteoclast progenitor cells into pOCs and from its disruptive effect on actin rings in mature osteoclasts.