Kyushu University Academic Staff Educational and Research Activities Database
List of Papers
ATSUTA IKIRU Last modified date:2022.06.27

Associate Professor / Division of Advanced Dental Devices and Therapeutics, Faculty of Dental Science Kyushu University / Department of Dental Science / Faculty of Dental Science

1. Yu Feng Mei, Takayoshi Yamaza, Ikiru Atsuta, Atsushi Danjo, Yoshio Yamashita, Mizuho A Kido, Masaaki Goto, Akifumi Akamine, Teruo Tanaka, Sequential expression of endothelial nitric oxide synthase, inducible nitric oxide synthase, and nitrotyrosine in odontoblasts and pulp cells during dentin repair after tooth preparation in rat molars., Cell and tissue research, 328, 1, 117-27, 2007.04, Nitric oxide (NO) stimulates osteoblast differentiation, but whether NO contributes to odontoblast differentiation during dentin repair is unknown. By using reverse transcription/polymerase chain reaction and immunostaining, we investigated the gene expression and/or immunolocalization of endothelial NO synthase (eNOS), inducible NOS (iNOS), and nitrotyrosine (a biomarker for NO-derived peroxinitrite), and alkaline phosphatase (ALP) and osteocalcin (early and terminal differentiation markers of odontoblasts, respectively) in dental pulp tissue after rat tooth preparation. At the early stage (1-3 days) post-preparation, markedly increased expression of iNOS and nitrotyrosine was found in odontoblasts and pulp cells beneath the cavity, whereas eNOS expression was significantly decreased. ALP mRNA expression was significantly increased after 1 day but decreased after 3 days, whereas ALP activity was weak in the dentin-pulp interface under the cavity after 1 day but strong after 3 days. Osteocalcin mRNA expression was significantly increased at this stage. At 7 days post-preparation, tertiary dentin was formed under the cavity. All the molecules studied were expressed at control levels in odontoblasts/pulp cells beneath the cavity. These findings show that abundant NO is released from odontoblasts and pulp cells at an early stage after tooth preparation and indicate that, after tooth preparation, the up-regulation of iNOS and nitrotyrosine in odontoblasts is synchronized with increased cellular expression of ALP and osteocalcin. Therefore, the NO synthesized by iNOS after tooth preparation probably participates in regulating odontoblast differentiation during tertiary dentinogenesis..
2. Yasuko Moriyama, Yasunori Ayukawa, Yoichiro Ogino, Ikiru Atsuta, Kiyoshi Koyano, Topical application of statin affects bone healing around implants., Clinical oral implants research, 10.1111/j.1600-0501.2007.01508.x, 19, 6, 600-5, 2008.06, OBJECTIVES: 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) are widely used for hyperlipidemia. Recent studies demonstrate that statins stimulate bone morphogenetic protein-2 expression and lead to bone formation. The aim of this study was to evaluate whether the topical application of statin enhances the osteogenesis around a titanium implant. MATERIALS AND METHODS: Ten-week-old female rats received pure titanium rods in both tibiae with or without fluvastatin. Propylene glycol alginate (PGA) was used as a carrier. The rats were divided into five groups: implant-only group, implant with PGA group, low-dose group [implant+PGA containing 3 microg of fluvastatin (FS)], medium-dose group (15 microg of FS), and high-dose group (75 microg of FS). The animals were sacrificed at 1 and 2 weeks after implantation. Peri-implant bone formation was assessed by histomorphometric procedures, i.e., measuring the bone-implant contact (BIC) and peri-implant bone volume (BV). A mechanical push-out test was also performed to evaluate the implant fixation strength. Statistical differences among the groups were determined by ANOVA and P
3. Yohei Jinno, Yasunori Ayukawa, Yoichiro Ogino, Ikiru Atsuta, Yoshihiro Tsukiyama, Kiyoshi Koyano, Vertical bone augmentation with fluvastatin in an injectable delivery system: a rat study., Clinical oral implants research, 10.1111/j.1600-0501.2008.01665.x, 20, 8, 756-60, 2009.08, OBJECTIVES: HMG-CoA reductase inhibitors (statins) are widely used for hyperlipidemia. Previous studies demonstrate that statins stimulate bone morphogenetic protein-2 (BMP-2) expression and lead to bone formation. The aim of this study was to evaluate whether percutaneously injected statin with a novel statin delivery system achieved vertical bone augmentation. MATERIAL AND METHODS: As experimental groups, atelocollagen-alpha-tricalcium phosphate (alphaTCP) composites containing 3.3 mg (low dose) or 6.7 mg (high dose) of fluvastatin were injected (one shot) subcutaneously over the calvarial periosteum of rats. The animals were then sacrificed 1, 2, and 4 weeks after injection. Vertically augmented bone was assessed by histomorphometric procedures, i.e., by measuring new bone thickness (NBT) and bone density (BD). RESULTS: In control groups, no newly formed bone could be seen over the calvarial bone. In the experimental groups, in contrast, a large amount of newly formed bone could be seen over the preexisting calvarial bone. The newly formed bone was seen to be in direct contact with the preexisting bone. During the entire observation, significant NBT was observed in the experimental groups (P
4. Yasunori Ayukawa, Eisuke Yasukawa, Yasuko Moriyama, Yoichiro Ogino, Hiroko Wada, Ikiru Atsuta, Kiyoshi Koyano, Local application of statin promotes bone repair through the suppression of osteoclasts and the enhancement of osteoblasts at bone-healing sites in rats., Oral surgery, oral medicine, oral pathology, oral radiology, and endodontics, 10.1016/j.tripleo.2008.07.013, 107, 3, 336-42, 2009.03, OBJECTIVE: We investigated whether the local administration of simvastatin affected both the cellular events and the bone formation at surgically created bone defects in rat. STUDY DESIGN: Simvastatin (or a vehicle) was injected into a rat bony defect for 3 consecutive days from the day of surgery. Five or ten days after the injection, new bone tissue was collected, and the gene expressions of bone-related proteins were examined. For the histomorphometry, new bone area was measured. RESULTS: At day 5, the statin group demonstrated significantly larger new bone area. The number of tartrate-resistant acid phosphatase-positive multinucleated cells in the statin group was less than in the control group. In the statin group, the expressions of both alkaline phosphatase and bone morphogenetic protein 2 mRNA significantly increased. In contrast, the expression of cathepsin K was significantly suppressed in the statin group. Although the levels of both RANK and osteoprotegerin were not affected by statin, the expression of RANKL was depressed. At day 10, there were no significant differences among the groups in either histomorphometric or reverse-transcription polymerase chain reaction analyses. CONCLUSION: New bone area increased under the influence of simvastatin; however, the effect did not continue when the administration was terminated. Osteoclast suppression may be the consequence of RANKL depression..
5. Tomohiro Masuzaki, Yasunori Ayukawa, Yasuko Moriyama, Yohei Jinno, Ikiru Atsuta, Yoichiro Ogino, Kiyoshi Koyano, The effect of a single remote injection of statin-impregnated poly (lactic-co-glycolic acid) microspheres on osteogenesis around titanium implants in rat tibia., Biomaterials, 10.1016/j.biomaterials.2010.01.016, 31, 12, 3327-34, 2010.04, The aim of this study was to evaluate the effects of newly developed injectable poly (lactic-co-glycolic acid) (PLGA) microspheres containing fluvastatin on osteogenesis around titanium implants in the rat tibia. After confirmation of the sustained-release profile of fluvastatin from the microspheres by an in vitro assay, the microspheres were administered to the back skin of the rats by a single transdermal injection. At 2 and 4 weeks after the implant surgery, the fluvastatin groups showed enhanced new bone formation around the titanium implants without any influence on the serum biochemistry. In addition, the fluvastatin groups showed increased three-point bending strengths of their femurs. The results of this study indicate that a single remote injection of PLGA/fluvastatin microspheres safely and successfully stimulated bone formation around titanium implants and increased the mechanical properties of bone..
6. Yasuko Moriyama, Yasunori Ayukawa, Yoichiro Ogino, Ikiru Atsuta, Mitsugu Todo, Yoshihiro Takao, Kiyoshi Koyano, Local application of fluvastatin improves peri-implant bone quantity and mechanical properties: a rodent study., Acta biomaterialia, 10.1016/j.actbio.2009.10.045, 6, 4, 1610-8, 2010.04, Statins are known to stimulate osteoblast activity and bone formation. This study examines whether local application of fluvastatin enhances osteogenesis around titanium implants in vivo. Ten-week-old rats received a vehicle gel (propylene glycol alginate (PGA)) or PGA containing fluvastatin (3, 15, 75 or 300 microg) in their tibiae just before insertion of the implants. For both histological and histomorphometric evaluations undecalcified ground sections were obtained and the bone-implant contact (BIC), peri-implant osteoid volume and mineralized bone volume (MBV) were calculated after 1, 2 and 4 weeks. Using the same models mechanical push-in tests were also performed to evaluate the implant fixation strength. After 1 week the MBV and push-in strength were significantly lower in the 300 microg fluvastatin-treated group than in the other groups (P
7. Akihiro Furuhashi, Yasunori Ayukawa, Ikiru Atsuta, Hideyuki Okawachi, Kiyoshi Koyano, The difference of fibroblast behavior on titanium substrata with different surface characteristics., Odontology, 10.1007/s10266-011-0029-y, 100, 2, 199-205, 2012.07, Connective tissue, one of the main components of peri-implant soft tissue, is key to the formation of the peri-implant mucosal seal and helping to prevent epithelial ingrowth. Rough surfaces (Rs), machined surfaces (Ms) or microgrooved surface (MG) are used in the neck area of commercially available titanium implants. In this paper, we aimed to evaluate the influence of surface topography of titanium substratum on connective tissue fibroblasts to gain a better understanding of this effect. Fibroblasts were cultured on titanium plates with Rs, Ms and MG. Adhesion cell number at day 3 was compared and protein distribution of both F-actin and vinculin was determined to observe cellular structure and adhesion. Cell adhesion strength was compared on each surface. At day 3, the number of fibroblasts attached on each substratum was in the order of MG ≈ Ms > Rs. Fibroblasts strongly expressed vinculin in the peripheral area on Ms and MG, and showed strong F-actin architecture. Decreased expression of vinculin and weaker continuity of F-actin were observed on Rs. Fibroblasts on MG were aligned along the grooves, with a significantly higher cell density, whereas cells on Ms and Rs had no clear orientation. The cell adhesion strength was significantly lower on Rs, and no significant difference was seen between MG and Ms. Both MG and Ms showed greater adhesion cell numbers and adhesion strength of fibroblasts when compared with Rs at day 3. The cell density on MG was greater than those on other substrata..
8. Yinghua Zhao, Lei Wang, Yi Liu, Kentaro Akiyama, Chider Chen, Ikiru Atsuta, Tao Zhou, Xiaohong Duan, Yan Jin, Songtao Shi, Technetium-99 conjugated with methylene diphosphonate ameliorates ovariectomy-induced osteoporotic phenotype without causing osteonecrosis in the jaw., Calcified tissue international, 10.1007/s00223-012-9649-7, 91, 6, 400-8, 2012.12, Technetium-99 conjugated with methylene diphosphonate ((99)Tc-MDP) is a novel bisphosphonate derivative without radioactivity and has been successfully used to treat arthritis in China for years. Since bisphosphonate therapy has the potential to induce bisphosphonate-related osteonecrosis of the jaw (BRONJ), we examined whether (99)Tc-MDP represents a new class of bisphosphonate for antiresorptive therapy to ameliorate estrogen deficiency-induced bone resorption with less risk of causing BRONJ. We showed that (99)Tc-MDP-treated, ovariectomized (OVX) mice had significantly improved bone mineral density and trabecular bone volume in comparison to the untreated OVX group by inhibiting osteoclasts and enhancing osteogenic differentiation of bone marrow mesenchymal stem cells. To determine the potential of inducing BRONJ, (99)Tc-MDP/dexamethasone (Dex) or zoledronate/Dex was administered into C57BL/6J mice via the tail vein, followed by extraction of maxillary first molars. Interestingly, (99)Tc-MDP treatment showed less risk to induce osteonecrosis in the maxillary bones compared to zoledronate treatment group, partially because (99)Tc-MDP neither suppressed adaptive regulatory T cells nor activated the inflammatory T-helper-producing interleukin-17 cells. Taken together, our findings demonstrate that (99)Tc-MDP therapy may be a promising approach in the treatment of osteoporosis with less risk of causing BRONJ..
9. Hiroshi Egusa, Wataru Sonoyama, Masahiro Nishimura, Ikiru Atsuta, Kentaro Akiyama, Stem cells in dentistry--part I: stem cell sources., Journal of prosthodontic research, 10.1016/j.jpor.2012.06.001, 56, 3, 151-65, 2012.07, Stem cells can self-renew and produce different cell types, thus providing new strategies to regenerate missing tissues and treat diseases. In the field of dentistry, adult mesenchymal stem/stromal cells (MSCs) have been identified in several oral and maxillofacial tissues, which suggests that the oral tissues are a rich source of stem cells, and oral stem and mucosal cells are expected to provide an ideal source for genetically reprogrammed cells such as induced pluripotent stem (iPS) cells. Furthermore, oral tissues are expected to be not only a source but also a therapeutic target for stem cells, as stem cell and tissue engineering therapies in dentistry continue to attract increasing clinical interest. Part I of this review outlines various types of intra- and extra-oral tissue-derived stem cells with regard to clinical availability and applications in dentistry. Additionally, appropriate sources of stem cells for regenerative dentistry are discussed with regard to differentiation capacity, accessibility and possible immunomodulatory properties..
10. Hiroshi Egusa, Wataru Sonoyama, Masahiro Nishimura, Ikiru Atsuta, Kentaro Akiyama, Stem cells in dentistry--Part II: Clinical applications., Journal of prosthodontic research, 10.1016/j.jpor.2012.10.001, 56, 4, 229-48, 2012.10, New technologies that facilitate solid alveolar ridge augmentation are receiving considerable attention in the field of prosthodontics because of the growing requirement for esthetic and functional reconstruction by dental implant treatments. Recently, several studies have demonstrated potential advantages for stem-cell-based therapies in regenerative treatments. Mesenchymal stem/stromal cells (MSCs) are now an excellent candidate for tissue replacement therapies, and tissue engineering approaches and chair-side cellular grafting approaches using autologous MSCs represent the clinical state of the art for stem-cell-based alveolar bone regeneration. Basic studies have revealed that crosstalk between implanted donor cells and recipient immune cells plays a key role in determining clinical success that may involve the recently observed immunomodulatory properties of MSCs. Part II of this review first overviews progress in regenerative dentistry to consider the implications of the stem cell technology in dentistry and then highlights cutting-edge stem-cell-based alveolar bone regenerative therapies. Factors that affect stem-cell-based bone regeneration as related to the local immune response are then discussed. Additionally, pre-clinical stem cell studies for the regeneration of teeth and other oral organs as well as possible applications of MSC-based immunotherapy in dentistry are outlined. Finally, the marketing of stem cell technology in dental stem cell banks with a view toward future regenerative therapies is introduced..
11. Ikiru Atsuta, Yasunori Ayukawa, Yoichiro Ogino, Yasuko Moriyama, Yohei Jinno, Kiyoshi Koyano, Evaluations of epithelial sealing and peri-implant epithelial down-growth around "step-type" implants., Clinical oral implants research, 10.1111/j.1600-0501.2011.02163.x, 23, 4, 459-66, 2012.04, OBJECTIVE: Implant designs that can stimulate and integrate with an epithelial wound-healing process may significantly enhance the efficacy of dental implants. Here, we evaluated the potential of "step-type" implant systems to improve the sealing between the peri-implant epithelium (PIE) and the implant surface, and investigated the effect of implant structure on PIE down-growth. MATERIALS AND METHODS: Right maxillary first molars were extirpated from rats and implanted with either a straight-type or a step-type implant varying in step height and/or width (N(s): 0.8 mm height, 0.1 mm width; W(s): 0.8 mm height, 0.2 mm width; H(s): 0.4 mm height, 0.1 mm width). Maxillae were harvested at various time points over 16 weeks to evaluate laminin-5 distribution as an indicator of wound healing and PIE formation, horse-radish peroxidase (HRP) penetration as a measurement of epithelial sealing, and PIE down-growth formation. RESULTS: In all implant models, the PIE formed from the oral sulcular epithelium and spread apically along the implant surface. In the W(s) group, HRP penetration was detected only in the coronal region of the PIE at 4 weeks, whereas in the straight-type, it was observed in the apical region and the connective tissue. At 16 weeks, the W(s) implants exhibited markedly less PIE down-growth than the Con, N(s) or H(s) implants, and were equivalent to that observed in natural teeth. CONCLUSION: The step-type implant system may have the potential for improving epithelial sealing at the tissue-implant interface, as well as reducing apical PIE down-growth, thus enhancing dental implant efficacy..
12. Hideyuki Okawachi, Yasunori Ayukawa, Ikiru Atsuta, Akihiro Furuhashi, Mami Sakaguchi, Koichi Yamane, Kiyoshi Koyano, Effect of titanium surface calcium and magnesium on adhesive activity of epithelial-like cells and fibroblasts., Biointerphases, 10.1007/s13758-012-0027-9, 7, 1-4, 27-27, 2012.12, In the present study, we investigated the hydrothermal treatment of titanium with divalent cation solutions and its effect in promoting the adhesion of gingival epithelial cells and fibroblasts in vitro. Gingival keratinocyte-like Sa3 cells or fibroblastic NIH3T3 cells were cultured for 1 h on experimental titanium plates hydrothermally-treated with CaCl(2) (Ca) or MgCl(2) (Mg) solution, or distilled water (DW). The number and adhesive strengths of attached cells on the substrata were then analyzed. The number of Sa3 cells adhering to the Ca- and Mg-treated plates was significantly larger than in the DW group, but the strength of this adhesion did not differ significantly between groups. In contrast, NIH3T3 cell adhesion number and strength were increased in both the Ca and Mg groups compared to the DW group. Fluorescent microscopic observation indicated that, in all groups, Sa3 had identical expression levels of integrin β4 and development of actin filaments, whereas NIH3T3 cells in the Ca and Mg groups displayed much stronger punctate cytoplasmic signals for vinculin and more bundle-shaped actin filaments than cells in the DW group. As a result, it was indicated that the hydrothermal treatment of titanium with Ca or Mg solution improved the integration of soft tissue cells with the substrata, which may facilitate the development of a soft tissue barrier around the implant..
13. Ikiru Atsuta, Yasunori Ayukawa, Takayoshi Yamaza, Akihiro Furuhashi, Kiyoshi Koyano, The role of phosphoinositide 3-kinase in adhesion of oral epithelial cells to titanium., Archives of oral biology, 10.1016/j.archoralbio.2013.07.013, 58, 11, 1696-708, 2013.11, BACKGROUND: Oral epithelial cells (OECs) adhesion to titanium may improve the success rate of implant restoration. PURPOSE: We investigated the mechanism by which OECs adhere to titanium dental implants. MATERIALS AND METHODS: (1) After culturing rat OECs on titanium plates (Ti) or culture dishes in the presence or absence of a phosphoinositide 3-kinase (PI3K) activator or inhibitors and/or growth factors, and OEC morphology under these conditions were analyzed. (2) Right maxillary first molars were extracted and replaced with experimental implants. The rats were treated with or without growth factors. RESULTS: (1) Cell adherence was lower of OECs on Ti than in those on culture dishes, as were the levels of integrin β4 and the continuity of F-actin structures. After PI3K inhibition, markedly reducing adherence to both substrates. In contrast, PI3K activation with activator or insulin-like growth factor restored the OEC adherence and the expression of adhesion molecules on Ti to the levels seen in OECs cultured on dishes. Cell migration was inhibited by PI3K activation. (2) High expression of integrin β4 was observed in the peri-implant epithelia of PI3K-activated rats. CONCLUSION: These findings suggest that PI3K plays an important role in the adhesion of OECs to Ti..
14. Ikiru Atsuta, Yasunori Ayukawa, Akihiro Furuhashi, Takayoshi Yamaza, Yoshihiro Tsukiyama, Kiyoshi Koyano, Promotive effect of insulin-like growth factor-1 for epithelial sealing to titanium implants., Journal of biomedical materials research. Part A, 10.1002/jbm.a.34608, 101, 10, 2896-904, 2013.10, Improvement of oral epithelial adhesion to titanium (Ti) may significantly enhance the efficacy of dental implants. Here, we investigated whether insulin-like growth factor-1 (IGF-1) improved the sealing of the peri-implant epithelium (PIE) around the implant. Right maxillary first molars were extracted from rats and replaced with experimental implants. After 4 weeks of IGF-1 treatment, the implant-PIE interface exhibited a band of immunoreactive laminin-332 (Ln-5), similar to the tooth-junctional epithelium interface, that was partially absent in the untreated group. Immunoelectron microscopy showed a characteristic Ln-5-positive band including hemidesmosomes at both the apical and upper portions of the implant-PIE interface in the IGF-1-treated group. We also investigated the effects of IGF-1/PI3K inhibitors on the dynamics of rat oral epithelial cells (OECs) grown on Ti plates. In OECs cultured with IGF-1, adhesion protein expression increased, cell adherence to Ti plates was higher, and proliferation was faster, whereas migration and apoptosis were induced in the absence of IGF-1 or in the presence of both IGF-1 and a PI3K inhibitor. These data suggest that PI3K mediates the promotive effects of IGF-1, and that IGF-1 is effective at enhancing epithelial integration around Ti implants..
15. Qunzhou Zhang, Ikiru Atsuta, Shiyu Liu, Chider Chen, Shihong Shi, Songtao Shi, Anh D Le, IL-17-mediated M1/M2 macrophage alteration contributes to pathogenesis of bisphosphonate-related osteonecrosis of the jaws., Clinical cancer research : an official journal of the American Association for Cancer Research, 10.1158/1078-0432.CCR-13-0042, 19, 12, 3176-88, 2013.06, PURPOSE: Osteonecrosis of the jaw (ONJ) is emerging as one of the important complications in cancer patients treated with antiresorptive agents. This study explored the potential role of interleukin (IL)-17-mediated M1/M2 macrophage alterations in the pathogenesis of bisphosphonate-related osteonecrosis of the jaw (BRONJ). EXPERIMENTAL DESIGN: The expression of IL-17 and M1 and M2 macrophage markers at the local mucosal site of human BRONJ lesions was examined by immunofluorescence studies. BRONJ-like disease was induced in C57BL/6 mice and multiple myeloma-burdened mice by intravenous injection of zoledronate to evaluate the correlation of elevated IL-17 levels with changes in M1 and M2 macrophage phenotypes and the therapeutic effects of blocking IL-17 on pathogenesis of BRONJ-like disease. RESULTS: Increased T-helper (TH)17 cells and IL-17 cytokine correlate with an increase in M1/M2 macrophages ratio at the local mucosal site of both murine and human BRONJ lesion. Convincingly, in mice burdened with multiple myeloma, a combination of elevated suprabasal level and drug-induced IL-17 activity augmented the incidence of BRONJ; both systemic increase of IL-17 and disease severity could be reversed by adoptive transfer of ex vivo expanded M2 macrophages. Targeting IL-17 via specific neutralizing antibodies or a small inhibitory molecule, laquinimod, significantly decreased M1/M2 ratio and concomitantly suppressed BRONJ-like condition in mice. Mechanistically, IL-17 enhanced IFN-γ-induced M1 polarization through augmenting STAT-1 phosphorylation while suppressing IL-4-mediated M2 conversion via inhibiting STAT-6 activation. CONCLUSIONS: These findings have established a compelling linkage between activated IL-17-mediated polarization of M1 macrophages and the development of BRONJ-like conditions in both human disease and murine models..
16. Ruili Yang, Yi Liu, Peyman Kelk, Cunye Qu, Kentaro Akiyama, Chider Chen, Ikiru Atsuta, WanJun Chen, Yanheng Zhou, Songtao Shi, A subset of IL-17(+) mesenchymal stem cells possesses anti-Candida albicans effect., Cell research, 10.1038/cr.2012.179, 23, 1, 107-21, 2013.01, Bone marrow mesenchymal stem cells (MSCs) comprise a heterogeneous population of postnatal progenitor cells with profound immunomodulatory properties, such as upregulation of Foxp3(+) regulatory T cells (Tregs) and downregulation of Th17 cells. However, it is unknown whether different MSC subpopulations possess the same range of immunomodulatory function. Here, we show that a subset of single colony-derived MSCs producing IL-17 is different from bulk MSC population in that it cannot upregulate Tregs, downregulate Th17 cells, or ameliorate disease phenotypes in a colitis mouse model. Mechanistically, we reveal that IL-17, produced by these MSCs, activates the NFκB pathway to downregulate TGF-β production in MSCs, resulting in abolishment of MSC-based immunomodulation. Furthermore, we show that NFκB is able to directly bind to TGF-β promoter region to regulate TGF-β expression in MSCs. Moreover, these IL-17(+) MSCs possess anti-Candida albicans growth effects in vitro and therapeutic effect in C. albicans-infected mice. In summary, this study shows that MSCs contain an IL-17(+) subset capable of inhibiting C. albicans growth, but attenuating MSC-based immunosuppression via NFκB-mediated downregulation of TGF-β..
17. Ikiru Atsuta, Yasunori Ayukawa, Akihiro Furuhashi, Yoichiro Ogino, Yasuko Moriyama, Yoshihiro Tsukiyama, Kiyoshi Koyano, In vivo and in vitro studies of epithelial cell behavior around titanium implants with machined and rough surfaces., Clinical implant dentistry and related research, 10.1111/cid.12043, 16, 5, 772-81, 2014.10, BACKGROUND: The surface roughness of a dental implant affects the epithelial wound healing process and may significantly enhance implant prognosis. PURPOSE: We explored the influence of surface roughness on peri-implant epithelium (PIE) sealing and down-growth by comparing machine-surfaced (Ms) and rough-surfaced (Rs) implants. MATERIALS AND METHODS: (1) Maxillary first molars were extracted from rats and replaced with Ms or Rs implants. (2) We also compared changes in the morphology of cultured rat oral epithelial cells (OECs) grown on Ms or Rs titanium (Ti) plates. RESULTS: (1) After 4 weeks, the PIE around Ms and Rs implants showed a similar structure to junctional epithelium (JE). At 16 weeks, Rs implants appeared to form a weak epithelial seal at the tissue-implant interface and exhibited markedly less PIE down-growth than Ms implants but was deeper than that observed in natural teeth. (2) We observed less expression of adhesion proteins in OECs cultured on Rs plates than in cells grown on Ms plates. Additionally, cell adherence, migration, and proliferation on Rs plates were lower, whereas apoptosis was reduced on Ms plates. CONCLUSION: Ms implants are a better choice for integration with an epithelial wound healing process..
18. Yunia Dwi Rakhmatia, Yasunori Ayukawa, Ikiru Atsuta, Akihiro Furuhashi, Kiyoshi Koyano, Fibroblast attachment onto novel titanium mesh membranes for guided bone regeneration., Odontology, 10.1007/s10266-014-0151-8, 103, 2, 218-26, 2015.05, Titanium mesh is used in orthopedic surgery as a barrier membrane, as it offers suitable characteristics, which allow mechanical support during the formation of new bone. An ideal membrane would facilitate cell attachment onto its surface, thereby helping to stabilize the blood clot and integrate the membrane into the tissue. However, currently available titanium mesh has millimeter-level pore sizes, which lead to soft tissue ingrowth through the pores. Therefore, the aim of this study was to investigate the fibroblast attachment and migration on different designs of novel titanium mesh with micrometer pore size for guided bone regeneration treatment. Six types of novel titanium mesh membrane and three groups of commercially available membranes were used in this study. Fibroblasts were isolated from 4-day-old green fluorescence protein rats and seeded onto membrane surfaces. At 24 h, the cells attached to the membrane surfaces were fixed and stained with DAPI. The blue-stained nuclei on membrane surfaces, and both upper and lower sides were counted. It was shown that different membrane materials, structure and design differ considerably in their capacity for cell attachment to the membrane surface. The novel membranes, especially mesh with 12 pores compared with mesh with multi-pores, allowed the fibroblast attachment on the membrane surface, but hindered the fibroblast migration through the pores into the lower side of the membrane, which is associated with the defect area in the clinical condition..
19. Wakana Oshiro, Yasunori Ayukawa, Ikiru Atsuta, Akihiro Furuhashi, Jyunichi Yamazoe, Ryosuke Kondo, Mami Sakaguchi, Yuri Matsuura, Yoshihiro Tsukiyama, Kiyoshi Koyano, Effects of CaCl2 hydrothermal treatment of titanium implant surfaces on early epithelial sealing., Colloids and surfaces. B, Biointerfaces, 10.1016/j.colsurfb.2015.04.025, 131, 141-7, 2015.07, Improvement of oral epithelial adhesion to titanium (Ti) may significantly enhance the efficacy of dental implants. We aimed to investigate whether calcium chloride (CaCl2) hydrothermally treated (HT) Ti could promote sealing of the peri-implant epithelium (PIE) around the implant. Right maxillary first molars were extracted from rats and replaced with either CaCl2-HT implants (Ca-HT group), distilled water-HT implants (DW-HT group), or untreated implants (Cont group). After 4 weeks, the implant-PIE interface of the Ca-HT group exhibited a band of immunoreactive laminin-332, similar to the tooth-junctional epithelium interface, which was absent in the Cont and DW-HT groups at the upper portion. We also investigated the effect of Ca-HT on the attachment of rat oral epithelial cells (OECs). OEC adherence onto Ca-HT Ti plates was stronger with higher expression levels of adhesion proteins compared with Cont and DW-HT groups. These results indicate that HT with CaCl2 improves the integration of soft tissue cells with the Ti implant at 4 weeks after implantation, which might facilitate the development of a soft tissue barrier around the implant..
20. Hideaki Tanaka, Takeshi Toyoshima, Ikiru Atsuta, Yasunori Ayukawa, Masanori Sasaki, Yasuyuki Matsushita, Ryu Hiraoka, Kiyoshi Koyano, Seiji Nakamura, Additional Effects of Platelet-Rich Fibrin on Bone Regeneration in Sinus Augmentation With Deproteinized Bovine Bone Mineral: Preliminary Results., Implant dentistry, 10.1097/ID.0000000000000306, 24, 6, 669-74, 2015.12, PURPOSE: Platelet-rich fibrin (PRF) has been reported to contribute to bone regeneration; however, little is known about details with deproteinized bovine bone mineral (DBBM). This study aims to evaluate additional effects of PRF on bone regeneration in sinus augmentation with DBBM. MATERIALS AND METHODS: Sinus augmentations were made with DBBM/PRF mixture as lateral approach for 4 sinuses from 4 patients. Bone biopsies were obtained from posterior maxilla at the implant placements 7 or 10 months after sinus augmentations. Histological observations and histomorphometric analyses from augmented areas were performed. RESULTS: The new bone formation was found around the DBBMs with very good contact while surfaces of DBBMs were partly resorbed. Osteoclasts recognized the DBBMs for remodeling, followed by new bone running. The histomorphometric analyses revealed that mean percentages of newly formed bone were 31.7 ± 1.2%, 21.0 ± 1.0%, 38.0 ± 0.6%, and 47.0 ± 0.6%, respectively (mean 34.5 ± 5.7%). CONCLUSIONS: Additional effects of PRF could be found because of higher percentages of newly bone formation by DBBM/PRF mixture than those by DBBM individual in previous reports..
21. Noriyuki Yasunami, Yasunori Ayukawa, Akihiro Furuhashi, Ikiru Atsuta, Yunia Dwi Rakhmatia, Yasuko Moriyama, Tomohiro Masuzaki, Kiyoshi Koyano, Acceleration of hard and soft tissue healing in the oral cavity by a single transmucosal injection of fluvastatin-impregnated poly (lactic-co-glycolic acid) microspheres. An in vitro and rodent in vivo study., Biomedical materials (Bristol, England), 10.1088/1748-6041/11/1/015001, 11, 1, 015001-015001, 2015.12, Antihyperlipidemic drug statins reportedly promote both bone formation and soft tissue healing. We examined the effect of sustained-release, fluvastatin-impregnated poly(lactic-co-glycolic acid) (PLGA) microspheres on the promotion of bone and gingival healing at an extraction socket in vivo, and the effect of fluvastatin on epithelial cells and fibroblasts in vitro. The maxillary right first molar was extracted in rats, then one of the following was immediately injected, as a single dose, into the gingivobuccal fold: control (no administration), PLGA microspheres without a statin (active control), or PLGA microspheres containing 20 or 40 μg kg(-1) of fluvastatin. At days 1, 3, 7, 14, and 28 after injection, bone and soft tissue healing were histologically evaluated. Cell proliferation was measured under the effect of fluvastatin at dosages of 0, 0.01, 0.1, 1.0, 10, and 50 μM. Cell migration and morphology were observed at dosages of 0 and 0.1 μM. Following tooth extraction, the statin significantly enhanced bone volume and density, connective tissue volume, and epithelial wound healing. In the in vitro study, it promoted significant proliferation and migration of epithelial cells and fibroblasts. A single dose of topically administered fluvastatin-impregnated PLGA microspheres promoted bone and soft tissue healing at the extraction site..
22. Hideo Shimizu, Yohei Jinno, Yasunori Ayukawa, Ikiru Atsuta, Takaaki Arahira, Mitsugu Todo, Kiyoshi Koyano, Tissue Reaction to a Novel Bone Substitute Material Fabricated With Biodegradable Polymer-Calcium Phosphate Nanoparticle Composite., Implant dentistry, 10.1097/ID.0000000000000447, 25, 5, 567-74, 2016.10, PURPOSE: The aim of this study was to evaluate the effectiveness of a novel bone substitute material fabricated using a biodegradable polymer-calcium phosphate nanoparticle composite. METHODS: Porous structured poly-L-lactic acid (PLLA) and hydroxyapatite (HA) nanoparticle composite, which was fabricated using solid-liquid phase separation and freeze-drying methods, was grafted into bone defects created in rat calvarium or tibia. Rats were killed 4 weeks after surgery, and histological analyses were performed to evaluate new bone formation. RESULTS: Scanning electron microscopic observation showed the interconnecting pores within the material and the pore diameter was approximately 100 to 300 μm. HA nanoparticles were observed to be embedded into the PLLA beams. In the calvarial implantation model, abundant blood vessels and fibroblastic cells were observed penetrating into pores, and in the tibia model, newly formed bone was present around and within the composite. CONCLUSIONS: The PLLA-HA nanoparticle composite bone substitute developed in this study showed biocompatibility, elasticity, and operability and thus has potential as a novel bone substitute..
23. Yuri Matsuura, Ikiru Atsuta, Yasunori Ayukawa, Takayoshi Yamaza, Ryosuke Kondo, Akira Takahashi, Nobuyuki Ueda, Wakana Oshiro, Yoshihiro Tsukiyama, Kiyoshi Koyano, Therapeutic interactions between mesenchymal stem cells for healing medication-related osteonecrosis of the jaw., Stem cell research & therapy, 10.1186/s13287-016-0367-3, 7, 1, 119-119, 2016.08, BACKGROUND: Mesenchymal stem cells (MSCs) have been isolated from a variety of tissues, including bone marrow, adipose, and mucosa. MSCs have the capacity for self-renewal and differentiation. Reports have been published on the systemic administration of MSCs leading to functional improvements by engraftment and differentiation, thus providing a new strategy to regenerate damaged tissues. Recently, it has become clear that MSCs possess immunomodulatory properties and can therefore be used to treat diseases. However, the therapeutic effect mechanisms of MSCs are yet to be determined. Here, we investigated these mechanisms using a medication-related osteonecrosis of the jaw (MRONJ)-like mouse model. METHODS: To generate MRONJ-like characteristics, mice received intravenous zoledronate and dexamethasone two times a week. At 1 week after intravenous injection, maxillary first molars were extracted, and at 1 week after tooth extraction, MSCs were isolated from the bone marrow of the mice femurs and tibias. To compare "diseased MSCs" from MRONJ-like mice (d-MSCs) with "control MSCs" from untreated mice (c-MSCs), the isolated MSCs were analyzed by differentiation and colony-forming unit-fibroblast (CFU-F) assays and systemic transplantation of either d-MSCs or c-MSCs into MRONJ-like mice. Furthermore, we observed the exchange of cell contents among d-MSCs and c-MSCs during coculture with all combinations of each MSC type. RESULTS: d-MSCs were inferior to c-MSCs in differentiation and CFU-F assays. Moreover, the d-MSC-treated group did not show earlier healing in MRONJ-like mice. In cocultures with any combination, MSC pairs formed cell-cell contacts and exchanged cell contents. Interestingly, the exchange among c-MSCs and d-MSCs was more frequently observed than other pairs, and d-MSCs were distinguishable from c-MSCs. CONCLUSIONS: The interaction of c-MSCs and d-MSCs, including exchange of cell contents, contributes to the treatment potential of d-MSCs. This cellular behavior might be one therapeutic mechanism used by MSCs for MRONJ..
24. Ikiru Atsuta, Yasunori Ayukawa, Ryosuke Kondo, Wakana Oshiro, Yuri Matsuura, Akihiro Furuhashi, Yoshihiro Tsukiyama, Kiyoshi Koyano, Soft tissue sealing around dental implants based on histological interpretation., Journal of prosthodontic research, 10.1016/j.jpor.2015.07.001, 60, 1, 3-11, 2016.01, PURPOSE: The aim of this study was to provide an overview on the biology and soft tissue sealing around dental implants and teeth. STUDY SELECTION: This is a narrative review performed through scientific articles published between 1977 and 2014, indexed in MEDLINE and PubMed databases. The study selected articles that focused on epithelial sealing around dental implant or teeth with cell biology and histology of soft tissue. RESULTS: Implant therapy has been widely applied in dental rehabilitation for many years, with predictable long-term results. The longevity and functionality of dental implants is dependent on both osseointegration around the implant body and the establishment of a soft tissue barrier that protects the underlying hard tissue structures and the implant itself. The health and stability of the peri-implant mucosa also affects the esthetics of the implant. The healing and maintenance of the epithelial and connective tissues around implants are increasingly recognized as being fundamental to implant success. However, there has been little research into the function or formation of the soft tissue seal around dental implants, and the roles of this unique mucosal interface remain unclear. CONCLUSIONS: This narrative review explores the extent of the current knowledge of soft tissue barriers around implants from both a basic and clinical perspective, and aims to consolidate this knowledge and highlight the most pertinent questions relating to this area of research..
25. Shinichiro Kuroshima, Masaru Kaku, Takashi Matsuura, Ikiru Atsuta, Yasunori Ayukawa, Takashi Sawase, Medication-related osteonecrosis of the jaw; what should we do as prosthodontists?, Journal of prosthodontic research, 10.1016/j.jpor.2016.01.004, 60, 4, 229-230, 2016.10.
26. Yuzo Takamori, Ikiru Atsuta, Hirotaka Nakamura, Takashi Sawase, Kiyoshi Koyano, Yoshitaka Hara, Histopathological comparison of the onset of peri-implantitis and periodontitis in rats., Clinical oral implants research, 10.1111/clr.12777, 28, 2, 163-170, 2017.02, BACKGROUND AND OBJECTIVE: There are a few experimental models that clearly describe the pathological differences in tissue destruction between periodontitis and peri-implantitis. We recently reported that the formation of immune complexes accelerates site-specific loss of attachment and alveolar bone resorption when an antigen is topically applied in the gingival sulcus of an immunized rat. We applied this model to the peri-implant tissues and compared peri-implant destruction to periodontitis without using a ligature. MATERIAL AND METHODS: Twenty-five rats were used in this study and were divided into five groups. Implantation was performed immediately after extraction of right first molars in rats. The left first molars were left untreated to be examined as natural teeth. The immunized group consisted of rats that had received intraperitoneal lipopolysaccharide (LPS), whereas the nonimmunized group received only phosphate-buffered saline (PBS). The untreated baseline group received only implantation. After intraperitoneal booster injection, half of each group received topical application of LPS in the palatal gingival sulcus daily for 3 days. The other half of the groups received PBS. Histopathological and histometrical findings were observed with hematoxylin and eosin staining, collagen fibers were observed with Azan staining, and formation of immune complexes was immunohistologically evaluated by C1qB expression. RESULT: Peri-implant tissue destruction was greater in the immunized and LPS-applied groups than in the other groups. No periodontal destruction was observed. Formation of immune complexes was observed in the junctional epithelium and adjacent connective tissue in the immunized groups. CONCLUSION: Antigen-induced peri-implant tissue destruction occurs faster than periodontal tissue destruction..
27. Miya Kanazawa, Ikiru Atsuta, Yasunori Ayukawa, Takayoshi Yamaza, Ryosuke Kondo, Yuri Matsuura, Kiyoshi Koyano, The influence of systemically or locally administered mesenchymal stem cells on tissue repair in a rat oral implantation model., International journal of implant dentistry, 10.1186/s40729-017-0112-4, 4, 1, 2-2, 2018.01, BACKGROUND: Multipotent mesenchymal stem cells (MSCs) are used clinically in regenerative medicine. Our previous report showed systemically injected MSCs improved peri-implant sealing and accelerated tissue healing. However, the risks of systemic MSC administration, including lung embolism, must be considered; therefore, their local application must be assessed for clinical safety and efficacy. We investigated differences in treatment effect between local and systemic MSC application using a rat oral implantation model. METHODS: Rat bone marrow-derived MSCs were isolated and culture-expanded. The rat's right maxillary first molars were extracted and replaced with experimental titanium implants. After 24 h, MSCs (1 × 106/ml) were systemically or locally injected into recipient rats via the tail vein (systemic group) or buccal subcutaneous tissue (local group), respectively. Rats treated in the absence of MSCs were included as a control (control group). The maxillary epithelium was assessed histologically after 4 weeks to evaluate laminin-332 (Ln-332) distribution and horseradish peroxidase invasion, as indicators of peri-implant epithelium (PIE) formation and PIE sealing to the implant surface, respectively. The effect of MSCs on rat oral epithelial cell (OEC) morphology was determined by coculture. RESULTS: Systemic group MSCs accumulated early at the peri-implant mucosa, while local group MSCs were observed in various organs prior to later accumulation around the implant surface. PIE formation and Ln-332-positive staining at the implant interface were enhanced in the systemic group compared with the local and control groups. Furthermore, OEC adherence on implants was reduced in high-density compared with low-density MSC cocultures. CONCLUSIONS: Local MSC injection was more ineffective than systemic MSC injection at enhancing PIE sealing around titanium implants. Thus, although local MSC administration has a wide range of applications, further investigations are needed to understand the exact cellular and molecular mechanisms of this approach prior to clinical use..
28. Maiko Mieda, Ikiru Atsuta, Yasuyuki Matsushita, Takehiro Morita, Yasunori Ayukawa, Yoshihiro Tsukiyama, Yoshinori Sawae, Kiyoshi Koyano, The effective design of zirconia coping on titanium base in dental implant superstructure., Dental materials journal, 10.4012/dmj.2017-022, 37, 2, 237-243, 2018.03, Zirconia exhibits good tissue compatibility and nontoxicity, making it a widely used esthetic replacement material for implant abutments. To avoid abutment-fracture, the parts composed of zirconia with a bonded metal component connected to the implant can be used. The purpose of this study was to design titanium and zirconia components with high fracture resistance at the zirconia component's edge line. Three edge line designs of the titanium base and zirconia sleeve were made: chamfer, shoulder, and back-taper. To assess the strength of the abutment design, static loads were applied vertically and 30 degrees from the vertical axis. A test of tensile strength was also performed after chewing simulation. Conventional zirconia components mounted on a chamfer-type titanium base showed significantly lower fracture resistance than shoulder and back-taper types. This study suggests that to improve the durability of zirconia abutments with a titanium base, a back-tapered edge design is recommended..
29. Sonoda S, Mei YF, Atsuta I, Danjo A, Yamaza H, Hama S, Nishida K, Tang R, Kyumoto-Nakamura Y, Uehara N, Kukita T, Nishimura F, Yamaza T, Exogenous nitric oxide stimulates the odontogenic differentiation of rat dental pulp stem cells., Scientific reports, 10.1038/s41598-018-21183-6, 8, 1, 3419-3419, 2018.02.
30. Yasunori Ayukawa, Wakana Oshiro, Ikiru Atsuta, Akihiro Furuhashi, Ryosuke Kondo, Yohei Jinno, Kiyoshi Koyano, Long Term Retention of Gingival Sealing around Titanium Implants with CaCl2 Hydrothermal Treatment: A Rodent Study., Journal of clinical medicine, 10.3390/jcm8101560, 8, 10, 2019.09, We previously reported that CaCl2 hydrothermal-treated (Ca-HT) titanium (Ti) implants induced a tight sealing at the interface between the implant and peri-implant epithelium (PIE) after implantation. However, it is not clear how long this improved epithelium sealing can be maintained. We subsequently investigated whether the positive effect of Ca-HT to promote sealing between the PIE and implant was sustained longer term. Maxillary molars were extracted from rats and replaced with either Ca-HT implants (Ca-HT group), distilled water-HT implants (DW-HT group) or non-treated implants (control group). After 16 weeks, the majority of implants in the Ca-HT group remained at the maxillary with no apical extension of the PIE. Conversely, half the number of control implants was lost following down-growth of the PIE. The effect of Ca-HT on migration and proliferation of rat oral epithelial cells (OECs) was also investigated. In OECs cultured on Ca-HT Ti plates, protein expression in relation to cell migration decreased, and proliferation was higher than other groups. Surface analysis indicated HT enhanced the formation of surface TiO2 layer without altering surface topography. Consequently, Ca-HT of Ti reduced PIE down-growth via tight epithelial attachment to the surface, which may enhance implant capability for a longer time post-implantation..
31. Ikiru Atsuta, Yasunori Ayukawa, Akihiro Furuhashi, Ikue Narimatsu, Ryosuke Kondo, Wakana Oshiro, Kiyoshi Koyano, Epithelial sealing effectiveness against titanium or zirconia implants surface., Journal of biomedical materials research. Part A, 10.1002/jbm.a.36651, 107, 7, 1379-1385, 2019.07, The aims of implant treatment now involve not only restoration of mastication function, but also recovery of esthetics. Currently, zirconia is widely used as an esthetic material for implant abutment. Therefore, it is very important to understand the efficacy of zirconia for epithelial sealing as an implant material. We compared the effects of materials on the sealing of the peri-implant epithelium (PIE) to titanium (Ti) or zirconia (Zr) implants, for application to clinical work. Maxillary first molars were extracted from rats and replaced with Ti or Zr implants. The sealing of the PIE to the implants was evaluated with immunohistochemistry observation and HRP analysis. The morphological and functional changes in rat oral epithelial cells (OECs) cultured on Ti or Zr plates were also evaluated. After 4 weeks, the PIE on the Ti and Zr implants showed similar structures. The Zr implants appeared to form a weak epithelial seal at the tissue-implant interface, and exhibited markedly less adhesive structures than the Ti implants under electron microscopic observation. In the in vitro experiments, decreased expression levels of adhesion proteins were observed in OECs cultured on Zr plates compared with those cultured on Ti plates. In addition, the cell adherence on Zr plates was reduced, while the cell migration was low on Ti plates. Zr is a better choice for an esthetic implant material, but needs further improvement for integration with the epithelial wound healing process around a dental implant. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2019..
32. Ikue Narimatsu, Ikiru Atsuta, Yasunori Ayukawa, Wakana Oshiro, Noriyuki Yasunami, Akihiro Furuhashi, Kiyoshi Koyano, Epithelial and Connective Tissue Sealing around Titanium Implants with Various Typical Surface Finishes., ACS biomaterials science & engineering, 10.1021/acsbiomaterials.9b00499, 5, 10, 4976-4984, 2019.10, Soft tissue barrier around a dental implant plays a crucial role in the success of dental implants because it protects underlying hard tissue structures. A number of surface alteration procedures of implants have been introduced to improve bone-implant contact, but there has been little research on the peri-implant soft tissue (PIS) seal. The present study focuses on the "biologic width" of epithelial and connective tissue seals around implants with various typical surface finishes by testing surfaces that have been machined (Ms), roughened by sandblasting and acid etching (Rs), treated hydrothermally with CaCl2 (Cs), or anodized (As). Ms, Rs, and As techniques are commonly used to finish surfaces of commercially available dental implants. The Cs technique was reported to produce strong epithelial cell-titanium adhesion. For culture study, rat oral epithelial cells (OECs) and fibroblasts were cultured on Ms, Rs, Cs, and As titanium plates. There was less cell adherence of OECs and more collagen expression when cultured on Rs and As plates than when cultured on Ms and Cs plates. For the in vivo study, implants with Ms, Rs, Cs, and As surfaces were placed in the rats' oral cavity. Although the PIS structure was similar to that around natural teeth, a horseradish peroxide assay revealed that the sealing ability around the Ms and Rs implants was weaker than that around Cs implants. After 16 weeks, Rs implants exhibited peri-implant epithelial apical down-growth and had lost bone support. Thus, although a smooth surface (Ms and Cs) showed better epithelial attachment, rough surfaces (Rs and As) are more suitable for binding to the connective tissue. Strong epithelium-implant attachment seems to be a fundamental defense against foreign body penetration. Selecting suitable surfaces to ensure strong sealing is important for implant success..
33. Mikio Imai, Yasunori Ayukawa, Noriyuki Yasunami, Akihiro Furuhashi, Yoko Takemura, Naomi Adachi, Jiangqi Hu, Xudiyang Zhou, Yasuko Moriyama, Ikiru Atsuta, Kosaku Kurata, Kiyoshi Koyano, Effect of a Single Injection of Benidipine-Impregnated Biodegradable Microcarriers on Bone and Gingival Healing at the Tooth Extraction Socket., Advances in wound care, 10.1089/wound.2018.0834, 8, 3, 108-117, 2019.03, Objective: A dihydropyridine-type calcium channel blocker, benidipine (BD), is extensively used in hypertension therapy. In vitro study reported BD promoting bone metabolism. We evaluated the effect of sustained release of BD-loaded poly(lactic-co-glycolic acid) (PLGA) microcarriers on the promotion of bone and gingival healing at an extraction socket in vivo. In addition, the effect of BD on osteoblasts, osteocytes, fibroblasts, and epithelial cells was evaluated in vitro. Approach: The maxillary first molar of rats was extracted. Next, PLGA microcarriers containing BD were directly injected into the gingivobuccal fold as a single dose. After injection, bone and soft-tissue healing was histologically evaluated. Effect of BD on proliferation, migration, and gene expression of gingival and bone cell was also examined in vitro. Results: After tooth extraction, BD significantly augmented bone volume and density, and also epithelial wound healing. During in vitro studies, BD promoted significant proliferation and migration of fibroblasts and epithelial cells. Real-time RT-PCR revealed that BD upregulated messenger RNA expression of Ahsg (alpha 2-HS glycoprotein) and Csf2 (colony-stimulating factor 2) in osteoblasts. Innovation: The prevention of bone and soft-tissue reduction associated with tooth extraction has been eagerly anticipated in the field of dentistry. This study first reported the effect of BD on extraction socket healing. Conclusion: A single dose of topically administered BD-loaded PLGA microcarriers promoted bone and soft-tissue healing at the extraction site of tooth..
34. Yasushige Sakamoto, Yasunori Ayukawa, Akihiro Furuhashi, Michimasa Kamo, Junji Ikeda, Ikiru Atsuta, Takuya Haraguchi, Kiyoshi Koyano, Effect of Hydrothermal Treatment with Distilled Water on Titanium Alloy for Epithelial Cellular Attachment., Materials (Basel, Switzerland), 10.3390/ma12172748, 12, 17, 2019.08, The enhancement of oral epithelial adhesion to the trans-mucosal material of dental implants may improve their long-term stability. The aim of this study is to investigate whether hydrothermal treatment with distilled water (HT-DW) applied to a Ti-6Al-4V (Ti64) alloy could improve epithelial cellular attachment. We hypothesized that this treatment would enhance the adsorption of proteins and the adhesion of gingival epithelial GE1 cells. This treatment changed the surface crystal structure into an anatase type of titanium oxide without an apparent change of surface roughness or topography. Nitrogen was not detected on the HT-DW-treated Ti64, which indicates decontamination. HT-DW-treated Ti64 exhibited a hydrophilic surface with a less than 10° angle of water contact. Adsorption of laminin-332 to the HT-DW-treated Ti64 was significantly greater than that of the untreated Ti64 plates (64). The number of GE1 cells on the HT-DW-treated Ti64 at 1 and 3 days was significantly lower than that on 64; however, cell adhesion strength on HT-DW was greater, with a higher expression of integrin β4, compared with 64. This indicates that the HT-DW treatment of Ti64 improves the integration of GE1 cells, which might facilitate the development of a soft tissue barrier around the implant..
35. Jiangqi Hu, Ikiru Atsuta, Yasunori Ayukawa, Xudiyang Zhou, Yunia Dwi Rakhmatia, Kiyoshi Koyano, The impact of surface alteration on epithelial tissue attachment after the mechanical cleaning of titanium or zirconia surface., Journal of oral rehabilitation, 10.1111/joor.12920, 47, 9, 1065-1076, 2020.09, OBJECTIVE: Mechanical plaque removal may alter the surface morphology of the gingival penetration part of the implant. We applied an air-powered abrasive system (AP), titanium curette (TC), stainless curette (SC), ultrasound scaler (US), and titanium brush (TB) which are commonly used to remove plaque, to titanium or zirconia and the changes in surface morphology and the epithelial attach against substrata. MATERIALS AND METHODS: (a) The morphological changes of titanium and zirconia after mechanical cleaning were assessed by scanning electron microscopy and a roughness analyser. (b) Oral epithelial cells of rats were inoculated on the surface of the materials after mechanical cleaning, and the adherence of epithelial cells was observed. (c) The maxillary first molars were extracted from the rats and replaced by experimental titanium or zirconia implants. The length of the immunoreactive laminin-332 band was observed at the implant-peri-implant epithelium interface. RESULTS: (a) The surface roughness increased in experimental groups except the AP group. (b) Among the experimental groups, the AP group showed the highest number of attached cells. (c) The length of the immunoreactive laminin-332 band was longer in the control group than those in all five experimental groups. Among the experimental groups, the AP group showed the longest band. CONCLUSION: All mechanical cleaning methods increased the surface roughness of the materials except AP. AP did not cause distinct implant surface alterations. Surface alteration caused by mechanical cleaning may evoke inferior for epithelial attachment and reduce resistance against foreign infiltration..
36. Hiroki Suzuki, Yasunori Ayukawa, Yoko Ueno, Ikiru Atsuta, Akio Jinnouchi, Kiyoshi Koyano, Relationship between Maximum Tongue Pressure Value and Age, Occlusal Status, or Body Mass Index among the Community-Dwelling Elderly., Medicina (Kaunas, Lithuania), 10.3390/medicina56110623, 56, 11, 1-8, 2020.11, Background and objectives: In an aging society, the maintenance of the oral function of the elderly is of importance for the delay or prevention of frailty and long-term care. In the present study, we focused on the maximum tongue pressure (MTP) value and analyzed the relationship between MTP and age, occlusal status, or body mass index (BMI). Materials and Methods: This one-center observatory study was conducted using a cohort consisting of 205 community-dwelling outpatients over 65 years old. The MTP values of all subjects were measured using a commercially available tongue pressure measurement device and statistically analyzed. In addition, the correlation between MTP value and BMI was analyzed. Results: The MTP value decreased with age, especially in subjects classified as Eichner B and C. The difference in occlusal status did not show any statistically significant influence on MTP value. The correlation between BMI and MTP value was indicated in the tested groups other than an age of 65-74 and Eichner A groups. Conclusions: Although MTP value decreased with age, the difference in occlusal status did not have an impact on MTP value. The correlation between BMI and MTP value was not shown in the youngest group or a group with sufficient occlusal units. The results presented in the present study may imply that, even if MTP is low, younger age and/or better occlusal status compensate for the inferior MTP value in the cohort studied..
37. Naomi Adachi, Yasunori Ayukawa, Noriyuki Yasunami, Akihiro Furuhashi, Mikio Imai, Koma Sanda, Ikiru Atsuta, Kiyoshi Koyano, Preventive effect of fluvastatin on the development of medication-related osteonecrosis of the jaw., Scientific reports, 10.1038/s41598-020-61724-6, 10, 1, 5620-5620, 2020.03, Medication-related osteonecrosis of the jaw (MRONJ) occurs in patients undergoing oral surgery while medicated with bisphosphonate, denosumab or anti-angiogenic agents. We employed a MRONJ-like rat model to investigate whether injecting fluvastatin at extraction sites prevents MRONJ-like lesion. A MRONJ-like model was created by treating rats with zoledronate and dexamethasone, extracting teeth, and immediately injecting fluvastatin at the extraction site. The experimental group comprised three subgroups treated with low (0.1 mg/kg; FS-L), medium (1.0 mg/kg; FS-M) and high concentrations (10 mg/kg; FS-H) of fluvastatin. Necrotic bone exposure was significantly lower in the FS-M (p = 0.028) and FS-H (p = 0.041) groups than in the MRONJ group. The distance between the edges of the epithelial surfaces was significantly shorter in the FS-M (p = 0.042) and FS-H (p = 0.041) groups. The area of necrotic bone and the necrotic bone ratio were significantly smaller in the FS-H group (p = 0.041 and p = 0.042 respectively). Bone volume fraction calculated on μ-CT images was significantly larger in the FS-H group than in the MRONJ group (p = 0.021). Our findings suggest that a single local injection of fluvastatin following tooth extraction can potentially reduce the chance of developing MRONJ-like lesion in rats..
38. Yasunori Ayukawa, Ikiru Atsuta, Yasuko Moriyama, Yohei Jinno, Kiyoshi Koyano, Localization of Integrin Beta-4 Subunit at Soft Tissue-Titanium or Zirconia Interface., Journal of clinical medicine, 10.3390/jcm9103331, 9, 10, 1-7, 2020.10, Currently, along with titanium (Ti), zirconia is widely used as an abutment material for dental implants because it makes it possible to avoid gingival discoloration; however, the epithelial sealing capability of zirconia remains unknown. The purpose of the present study is to elucidate the localization of integrin β4 subunit (Inβ4), one of the main proteins in the attachment structure between gingival junctional epithelial (JE) cells and substrata. Maxillary first molars were extracted from rats, and implants were placed with Ti or zirconia transgingival parts; then, the localization of Inβ4 was observed. Morphological and functional changes in rat oral epithelial cells (OECs) cultured on a culture dish (Dish) and Ti and zirconia plates were also evaluated with Inβ4 immunofluorescence histochemistry and Western blotting. After four weeks of implant placement, the morphology of the peri-implant epithelium (PIE) and the localization of Inβ4 around the Ti and zirconia transgingival parts were similar. However, both exhibited markedly shorter Inβ4-positive bands in the PIE than in the JE around natural teeth. Decreased expression levels of Inβ4 were observed in OECs cultured on Ti and zirconia plates compared with those cultured on Dish. In conclusion, although inferior to natural teeth, zirconia implants are thought to have epithelial sealing properties comparable to those of titanium..
39. Xudiyang Zhou, Ikiru Atsuta, Yasunori Ayukawa, Ikue Narimatsu, Tianren Zhou, Jiangqi Hu, Kiyoshi Koyano, Effects of Different Divalent Cation Hydrothermal Treatments of Titanium Implant Surfaces for Epithelial Tissue Sealing., Materials (Basel, Switzerland), 10.3390/ma13092038, 13, 9, 2020.04, The improvement of peri-implant epithelium (PIE) adhesion to titanium (Ti) may promote Ti dental implant stability. This study aims to investigate whether there is a positive effect of Ti hydrothermally treated (HT) with calcium chloride (CaCl2), zinc chloride (ZnCl2), and strontium chloride (SrCl2) on promoting PIE sealing. We analyzed the response of a rat oral epithelial cell (OEC) culture and performed an in vivo study in which the maxillary right first molars of rats were extracted and replaced with calcium (Ca)-HT, zinc (Zn)-HT, strontium (Sr)-HT, or non-treated control (Cont) implants. The OEC adhesion on Ca-HT and Zn-HT Ti plates had a higher expression of adhesion proteins than cells on the Cont and Sr-HT Ti plates. Additionally, the implant PIE of the Ca-HT and Zn-HT groups revealed better expression of immunoreactive laminin-332 (Ln-322) at 2 weeks after implantation. The Ca-HT and Zn-HT groups also showed better attachment at the implant-PIE interface, which inhibited horseradish peroxidase penetration. These results demonstrated that the divalent cations of Ca (Ca2+) and Zn (Zn2+)-HT improve the integration of epithelium around the implant, which may facilitate the creation of a soft barrier around the implant to protect it from foreign body penetration..
40. Takuya Haraguchi, Yasunori Ayukawa, Yukie Shibata, Toru Takeshita, Ikiru Atsuta, Yoichiro Ogino, Noriyuki Yasunami, Yoshihisa Yamashita, Kiyoshi Koyano, Effect of Calcium Chloride Hydrothermal Treatment of Titanium on Protein, Cellular, and Bacterial Adhesion Properties., Journal of clinical medicine, 10.3390/jcm9082627, 9, 8, 1-12, 2020.08, Topographical modification of the dental implant surface is one of the main topics for the improvement of the material, however, the roughened surface has some risks for peri-implantitis. A hydrothermal treatment (HT) of titanium with calcium chloride solution was reported to improve osseointegration and soft tissue sealing without changing the surface topography; however, its mechanism is unclear. We herewith investigated the interaction between extracellular matrix (ECM) protein and HT titanium. Furthermore, we also clarified the bacterial interaction. We employed two kinds of HT, HT with water (DW-HT) and HT with calcium chloride solution (Ca-HT). As a result, the adsorptions of both laminin-332 and osteopontin onto the Ca-HT surface were enhanced. In contrast, the adsorption of albumin, which was reported to have no cell adhesion capacity, was not influenced by Ca-HT. Osteoblast adhesion onto Ca-HT was also enhanced. Although Ca-HT was reported to enhance both epithelial cell attachment strength and in vivo peri-implant epithelial bonding, the number of epithelial cell attachment was not increased even after HT. Ca-HT had no impact in the adhesion of Streptococcus gordonii. These results suggest that Ca-HT enhances cell adhesion onto titanium without increasing bacterial adhesion, and the improvement of ECM protein adsorption is supposed to contribute to cell adhesion..
41. Keishu Tachibana, Ikiru Atsuta, Yoshihiro Tsukiyama, Rika Kuwatsuru, Takehiro Morita, Hiroya Yoshimatsu, Yasuyuki Matsushita, Ikue Narimatsu, Yasunori Ayukawa, Yoshinori Sawae, Kiyoshi Koyano, The need for polishing and occlusal adjustment of zirconia prostheses for wear on antagonist teeth., Dental materials journal, 10.4012/dmj.2020-190, 40, 3, 650-656, 2021.05, The attrition of enamel when opposed by ceramics is of great concern. The purpose of this study was to evaluate enamel wear against high translucent zirconia (Zr), lithium disilicate (LD), gold (Au), and enamel (E) with different surface and contact conditions. The materials were divided into two groups: polished and ground (n=8 each). Two-body wear tests were performed against human enamel with vertical and horizontal, horizontal, and vertical repetitive movements as experiments 1 to 3 respectively. The surface roughness of all materials except Zr changed throughout the experiments. In experiment 1, Zr and Au showed less antagonist wear when polished than when ground. In experiment 2, polished groups showed less antagonist wear than ground groups in all materials. In experiment 3, Zr and LD exerted greater antagonist wear than E, regardless of Ra. These findings confirm the importance of polishing and occlusal adjustment of zirconia..
42. Akihiro Furuhashi, Yasunori Ayukawa, Ikiru Atsuta, Yunia Dwi Rakhmatia, Kiyoshi Koyano, Soft Tissue Interface with Various Kinds of Implant Abutment Materials., Journal of clinical medicine, 10.3390/jcm10112386, 10, 11, 2021.05, Various materials, such as titanium, zirconia and platinum-gold (Pt-Au) alloy, have been utilized for dental implant trans-mucosal parts. However, biological understanding of soft tissue reaction toward these materials is limited. The aim of this study was to compare the response of cell lines and soft tissue to titanium, zirconia and Pt-Au substrata. The surface hydroxyl groups and protein adsorption capacities of the substrata were measured. Next, gingival epithelial-like cells (Sa3) and fibroblastic cells (NIH3T3) were cultured on the materials, and initial cell attachment was measured. Immuno-fluorescent staining of cell adhesion molecules and cytoskeletal proteins was also performed. In the rat model, experimental implants constructed from various materials were inserted into the maxillary tooth extraction socket and the soft tissue was examined histologically and immunohistochemically. No significant differences among the materials were observed regarding the amount of surface hydroxyl groups and protein adsorption capacity. Significantly fewer cells of Sa3 and NIH3T3 adhered to the Pt-Au alloy compared to the other materials. The expression of cell adhesion molecules and a well-developed cytoskeleton was observed, both Sa3 and NIH3T3 on each material. In an animal model, soft tissue with supracrestal tissue attachment was observed around each material. Laminin-5 immuno-reactivity was seen in epithelia on both titanium and zirconia, but only in the bottom of epithelia on Pt-Au alloy. In conclusion, both titanium and zirconia, but not Pt-Au alloy, displayed excellent cell adhesion properties..
43. Xiaoxu Zhang, Ikiru Atsuta, Ikue Narimatsu, Nobuyuki Ueda, Ryosuke Takahashi, Yuki Egashira, Jing-Qi Zhang, Jiong-Yan Gu, Kiyoshi Koyano, Yasunori Ayukawa, Replacement Process of Carbonate Apatite by Alveolar Bone in a Rat Extraction Socket., Materials (Basel, Switzerland), 10.3390/ma14164457, 14, 16, 2021.08, The objective of this study was to investigate a bone graft substitute containing carbonate apatite (CO3Ap) to analyze bone replacement and the state of bone formation in vitro and in vivo compared with autogenous bone (AB) or control. An osteoclast precursor cell line was cultured with AB or CO3Ap, and morphological analysis using scanning electron microscopy and a tartrate-resistant acid phosphatase activity assay were performed. The right maxillary first and second molars of Wistar rats were extracted and compensated by AB or CO3Ap granules. Following implantation, the bone formation state was evaluated after 3, 5, 7, 14, and 28 days of surgery by micro-computed tomography and immunohistostaining. The osteoclast-like cell morphology was typical with many cell protrusions in the AB and CO3Ap groups. Additionally, the number of osteoclast-like cells formed in the culture increased in each group; however, there was no significant difference between the AB and CO3Ap groups. Five days after tooth extraction, osteoclasts were observed near CO3Ap. The bone thickness in the CO3Ap group was significantly increased than that in the control group and the bone formation in the CO3Ap group increased by the same level as that in the AB group. CO3Ap is gradually absorbed by osteoclasts in the extraction socket and is easily replaced by alveolar bone. The process of bone replacement by osteoclasts is similar to that of autologous bone. By observing the process of bone replacement in more detail, it may be possible to gain a better understanding of the bone formation and control the amount of bone after surgery..
44. Ikiru Atsuta, Ikue Narimatsu, Taichiro Morimoto, Chi-Hsiang Cheng, Kiyoshi Koyano, Yasunori Ayukawa, Assessment of the Soft-Tissue Seal at the Interface between the Base of the Fixed Denture Pontic and the Oral Mucosa., Materials (Basel, Switzerland), 10.3390/ma14143997, 14, 14, 2021.07, Fixed dentures (bridges) are often selected as a treatment option for a defective prosthesis. In this study, we assess the contact condition between the base of the pontic and oral mucosa, and examine the effect of prosthetic preparation and material biocompatibility. The molars were removed and replaced with experimental implants with a free-end type bridge superstructure after one week. In Experiment 1, we assessed different types of prosthetic pre-treatment: (1) the untreated control group (Con: mucosa recovering from the tooth extraction); (2) the laser irradiation group (Las: mucosa recovering after the damage caused by a CO2 laser); and (3) the tooth extraction group (Ext: mucosa recovering immediately after the teeth extraction). In Experiment 2, five materials (titanium, zirconia, porcelain, gold-platinum alloy, and self-curing resin) were placed at the base of the bridge pontic. Four weeks after the placement of the bridge, the mucosa adjacent to the pontic base was histologically analyzed. In Experiment 1, the Con and Las groups exhibited no formation of an epithelial sealing structure on the pontic base. In the Ext group, adherent epithelium was observed. In Experiment 2, the sealing properties at the pontic interface were superior for titanium and the zirconia compared with those made of porcelain or gold-platinum alloy. In the resin group, a clear delay in epithelial healing was observed..
45. Sonoda S, Mei YF, Atsuta I, Danjo A, Yamaza H, Hama S, Nishida K, Tang R, Kyumoto-Nakamura Y, Uehara N, Kukita T, Nishimura F, Yamaza T., Exogenous nitric oxide stimulates the odontogenic differentiation of rat dental pulp stem cells., 2018.06.
46. Kanazawa M, Atsuta I, Ayukawa Y, Yamaza T, Kondo R, Matsuura Y, Koyano K., The influence of systemically or locally administered mesenchymal stem cells on tissue repair in a rat oral implantation model., 2018.02.
47. 熱田 生, Soft tissue sealing around dental implants based on histological interpretation., J Prosthodont Res, 2016.01.
48. 熱田 生, Therapeutic interaction of systemically-administered mesenchymal stem cells with peri-implant mucosa., PLoS One, 2014.05.
49. ATSUTA IKIRU, IL-17-mediated M1/M2 macrophage alteration contributes to pathogenesis of bisphosphonate-related osteonecrosis of the jaws., Clin Cancer Res., 2013.05.
50. ATSUTA IKIRU, Mesenchymal stem cells inhibit multiple myeloma cells via the Fas-L/Fas pathway, Stem Cell Res Ther, 2013.04.
51. ATSUTA IKIRU, Yasunori AYUKAWA, kiyoshi koyano, The role of phosphoinositide 3-kinase in adhesion of oral epithelial cells to titanium, ARCHIVES OF ORAL BIOLOGY, 10.1016/j.archoralbio.2013.07.013, 58, 11, 1696-1708, 2013.11.
52. ATSUTA IKIRU, Evaluations of epithelial sealing and peri-implant epithelial down-growth around “step-type” implants, 2012.09.
53. ATSUTA IKIRU, Expression of Integrin beta-4 and alpha-3 subunits on the process of peri-implant epithelium formation, 2013.03.
54. ATSUTA IKIRU, In vivo and in vitro studies of epithelial cell behavior around titanium implants with machined- and rough-surface., 2013.01.
55. ATSUTA IKIRU, Promotive effect of insulin-like growth factor-1 for epithelial sealing to titanium implants, 2013.04.