Updated on 2024/05/26

写真a

 
YOSHIBA Nagako
 
Organization
Academic Assembly Institute of Medicine and Dentistry SHIGAKU KEIRETU Professor
Graduate School of Medical and Dental Sciences Oral Health and Welfare Science Oral Health and Welfare Professor
Title
Professor
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Degree

  • 博士(歯学) ( 1996.6   新潟大学 )

Research Areas

  • Life Science / Conservative dentistry

Research History (researchmap)

  • 新潟大学大学院医歯学総合研究科 口腔保健学分野 教授

    2024.4

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  • Niigata University   研究推進機構研究准教授

    2022.11 - 2024.3

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  • 2021年度拠点事業「大阪大学蛋白質研究所共同研究員」

    2021.4 - 2022.3

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  • 新潟大学医歯学総合病院 講師

    2009.4 - 2024.3

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  • 日本学術振興会海外特別研究員 ルイパスツール・ストラスブルブ大学(フランス)

    2002.4 - 2003.1

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  • 新潟大学歯学部 助手

    2000.1 - 2009.3

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  • 日本学術振興会特別研究員(PD)

    1997.4 - 1999.12

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  • ルイパスツール・ストラスブルグ大学(フランス)リサーチフェロー

    1995.11 - 1996.10

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  • 新潟大学歯学部付属病院 医員

    1989.4 - 1995.10

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Research History

  • Niigata University   Institute of Medicine and Dentistry, Academic Assembly   Professor

    2024.4

  • Niigata University   Dental Educational Research Development, Master's Program of Oral Health and Welfare Science, Graduate School of Medical and Dental Sciences   Professor

    2024.4

  • Niigata University   University Medical and Dental Hospital Dental Health   Lecturer

    2009.4 - 2024.3

  • Niigata University   Graduate School of Medical and Dental Sciences Oral Life Science   Lecturer

    2009.4 - 2024.3

  • Niigata University   Graduate School of Medical and Dental Sciences Oral Life Science   Assistant Professor

    2004.4 - 2009.3

Education

  • Niigata University   Faculty of Dentistry   School of Dentistry

    - 1989

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    Country: Japan

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  • Niigata University   Faculty of Dentistry

    - 1989

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Professional Memberships

 

Papers

  • A novel macrolide–Del-1 axis to regenerate bone in old age Reviewed

    Kridtapat Sirisereephap, Hikaru Tamura, Jong-Hyung Lim, Meircurius Dwi Condro Surboyo, Toshihito Isono, Takumi Hiyoshi, Andrea L. Rosenkranz, Yurie Sato-Yamada, Hisanori Domon, Akari Ikeda, Tomoyasu Hirose, Toshiaki Sunazuka, Nagako Yoshiba, Hiroyuki Okada, Yutaka Terao, Takeyasu Maeda, Koichi Tabeta, Triantafyllos Chavakis, George Hajishengallis, Tomoki Maekawa

    iScience   27 ( 2 )   108798 - 108798   2024.2

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    Publishing type:Research paper (scientific journal)   Publisher:Elsevier BV  

    DOI: 10.1016/j.isci.2024.108798

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  • 歯根形成時におけるTissue nonspecific alkaline phosphataseの機能解析

    大倉 直人, 吉羽 永子, 高原 信太郎, Baldeon Gutierrez Rosa Edith, Gomez-Kasimoto Susan, 井田 貴子, 枝並 直樹, 竹中 彰治, 吉羽 邦彦, 野杁 由一郎

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   159回   92 - 92   2023.10

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  • 再生歯内療法後の治癒過程初期における各種間葉系幹細胞マーカー陽性細胞の局在性

    高原 信太郎, 大倉 直人, 吉羽 永子, 竹中 彰治, 枝並 直樹, 吉羽 邦彦, 野杁 由一郎

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   159回   47 - 47   2023.10

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  • 歯髄再生療法モデルラットを用いた歯根成長段階による治癒形態の比較解析

    高原 信太郎, 大倉 直人, 吉羽 邦彦, 吉羽 永子, 竹中 彰治, 枝並 直樹, 庭野 和明, 野杁 由一郎

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   158回   31 - 31   2023.5

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  • In Vivo assessment of the Apatite-Forming Ability of New-Generation Hydraulic Calcium Silicate Cements Using a Rat Subcutaneous Implantation Model Reviewed International journal

    Naoki Edanami, Shouji Takenaka, Razi Saifullah, Ibn Belal, Kunihiko Yoshiba, Shintaro Takahara, Nagako Yoshiba, Naoto Ohkura, Yuichiro Noiri

    Journal of Functional Biomaterials   14 ( 4 )   2023.4

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    Hydroxyapatite formation on endodontic hydraulic calcium silicate cements (HCSCs) plays a significant role in sealing the root canal system and elevating the hard-tissue inductivity of the materials. This study evaluated the in vivo apatite-forming ability of 13 new-generation HCSCs using an original HCSC (white ProRoot MTA: PR) as a positive control. The HCSCs were loaded into polytetrafluoroethylene tubes and implanted in the subcutaneous tissue of 4-week-old male Wistar rats. At 28 days after implantation, hydroxyapatite formation on the HCSC implants was assessed with micro-Raman spectroscopy, surface ultrastructural and elemental characterization, and elemental mapping of the material-tissue interface. Seven new-generation HCSCs and PR had a Raman band for hydroxyapatite (v1 PO43- band at 960 cm-1) and hydroxyapatite-like calcium-phosphorus-rich spherical precipitates on the surfaces. The other six HCSCs with neither the hydroxyapatite Raman band nor hydroxyapatite-like spherical precipitates did not show calcium-phosphorus-rich hydroxyapatite-layer-like regions in the elemental mapping. These results indicated that 6 of the 13 new-generation HCSCs possessed little or no ability to produce hydroxyapatite in vivo, unlike PR. The weak in vivo apatite-forming ability of the six HCSCs may have a negative impact on their clinical performance.

    DOI: 10.3390/jfb14040213

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  • In Vivo Assessment of the Calcium Salt-Forming Ability of a New Calcium Silicate-Based Intracanal Medicament: Bio-C Temp. Reviewed International journal

    Naoki Edanami, Razi Saifullah Ibn Belal, Shoji Takenaka, Kunihiko Yoshiba, Rosa Edith Baldeon Gutierrez, Shintaro Takahara, Nagako Yoshiba, Naoto Ohkura, Yuichiro Noiri

    Dentistry journal   11 ( 4 )   2023.3

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    Calcium salt precipitation induced by intracanal medicaments contributes to the formation of apical hard tissue during apexification. This study compared the calcium salt-forming ability of a new calcium silicate-based intracanal medicament (Bio-C Temp) with that of two commercial calcium hydroxide pastes (Calcipex Plane II and Vitapex) in a rat subcutaneous implantation model. Polytetrafluoroethylene tubes containing each of the three materials were subcutaneously implanted in 4-week-old male Wistar rats. After 28 days, the composition and amount of calcium salts formed at the material-tissue interface were assessed using micro-Raman spectroscopy, X-ray diffraction, and elemental mapping. The tested materials produced white precipitates that had Raman spectra with peaks corresponding to hydroxyapatite and calcite. X-ray diffraction detected hydroxyapatite formation on Calcipex Plane II and Vitapex implants, as well as calcite formation on all three materials. Elemental mapping revealed that Bio-C Temp generated significantly smaller calcium- and phosphorus-rich calcified regions within the subcutaneous connective tissue than Vitapex. These results indicate that Bio-C Temp produced less calcium salt in rat subcutaneous tissue than Vitapex, although all materials formed hydroxyapatite and calcite in rat subcutaneous tissue. Bio-C Temp could be less effective than Vitapex in promoting apical hard tissue formation during apexification.

    DOI: 10.3390/dj11040091

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  • Prostaglandin E2-Transporting Pathway and Its Roles via EP2/EP4 in Cultured Human Dental Pulp. Reviewed International journal

    Naoto Ohkura, Kunihiko Yoshiba, Nagako Yoshiba, Yohei Oda, Naoki Edanami, Hayato Ohshima, Shoji Takenaka, Takashi Okiji, Yuichiro Noiri

    Journal of endodontics   49 ( 4 )   Ahead of online - 418   2023.2

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    INTRODUCTION: Prostaglandin E2 (PGE2) exerts biological actions through its transport pathway involving intracellular synthesis, extracellular transport, and receptor binding. This study aimed to determine the localization of the components of the PGE2-transporting pathway in human dental pulp and explore the relevance of PGE2 receptors (EP2/EP4) to angiogenesis and dentinogenesis. METHODS: Protein localization of microsomal PGE2 synthase (mPGES), PGE2 transporters [(multidrug resistance-associated protein-4 (MRP4) and prostaglandin transporter (PGT)], and EP2/EP4 was analyzed using double immunofluorescence staining. Tooth slices from human third molars were cultured with or without butaprost (EP2 agonist) or rivenprost (EP4 agonist) for 1 week. Morphometric analysis of endothelial cell filopodia was performed to evaluate angiogenesis, and real-time polymerase chain reaction was performed to evaluate angiogenesis and odontoblast differentiation markers. RESULTS: MRP4 and PGT were colocalized with mPGES and EP2/EP4 in odontoblasts and endothelial cells. Furthermore, MRP4 was colocalized with mPGES and EP4 in human leukocyte antigen-DR-expressing dendritic cells. In the tooth slice culture, EP2/EP4 agonists induced significant increases in the number and length of filopodia and mRNA expression of angiogenesis markers (vascular endothelial growth factor and fibroblast growth factor-2) and odontoblast differentiation markers (dentin sialophosphoprotein and collagen type 1). CONCLUSIONS: PGE2-producing enzyme (mPGES), transporters (MRP4 and PGT), and PGE2-specific receptors (EP2/EP4) were immunolocalized in various cellular components of the human dental pulp. EP2/EP4 agonists promoted endothelial cell filopodia generation and upregulated angiogenesis- and odontoblast differentiation-related genes, suggesting that PGE2 binding to EP2/EP4 is associated with angiogenic and dentinogenic responses.

    DOI: 10.1016/j.joen.2023.01.009

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  • SVCT2-GLUT1-mediated ascorbic acid transport pathway in rat dental pulp and its effects during wound healing. Reviewed International journal

    Naoto Ohkura, Kunihiko Yoshiba, Nagako Yoshiba, Naoki Edanami, Hayato Ohshima, Shoji Takenaka, Yuichiro Noiri

    Scientific reports   13 ( 1 )   1251 - 1251   2023.1

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    Ascorbic acid (AA; vitamin C) plays a crucial role in the biosynthesis and secretion of collagen to produce the organic matrix of hard tissues. Nevertheless, the detailed mechanism by which AA induces reparative dentinogenesis is still unknown. This study aimed to investigate the pathway and function of AA during wound healing in a rat pulpotomy model. Sodium-dependent vitamin C transporter (SVCT) 2 and glucose transporter (GLUT) 1 were detected in odontoblasts, endothelial cells, and nerve fibers in normal pulp tissues. SVCT2 and GLUT1 were also expressed in odontoblast-like cells in pulpotomized tissues of Wistar rats, and immunopositive cells of SVCT2 were significantly increased at 5 days after pulpotomy (p < 0.05). By contrast, osteogenic disorder Shionogi (ODS) rats, which cannot generate AA, also expressed SVCT2 and GLUT1 in normal and wound healing conditions. However, in ODS rats, when compared with the AA-addition group, the formation of dentin bridges in the AA-loss group was not evident, a layer of osteopontin was significantly increased beneath the wound surface (p < 0.05), and alpha smooth muscle actin at the odontoblast-like cells observed along this layer was significantly increased (p < 0.05), but not Nestin. Moreover, the amounts of type 1 collagen generated in the reparative dentin and beneath the wound healing site were significantly diminished (p < 0.05). Macrophages expressing CD68 and CD206 increased beneath the wound site. Hence, AA may be involved in odontoblast-like cell differentiation and anti-inflammatory response during dental pulp wound healing. Our results provide new insights into the function of AA through SVCT2 and GLUT1 in reparative dentinogenesis and may help in developing new therapeutic targets for dental pulpal disease.

    DOI: 10.1038/s41598-023-28197-9

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  • 根尖孔外に漏出したバイオセラミック系シーラーと歯周組織の相互作用

    高原 信太郎, 枝並 直樹, 竹中 彰治, 吉羽 邦彦, 大倉 直人, 吉羽 永子, 野杁 由一郎

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   157回   125 - 125   2022.10

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  • ケイ酸カルシウム系貼薬剤と水酸化カルシウム系貼薬剤のBiomineralization Abilityの比較

    枝並 直樹, 竹中 彰治, 吉羽 邦彦, 大倉 直人, 吉羽 永子, 高原 信太郎, 野杁 由一郎

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   157回   126 - 126   2022.10

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  • Gli1+-PDL Cells Contribute to Alveolar Bone Homeostasis and Regeneration. Reviewed International journal

    N Shalehin, Y Seki, H Takebe, S Fujii, T Mizoguchi, H Nakamura, N Yoshiba, K Yoshiba, M Iijima, T Shimo, K Irie, A Hosoya

    Journal of dental research   220345221106921 - 220345221106921   2022.7

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    The periodontal ligament (PDL) contains mesenchymal stem cells (MSCs) that can differentiate into osteoblasts, cementoblasts, and fibroblasts. Nevertheless, the distribution and characteristics of these cells remain uncertain. Gli1, an essential hedgehog signaling transcription factor, functions in undifferentiated cells during embryogenesis. Therefore, in the present study, the differentiation ability of Gli1+ cells was examined using Gli1-CreERT2/ROSA26-loxP-stop-loxP-tdTomato (iGli1/Tomato) mice. In 4-wk-old iGli1/Tomato mice, Gli1/Tomato+ cells were only slightly detected in the PDL, around endomucin-expressing blood vessels. These cells had proliferated over time, localizing in the PDL as well as on the bone and cementum surfaces at day 28. However, in 8-wk-old iGli1/Tomato mice, Gli1/Tomato+ cells were quiescent, as most cells were not immunoreactive for Ki-67. These cells in 8-wk-old mice exhibited high colony-forming unit fibroblast activity and were capable of osteogenic, chondrogenic, and adipogenic differentiation in vitro. In addition, after transplantation of teeth of iGli1/Tomato mice into the hypodermis of wild-type mice, Tomato fluorescence indicating the progeny of Gli1+ cells was detected in the osteoblasts and osteocytes of the regenerated bone. These results demonstrate that Gli1+ cells in the PDL were MSCs and could contribute to the alveolar bone regeneration.

    DOI: 10.1177/00220345221106921

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  • 生体活性はケイ酸カルシウム系セメントに普遍的な特性か? 18種のケイ酸カルシウム系セメントの生体内評価

    枝並 直樹, イブンベラル・ラジサイフラー, 竹中 彰治, 吉羽 邦彦, 大倉 直人, 吉羽 永子, 野杁 由一郎

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   156回   24 - 24   2022.5

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  • Laminin Isoforms in Human Dental Pulp: Lymphatic Vessels Express Laminin-332 and Schwann Cell-Associated Laminin-211 Modulates CD163 Expression of M2-Like Macrophages Reviewed International journal

    Nagako Yoshiba, Naoki Edanami, Naoto Ohkura, Tomoki Maekawa, Naoki Takahashi, Takahiro Tsuzuno, Takeyasu Maeda, Koichi Tabeta, Kenji Izumi, Yuichiro Noiri, Kunihiko Yoshiba

    ImmunoHorizons   5 ( 12 )   1008 - 1020   2021.12

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    Laminin, a basement membrane heterotrimeric glycoprotein composed of α/β/γ subunits, has important tissue-specific functions in the control of cellular behavior. Our recent study showed the colocalization of CD163+ M2-like macrophages with Schwann cells in human dental pulp, leading us to hypothesize that the laminin isoform of Schwann cells is associated with CD163 expression. The present study investigated the distribution of laminin isoforms in human dental pulp and the underlying mechanisms that affect macrophage phenotypes. Immunofluorescence analysis indicated that blood vessels were exclusively positive for laminin α4 and α5, whereas laminin α2 was associated with Schwann cells. Unexpectedly, laminin α3/laminin-332 (α3β3γ2) was detected on lymphatic vessels. In intact and carious teeth, CD163+ cells were associated with laminin α2, whereas CD206 single-positive cells were present inside, outside, and along blood vessels. In vitro incubation of THP-1 macrophages in plates coated with laminin-211/511 or its functionally analogous E8 fragments of α-chain (E8-α) indicated that cell shapes differed between macrophages grown on laminin-211/E8-α2 and macrophages grown on laminin-511/E8-α5. Laminin-211/E8-α2-coated plates upregulated CD163 expression, compared with laminin-511/E8-α5-coated plates. Integrin α3- and integrin α6-neutralizing Abs altered the shape of THP-1 macrophages and upregulated mRNA levels of CD206 and CD163 in macrophages grown on laminin-511; the neutralizing Abs did not affect macrophages grown on laminin-211. These findings suggest that laminin isoforms differentially regulate macrophage behavior via distinct integrin-laminin affinities. Of note, laminin-332 is expressed by pulpal lymphatic vessels, the existence of which has been debated; laminin-211 might have a role in maintaining CD163 expression on macrophages.

    DOI: 10.4049/immunohorizons.2100110

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  • Effect of a resin-modified calcium silicate cement on inflammatory cell infiltration and reparative dentin formation after pulpotomy in rat molars. Reviewed International journal

    Naoki Edanami, Razi Saifullah Ibn Belal, Kunihiko Yoshiba, Nagako Yoshiba, Naoto Ohkura, Shoji Takenaka, Yuichiro Noiri

    Australian endodontic journal : the journal of the Australian Society of Endodontology Inc   48 ( 2 )   297 - 304   2021.10

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    Resin monomers and polymerisation initiators have been shown to be cytotoxic for pulp cells and to disturb odontoblast differentiation. This study aimed to compare the effect of a resin-modified calcium silicate cement (TheraCal LC; TC) and a resin-free calcium silicate cement (ProRoot MTA; PR) on pulpal healing after pulpotomy. Pulpotomy was performed on the maxillary first molars of 8-week-old rats using either PR or TC. After 1, 3, 7, 14 and 28 days, pulpal responses were assessed by micro-computed tomography, haematoxylin-eosin staining and immunostaining against CD68, which is a pan-macrophage marker. The results showed that pulpotomy with TC induced persistent infiltration of inflammatory cells, including CD68-positive macrophages, and delayed the formation of reparative dentin as compared with that with PR, although both materials allowed pulpal healing over the long term. Therefore, resin-modified TC was not as biocompatible nor bioinductive as resin-free PR when applied on the healthy pulp of rat molars.

    DOI: 10.1111/aej.12568

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  • Apatite-Forming Ability of Flowable vs. Putty Formulations of Newly Developed Bioactive Glass-Containing Endodontic Cement Reviewed

    Naoki Edanami, Razi Saifullah Ibn Belal, Shoji Takenaka, Kunihiko Yoshiba, Nagako Yoshiba, Naoto Ohkura, Shintaro Takahara, Yuichiro Noiri

    Applied Sciences   11 ( 19 )   8969 - 8969   2021.9

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    This study compared the apatite-forming ability (AFA) levels of flowable and putty formulations of Nishika Canal Sealer BG Multi (F-NBG and P-NBG, respectively) and attempted to clarify the cause of differences in the AFA levels of F-NBG and P-NBG. NBG samples were aged in simulated body fluid (SBF) or 1-, 5-, or 10-g/L bovine serum albumin-containing SBF (BSA-SBF) and analyzed in terms of their ultrastructures, elemental compositions, and Raman spectra to identify apatite formation. The phosphate ion consumption rates of NBG samples in the media were evaluated as an indicator of apatite growth. The original elemental composition, calcium ion release, and alkalizing ability levels of F-NBG and P-NBG were also evaluated. Apparent apatite formation was detected on all NBG samples except F-NBG aged in 10-g/L BSA-SBF. P-NBG consumed phosphate ions faster than F-NBG. As-prepared P-NBG showed more silicon elements on its surface than as-prepared F-NBG. P-NBG released more calcium ions than F-NBG, although their alkalizing ability levels did not differ statistically. In conclusion, the AFA of P-NBG was greater than that of F-NBG, probably because of the greater ability of P-NBG to expose silanol groups on the surface and release calcium ions.

    DOI: 10.3390/app11198969

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  • Comparison of calcium and hydroxyl ion release ability and in vivo apatite-forming ability of three bioceramic-containing root canal sealers. Reviewed International journal

    Razi Saifullah Ibn Belal, Naoki Edanami, Kunihiko Yoshiba, Nagako Yoshiba, Naoto Ohkura, Shoji Takenaka, Yuichiro Noiri

    Clinical oral investigations   26 ( 2 )   1443 - 1451   2021.8

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    OBJECTIVE: Bioceramic-containing root canal sealers promote periapical healing via Ca2+ and OH- release and apatite formation on the surface. This study aimed to compare Ca2+ and OH- release and in vivo apatite formation of three bioceramic-containing root canal sealers: EndoSequence BC sealer (Endo-BC), MTA Fillapex (MTA-F), and Nishika Canal Sealer BG (N-BG). MATERIALS AND METHODS: Polytetrafluoroethylene tubes filled with sealers were immersed in distilled water for 6 and 12 h and for 1, 7, 14, and 28 days to measure Ca2+ and OH- release. Additionally, tubes filled with sealers were implanted in the backs of rats for 28 days, and in vivo apatite formation was analyzed using an electron probe microanalyzer. RESULTS: Endo-BC released significantly more Ca2+ than the other sealers at 6 and 12 h and 1 day. Ca2+ release was significantly lower from N-BG than from Endo-BC and MTA-F at 14 and 28 days. OH- release was significantly higher from Endo-BC than from the other sealers throughout the experiment, except at 1 day. OH- release was lower from N-BG than from MTA-F at 6 h and 7 days. Only Endo-BC implants exhibited apatite-like calcium-, phosphorus-, oxygen-, and carbon-rich spherulites and apatite layer-like calcium- and phosphorus-rich, but radiopaque element-free, surface regions. CONCLUSIONS: Ca2+ and OH- release is ranked as follows: Endo-BC > MTA-F > N-BG. Only Endo-BC demonstrated in vivo apatite formation. CLINICAL RELEVANCE: Endo-BC could promote faster periapical healing than MTA-F and N-BG.

    DOI: 10.1007/s00784-021-04118-w

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  • Impact of remnant healthy pulp and apical tissue on outcomes after simulated regenerative endodontic procedure in rat molars. Reviewed International journal

    Naoki Edanami, Kunihiko Yoshiba, Mari Shirakashi, Razi Saifullah Ibn Belal, Nagako Yoshiba, Naoto Ohkura, Aiko Tohma, Ryosuke Takeuchi, Takashi Okiji, Yuichiro Noiri

    Scientific reports   10 ( 1 )   20967 - 20967   2020.12

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    When regenerative endodontic procedures (REPs) are performed on immature teeth diagnosed with pulp necrosis and apical periodontitis, various healing patterns occur. Furthermore, infected immature teeth with endodontic disorders often exhibit some remnant pulp and apical tissue. Therefore, this study investigated the impact of remnant healthy or fully functional pulp and apical tissue on healing patterns after REPs. Simulated REPs were performed on non-infected immature rat molars with different amounts of remnant pulp and apical tissue. Healing patterns in these teeth were assessed after 28 days. Teeth with 0.81-0.91 mm of remnant pulp healed with pulp-like tissue, dentin, and osteodentin-like dentin-associated mineralized tissue (OSD-DAMT); teeth with 0.60-0.63 mm of remnant pulp healed with pulp-like tissue and OSD-DAMT; teeth with 0.13-0.43 mm of remnant pulp healed with periodontal ligament (PDL)-like tissue, OSD-DAMT, and cementum-like dentin-associated mineralized tissue (CEM-DAMT); and teeth with disorganization of pulp and apical tissues at 0.15-0.38 mm beyond the root apex healed with PDL-like tissue, CEM-DAMT, and intracanal bone (IB). Loss of Hertwig's epithelial root sheath was observed with IB formation. These results showed that four distinct healing patterns occurred after REPs, depending on the preoperative amount of remnant healthy pulp and apical tissue.

    DOI: 10.1038/s41598-020-78022-w

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  • 各種バイオセラミック系覆髄材のアパタイト析出能に関する研究

    枝並 直樹, イブンベラル・ラジサイフラー, 白柏 麻里, 吉羽 邦彦, 大倉 直人, 吉羽 永子, 遠間 愛子, 竹内 亮祐, 野杁 由一郎

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   152回   97 - 97   2020.6

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  • in vitro・in vivoにおける各種バイオセラミック系シーラーのアパタイト析出能に関する研究(Apatite forming ability of different bioceramic based root canal sealers in vitro and in vivo)

    イブンベラル・ラジサイフラー, 枝並 直樹, 白柏 麻里, 吉羽 邦彦, 大倉 直人, 吉羽 永子, 遠間 愛子, 竹内 亮祐, 野杁 由一郎

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   152回   46 - 46   2020.6

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  • M2 Phenotype Macrophages Colocalize with Schwann Cells in Human Dental Pulp. Reviewed International journal

    Yoshiba N, Edanami N, Ohkura N, Maekawa T, Takahashi N, Tohma A, Izumi K, Maeda T, Hosoya A, Nakamura H, Tabeta K, Noiri Y, Yoshiba K.

    Journal of Dental Research   99 ( 3 )   329 - 338   2020.3

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    Macrophages are immune cells with high plasticity that perform many functions related to tissue injury and repair. They are generally categorized as 2 functional phenotypes: M1 (proinflammatory) and M2 (anti-inflammatory and prohealing). To investigate the role of macrophages in human dental pulp, we examined the localization and distributional alterations of macrophages in healthy dental pulp as well as during the reparative process of pulp capping with mineral trioxide aggregate (MTA) and in cariously inflamed pulp of adult human teeth. We also quantified the populations of M1/M2 macrophages in healthy dental pulp by flow cytometric analysis. CD68+CD86+ cells (M1 phenotype) and CD68+CD163+ cells (M2 phenotype) were 2.11% ± 0.50% and 44.99% ± 2.22%, respectively, of 2.96% ± 0.41% CD68+ cells (pan-macrophages) in whole healthy dental pulp. Interestingly, M2 phenotype macrophages were associated with Schwann cells in healthy pulp, during mineralized bridge formation, and in pulp with carious infections in vivo. Furthermore, the M2 macrophages associated with Schwann cells expressed brain-derived neurotrophic factor (BDNF) under all in vivo conditions. Moreover, we found that plasma cells expressed BDNF. Coculture of Schwann cells isolated from human dental pulp and human monocytic cell line THP-1 showed that Schwann cells induced M2 phenotypic polarization of THP-1 cell-derived macrophages. The THP-1 macrophages that maintained contact with Schwann cells were stimulated, leading to elongation of their cell shape and expression of M2 phenotype marker CD163 in cocultures. In summary, we revealed the spatiotemporal localization of macrophages and potent induction of the M2 phenotype by Schwann cells in human dental pulp. M2 macrophages protect neural elements, whereas M1 cells promote neuronal destruction. Therefore, suppressing the neurodestructive M1 phenotype and maintaining the neuroprotective M2 phenotype of macrophages by Schwann cells may be critical for development of effective treatment strategies to maintain the viability of highly innervated dental pulp.

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  • Immunohistochemistry and gene expression of GLUT1, RUNX2 and MTOR in reparative dentinogenesis. Reviewed International journal

    Ryosuke Takeuchi, Naoto Ohkura, Kunihiko Yoshiba, Aiko Tohma, Nagako Yoshiba, Naoki Edanami, Mari Shirakashi, Razi Saifullah Ibn Belal, Hayato Ohshima, Yuichiro Noiri

    Oral diseases   26 ( 2 )   341 - 349   2020.3

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    OBJECTIVES: To determine glucose transporter 1 (GLUT1) and runt-related transcription factor 2 (RUNX2) expression during reparative dentinogenesis after pulpotomy with mineral trioxide aggregate (MTA) capping. SUBJECTS AND METHODS: Eight-week-old male Wistar rats were used. Pulp of the upper left first molar was exposed and capped with MTA. The upper right first molar of the same animal was used as a control. After collecting molars at various time points, GLUT1, RUNX2 and mammalian target of rapamycin (MTOR) were examined by immunohistochemistry. mRNA levels of Slc2a1 (encoding GLUT1), Runx2, Nestin and Mtor were determined by real-time PCR. RESULTS: Pulp exhibited progressive formation of reparative dentine lined with GLUT1- and MTOR-immunoreactive odontoblast-like cells at 5 days after pulpotomy. RUNX2 was detected in nuclei of most pulp tissue cells at day 5 after pulpotomy. Double immunofluorescence staining revealed GLUT1 immunoreactivity on odontoblast-like cells positive for Nestin or RUNX2, 5 days after pulpotomy. Slc2a1, Runx2, Nestin and Mtor mRNA levels were significantly upregulated on days 3-5 after pulpotomy. CONCLUSIONS: After rat molar pulpotomy, dental pulp induced formation of reparative dentine with colocalization of GLUT1 and Nestin or RUNX2. Moreover, mRNA levels of Slc2a1, Runx2, Nestin and Mtor were significantly upregulated in pulpotomized dental pulp.

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  • Glucose Transporter 2 and 4 Are Involved in Glucose Supply during Pulpal Wound Healing after Pulpotomy with Mineral Trioxide Aggregate in Rat Molars. Reviewed International journal

    Aiko Tohma, Naoto Ohkura, Kunihiko Yoshiba, Ryosuke Takeuchi, Nagako Yoshiba, Naoki Edanami, Mari Shirakashi, Razi Saifullah Ibn Belal, Hayato Ohshima, Yuichiro Noiri

    Journal of endodontics   46 ( 1 )   81 - 88   2020.1

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    INTRODUCTION: Pulp capping materials allow healing of injured pulp with a layer of reparative dentin. Glucose is needed to cure the injured area. Glucose is transported by glucose transporter (Glut) 2 and Glut4, which are transmembrane proteins that act as gatekeepers. We hypothesized that the transport of glucose via Glut2/Glut4 might contribute to the production of a dentin bridge during wound healing. Therefore, we explored Glut2 and Glut4 expression during reparative dentinogenesis after mineral trioxide aggregate capping. METHODS: The upper left first molar of 8-week-old Wistar rats underwent pulpotomy with mineral trioxide aggregate. At 1, 3, 5, 7, and 14 days after treatment, localization and colocalization of Glut2, Glut4, nestin (odontoblast marker), and antiendothelial cell antigen 1 (RECA-1; endothelial cell marker) were analyzed with immunohistochemical staining. Messenger RNA expression levels of Slc2a2 (encoding Glut2), Slc2a4 (encoding Glut4), Igf-1r (encoding insulinlike growth factor 1 receptor), and nestin were analyzed in the extracted teeth using real-time polymerase chain reaction. RESULTS: Glut2 and Glut4 were localized within odontoblasts and endothelial cells in normal control teeth. Three days after pulpotomy, Glut2- and Glut4-positive cells were detected; 7 days after pulpotomy, immunoreactivity for Glut2 and Glut4 was confined to newly differentiated odontoblastlike cells arranged beneath reparative dentin. Messenger RNA expression levels of Slc2a2 and Slc2a4 were significantly up-regulated after pulpotomy. CONCLUSIONS: Glut2 and Glut4 regulate glucose transport during wound healing beneath the injured area. This may contribute to the development of new vital pulp therapy for patients with deep caries.

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  • Detection of bone marrow-derived fibrocytes in human dental pulp repair. Reviewed International journal

    N Yoshiba, N Edanami, A Tohma, R Takeuchi, N Ohkura, A Hosoya, Y Noiri, H Nakamura, K Yoshiba

    International endodontic journal   51 ( 11 )   1187 - 1195   2018.11

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    AIM: To explore the expression profile of CD45+/pro-collagen I+ fibrocytes in intact dental pulps as well as during wound healing in adult dental pulp tissue. METHODOLOGY: A total of 16 healthy permanent teeth were obtained from young patients (18 to 25 years) undergoing orthodontic treatment. Routine pulp capping with mineral trioxide aggregate (MTA) was performed under local anaesthesia to induce a mineralized barrier at the exposed surface. Teeth were extracted from patients after 7, 14 and 35 days. Sections of the extracted teeth were prepared and stained for various markers using indirect immunofluorescence. Fibrocytes were counted, and the data were statistically evaluated using the Dunnett test. RESULTS: In uninflammed pulp tissue, a pro-collagen I-positive reaction was detected in odontoblasts, as well as in perivascular cells. Most of the CD45-positive cells were negative for pro-collagen I in normal pulp tissue, whereas CD45+/pro-collagen I+ fibrocytes were detected 7 days after injury. At day 14, fibrocytes were recognized under the fibrous matrix in contact with MTA and had infiltrated into regions of new capillary formation, where the fibrocytes were positively stained for vascular endothelial growth factor. By 35 days, fibrocytes were few, coincident with the formation of dentine bridges. The number of fibrocytes peaked 7 days post-injury and decreased at 14 days. CONCLUSIONS: The presence of fibrocytes in human pulp wound healing was observed. The spatiotemporal distribution of fibrocytes suggests that fibrocytes are involved in the early stages of pulp wound healing, specifically by contributing to new blood vessel formation.

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  • Orthodontic force application upregulated pain-associated prostaglandin-I2/PGI2-receptor/TRPV1 pathway-related gene expression in rat molars. Reviewed

    Mariko Ohkura, Naoto Ohkura, Nagako Yoshiba, Kunihiko Yoshiba, Hiroko Ida-Yonemochi, Hayato Ohshima, Isao Saito, Takashi Okiji

    Odontology   106 ( 1 )   2 - 10   2018.1

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    This study aimed to analyze the mRNA expression and protein localization of prostaglandin I2 (PGI2) synthase (PGIS), the PGI2 receptor (IP receptor) and transient receptor potential cation channel, subfamily V, member 1 (TRPV1) in force-stimulated rat molars, toward the elucidation of the PGI2-IP receptor-TRPV1 pathway that is in operation in the pulp and possibly associated with orthodontic pain and inflammation. Experimental force was applied to the maxillary first and second molars by inserting an elastic band between them for 6-72 h. PGIS, PTGIR (the IP receptor gene), and TRPV1 mRNA levels in the coronal pulp were analyzed with real-time PCR. PGIS, IP receptor, and TRPV1 proteins were immunostained. The force stimulation induced significant upregulation of PGIS at 6-24 h, and PTGIR and TRPV1 at 6 and 12 h in the pulp. PGIS was immunolocalized in odontoblasts and some fibroblasts in the force-stimulated pulp. The IP receptor and TRPV1 immunoreactivities were detected on odontoblasts and some nerve fibers. It was concluded that PGIS, PTGIR, and TRPV1 in rat molar pulp were significantly upregulated shortly after the force application, and that the IP receptor was co-expressed on TRPV1-expressing nerves and odontoblasts. These findings suggest that the PGI2-IP receptor-TRPV1 pathway is associated with the acute phase of force-induced pulp changes involving odontoblasts and nerves.

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  • Bioactivity and biomineralization ability of calcium silicate-based pulp-capping materials after subcutaneous implantation Reviewed

    G. Hinata, K. Yoshiba, L. Han, N. Edanami, N. Yoshiba, T. Okiji

    INTERNATIONAL ENDODONTIC JOURNAL   50   E40 - E51   2017.12

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    AimTo evaluate the abilities of three calcium silicate-based pulp-capping materials (ProRoot MTA, TheraCal LC and a prototype tricalcium silicate cement) to produce apatite-like precipitates after being subcutaneously implanted into rats.
    MethodologyPolytetrafluoroethylene tubes containing each material were subcutaneously implanted into the backs of Wistar rats. At 7, 14 and 28days post-implantation, the implants were removed together with the surrounding connective tissue, and fixed in 2.5% glutaraldehyde in cacodylate buffer. The chemical compositions of the surface precipitates formed on the implants were analysed with scanning electron microscopy-electron probe microanalysis (SEM-EPMA). The distributions of calcium (Ca) and phosphorus (P) at the material-tissue interface were also analysed with SEM-EPMA. Comparisons of the thicknesses of the Ca- and P-rich areas were performed using the Friedman test followed by Scheffe's test at a significant level of 5%.
    ResultsAll three materials produced apatite-like surface precipitates containing Ca and P. For each material, elemental mapping detected a region of connective tissue in which the concentrations of Ca and P were higher than those in the surrounding connective tissue. The thickness of this Ca- and P-rich region exhibited the following pattern: ProRoot MTA &gt; prototype tricalcium silicate cementTheraCal LC. ProRoot MTA had a significantly thicker layer of Ca and P than the other materials at all time-points (P&lt;0.05), and a significant difference was detected between the prototype cement and TheraCal LC at 28days (P&lt;0.05).
    ConclusionAfter being subcutaneously implanted, all of the materials produced Ca- and P-containing surface precipitates and a Ca- and P-rich layer within the surrounding tissue. The thickness of the Ca- and P-rich layer of ProRoot MTA was significantly thicker than that of the other materials.

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  • Effects of pulpotomy using mineral trioxide aggregate on prostaglandin transporter and receptors in rat molars. Reviewed International journal

    Naoto Ohkura, Naoki Edanami, Ryosuke Takeuchi, Aiko Tohma, Mariko Ohkura, Nagako Yoshiba, Kunihiko Yoshiba, Hiroko Ida-Yonemochi, Hayato Ohshima, Takashi Okiji, Yuichiro Noiri

    Scientific reports   7 ( 1 )   6870 - 6870   2017.7

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    Mineral trioxide aggregate (MTA) is a commonly used dental pulp-capping material with known effects in promoting reparative dentinogenesis. However, the mechanism by which MTA induces dentine repair remains unclear. The aim of the present study was to investigate the role of prostaglandin E2 (PGE2) in dentine repair by examining the localisation and mRNA expression levels of its transporter (Pgt) and two of its receptors (Ep2 and Ep4) in a rat model of pulpotomy with MTA capping. Ep2 expression was detected in odontoblasts, endothelial cells, and nerve fibres in normal and pulpotomised tissues, whereas Pgt and Ep4 were immunolocalised only in the odontoblasts. Moreover, mRNA expression of Slco2a1 (encoding Pgt), Ptger2 (encoding Ep2), and Ptger4 (encoding Ep4) was significantly upregulated in pulpotomised dental pulp and trigeminal ganglia after MTA capping. Our results provide insights into the functions of PGE2 via Pgt and Ep receptors in the healing dentine/pulp complex and may be helpful in developing new therapeutic targets for dental disease.

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  • Characterization of Dental Pulp Myofibroblasts in Rat Molars after Pulpotomy Reviewed

    Naoki Edanami, Nagako Yoshiba, Naoto Ohkura, Ryosuke Takeuchi, Aiko Tohma, Yuichiro Noiri, Kunihiko Yoshiba

    JOURNAL OF ENDODONTICS   43 ( 7 )   1116 - 1121   2017.7

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    Introduction: Myofibroblasts express alpha smooth muscle actin (alpha-SMA) and play a critical role in wound healing. Myofibroblast differentiation is controlled by the joint actions of transforming growth factor beta 1 (TGF-beta 1) and the extradomain A fibronectin splice variant (EDA-FN). Currently, the contribution of myofi-broblasts to dental pulp healing is unknown. Therefore, we analyzed expressional characteristics of alpha-SMA positive cells and investigated TGF-beta 1, EDA-FN, and alpha-SMA expression levels after pulpotomy to better understand dental pulp healing. Methods: The maxillary first molars of 8-week-old Wistar rats were pulpotomized with mineral trioxide aggregate. After 1 to 14 days, localization and colocalization of alpha-SMA, rat endothelial cell antigen-1 (as a marker of endothelial cells), neuron-glial antigen 2 (as a marker of perivascular cells), prolyl-4-hydroxylase (P4H, as an additional marker of myofibroblasts), and EDA-FN were analyzed using immunohistochemistry and double immunofluorescence. Time-course changes in the messenger RNA expression levels of TGF-beta 1, EDA-FN, and alpha-SMA were evaluated using quantitative real-time polymerase chain reaction analysis. Results: Spindle-shaped alpha-SMA positive cells transiently appeared after pulpotomy. These cells initially emerged in the pulp core on day 3 and then accumulated at the wound site by day 5. These cells were isolated from rat endothelial cell antigen-1 positive cells and did not express neuron-glial antigen 2 but did express P4H. The messenger RNA levels of TGF-beta 1, EDA-FN, and alpha-SMA were significantly up-regulated after pulpotomy. EDA-FN and a-SMA were colocalized at the wound sites on day 5. Conclusions: In association with up regulation of TGF-beta 1 and EDA-FN expression, alpha-SMA and P4H double-positive cells accumulated at the wound sites after pulpotomy. This suggests that myofibroblasts participate in dental pulp healing.

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  • GaAlAs laser-induced pulp mineralization involves dentin matrix protein 1 and osteopontin expression Reviewed

    Y. Shigetani, N. Ohkura, K. Yoshiba, H. Ohshima, A. Hosoya, N. Yoshiba, T. Okiji

    ORAL DISEASES   22 ( 5 )   399 - 405   2016.7

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    ObjectivesGaAlAs lasers induce pulp mineralization by promoting reparative dentinogenesis. This study analyzed the expression of dentin matrix protein 1 (DMP1) and osteopontin in GaAlAs laser-irradiated rat molars, to examine the hypothesis that these proteins play a role in the laser-induced reparative dentinogenic process.
    Materials and methodsThe mesial surfaces of the upper first molars of 8-week-old Wistar rats were irradiated with a pulsed GaAlAs laser. After 1-14days, mRNA expression of DMP1 and osteopontin in the coronal pulp was analyzed using real-time PCR. DMP1, osteopontin, and heat shock protein 25 (HSP25) were immunolocalized at 1-21days.
    ResultsThe pulp exhibited a degenerative zone in its mesial portion on days 1-3, and progressive formation of reparative dentin lined with HSP25-immunoreactive odontoblast-like cells, from day 7 onwards. DMP1 and osteopontin mRNA expression were significantly upregulated on days 1-7 and 3-7, respectively. From day 7 onwards, DMP1 and osteopontin immunoreactivity colocalized along the boundary between the primary and reparative dentin.
    ConclusionGaAlAs laser irradiation of rat molars induced upregulated DMP1 and osteopontin mRNA expression in the coronal pulp, followed by the formation of reparative dentin and the colocalization of DMP1 and osteopontin immunoreactivity at the site at which this tissue first appeared.

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  • Correlation between Fibrillin-1 Degradation and mRNA Downregulation and Myofibroblast Differentiation in Cultured Human Dental Pulp Tissue Reviewed

    Nagako Yoshiba, Kunihiko Yoshiba, Naoto Ohkura, Erika Takei, Naoki Edanami, Youhei Oda, Akihiro Hosoya, Hiroaki Nakamura, Takashi Okiji

    JOURNAL OF HISTOCHEMISTRY & CYTOCHEMISTRY   63 ( 6 )   438 - 448   2015.6

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    Myofibroblasts and extracellular matrix are important components in wound healing. Alpha-smooth muscle actin (-SMA) is a marker of myofibroblasts. Fibrillin-1 is a major constituent of microfibrils and an extracellular-regulator of TGF-1, an important cytokine in the transdifferentiation of resident fibroblasts into myofibroblasts. To study the correlation between changes in fibrillin-1 expression and myofibroblast differentiation, we examined alterations in fibrillin-1 and -SMA expression in organotypic cultures of dental pulp in vitro. Extracted healthy human teeth were cut to 1-mm-thick slices and cultured for 7 days. In intact dental pulp, fibrillin-1 was broadly distributed, and -SMA was observed in pericytes and vascular smooth muscle cells. After 7 days of culture, immunostaining for fibrillin-1 became faint concomitant with a downregulation in its mRNA levels. Furthermore, fibroblasts, odontoblasts and Schwann cells were immunoreactive for -SMA with a significant increase in -SMA mRNA expression. Double immunofluorescence staining was positive for pSmad2/3, central mediators of TGF- signaling, and -SMA. The administration of inhibitors for extracellular matrix proteases recovered fibrillin-1 immunostaining; moreover, fibroblasts lost their immunoreactivity for -SMA along with a downregulation in -SMA mRNA. These findings suggest that the expression of -SMA is TGF-1 dependent, and fibrillin-1 degradation and downregulation might be implicated in the differentiation of myofibroblasts in dental pulp wound healing.

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  • Temporospatial localization of dentine matrix protein 1 following direct pulp capping with calcium hydroxide in rat molars Reviewed

    Y. Shigetani, K. Yoshiba, M. Kuratate, E. Takei, N. Yoshiba, Y. Yamanaka, H. Ohshima, T. Okiji

    INTERNATIONAL ENDODONTIC JOURNAL   48 ( 6 )   573 - 581   2015.6

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    AimTo examine the temporospatial expression of dentine matrix protein 1 (DMP1; a noncollagenous protein involved in mineralized tissue formation), osteopontin (another noncollagenous protein detected during reparative dentinogenesis) and nestin (a marker of differentiating/differentiated odontoblasts), following direct pulp capping with calcium hydroxide in rat molars.
    MethodologyThe maxillary first molars of 8-week-old Wistar rats had their pulps exposed and capped with calcium hydroxide. The pulp-capped teeth were collected from 6h to 14days postoperatively and processed for immunohistochemistry for DMP1, osteopontin and nestin. Cell proliferation was monitored using 5-bromo-2-deoxyuridine (BrdU) labelling.
    ResultsThe capped pulps initially exhibited superficial necrotic changes followed by the formation of new matrix and its mineralization. DMP1 immunoreactivity was observed in the matrix beneath the necrotic layer from 6h onwards and present in the outer portion of the newly formed mineralized matrix from 7days onwards. Osteopontin displayed a similar expression pattern, although it occupied a narrower area than DMP1 at 6 and 12h. Nestin-immunoreactive cells appeared beneath the DMP1-immunoreactive area at 1day, were distributed beneath the newly formed matrix at 5days and exhibited odontoblast-like morphology by 14days. BrdU-positive cells significantly increased at 2 and 3days (P&lt;0.05) and then decreased.
    ConclusionsThe deposition of DMP1 at exposed pulp sites preceded the appearance of nestin-immunoreactive cells, active cell proliferation and new matrix formation after pulp capping with calcium hydroxide in rat molars, suggesting that DMP1 acts as a trigger of pulp repair. The colocalization of DMP1 and osteopontin suggests that these two proteins play complementary roles.

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  • Initial Transient Accumulation of M2 Macrophage-associated Molecule-expressing Cells after Pulpotomy with Mineral Trioxide Aggregate in Rat Molars Reviewed

    Erika Takei, Yoshimi Shigetani, Kunihiko Yoshiba, Go Hinata, Nagako Yoshiba, Takashi Okiji

    JOURNAL OF ENDODONTICS   40 ( 12 )   1983 - 1988   2014.12

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    Introduction: M2 (alternatively activated) macrophages are known to participate in wound healing and tissue repair. This study aimed to analyze the temporospatial changes in the distribution and density of M2 macrophage associated molecule-expressing cells after pulpotomy with mineral trioxide aggregate (MTA) in rat molars to ascertain the role played by M2 macrophages in the healing of MTA-capped pulp tissue. Methods: The maxillary first molars of 8-week-old Wistar rats were pulpotomized and capped with MTA. After 1-14 days, the teeth were examined after hematoxylineosin staining or immunoperoxidase staining of CD68 (a general macrophage marker) and M2 macrophage markers (CD163 and CD204)1 The density of positively stained cells was enumerated in the surface and inner regions (0-100 mu m and 300-400 mu m, respectively, from the wound surface). Results: MTA capping initially caused mild inflammatory changes and the formation of a degenerative layer followed by progressive new matrix formation and calcified bridging. At 1-2 days, CD68-, CD163-, and CD204-positive cells started to accumulate beneath the degenerative layer, and the density of these cells was significantly higher in the surface region than in the inner region (P &lt; .05). From 7 days onward, the 3 types of cells displayed an almost normal distribution beneath the newly formed dentinlike matrix. Conclusions: After the pulpotomy of rat molars with MTA, M2 macrophage associated molecule-expressing cells transiently accumulated beneath the degenerative layer under the MTA. This suggests that M2 macrophages participate in the initial phases of the healing of MTA-capped pulp tissue.

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  • Prostaglandin Transporting Protein-mediated Prostaglandin E-2 Transport in Lipopolysaccharide-inflamed Rat Dental Pulp Reviewed

    Naoto Ohkura, Yoshimi Shigetani, Nagako Yoshiba, Kunihiko Yoshiba, Takashi Okiji

    JOURNAL OF ENDODONTICS   40 ( 8 )   1112 - 1117   2014.8

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    Introduction: The prostaglandin transporter (Pgt) and multidrug resistance-associated protein (Mrp) 4 are membrane transport proteins that play crucial roles in the transmembrane uptake and/or efflux of prostaglandins (PGs). This study attempted to analyze the protein expression of Pgt and Mrp4 and their involvement in PGE(2) efflux transport in lipopolysaccharide (LPS)-inflamed rat incisor pulp tissue. Methods: Pulpit's was induced in the upper incisors of Wistar rats by treating them with LPS for 24 hours. The protein expression levels of Pgt, Mrp4, and microsomal PGE synthase (mPGES) were analyzed with immunofluorescent staining. The amount of PGE(2) released from the inflamed pulp tissue in the presence or absence of dipyridamole (an Mrp4 inhibitor) was assessed by using an enzyme-linked immunosorbent assay. Results: Double immunofluorescence staining revealed that the Pgt, Mrp4, and mPGES immunoreactivity co-localized in CD31-expressing endothelial cells. Moreover, the Mrp4 inhibitor caused a significant decrease in the amount of PGE(2) released from the LPS-inflamed pulp (P &lt; .01 at 24 hours). Conclusion: Pgt, Mrp4, and mPGES expression was detected in the endothelial cells of normal and LPS-inflamed rat incisor pulp tissue, suggesting that these cells are associated with the biosynthesis and transmembrane transport of PGE(2). The significant decrease in PGE(2) release induced by the Mrp4 inhibitor suggests that Mrp4 contributes to the transport of PGE(2) in the transmembrane efflux pathway.

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  • M2 Macrophages Participate in the Biological Tissue Healing Reaction to Mineral Trioxide Aggregate Reviewed

    Takafumi Ito, Tomoatsu Kaneko, Yusuke Yamanaka, Yoshimi Shigetani, Kunihiko Yoshiba, Takashi Okiji

    JOURNAL OF ENDODONTICS   40 ( 3 )   379 - 383   2014.3

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    Introduction: This study examined the protein and messenger RNA (mRNA) expression of molecules associated with M2 (wound healing) macrophages in mineral trioxide aggregate (MTA)-implanted rat subcutaneous tissue to elucidate the involvement of M2 macrophages in the connective tissue response to MTA. Methods: Silicone tubes containing freshly mixed MTA or a calcium hydroxide cement (Life; Kerr, Romulus, MI) were subcutaneously implanted into the backs of Wistar rats. Solid silicone rods implanted in different animals served as controls. The specimens were then double immunostained for ED1 (CD68, a general macrophage marker) and ED2 (CD163, an M2 macrophage marker). Immunostaining for CD34 (a marker for vascularization and wound healing) was also performed. Expression levels of CD34, CD163, and mannose receptor c type 1 (an M2 macrophage marker) mRNAs were determined with real-time polymerase chain reaction. Results: MTA-implanted subcutaneous tissues showed significant increases in the density of ED1+ED2+ macrophages beneath the implantation site and expression levels of CD163 and MMR mRNAs compared with Life-implanted and control tissues. MTA-implanted subcutaneous tissues also showed a significant increase of CD34-immunostained areas and up-regulation of CD34 mRNAs compared with Life-implanted and control tissues. Conclusions: MTA implantation induced the accumulation of M2 macrophage marker (ED2)-expressing macrophages and enhanced the expression of M2 macrophage marker genes. MTA implantation also enhanced the expression of CD34, suggesting acceleration of the healing/tissue repair process. Taken together, biological connective tissue response to MTA may involve wound healing/tissue repair processes involving M2 macrophages.

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  • Localization of SUMOylation factors and Osterix in odontoblast lineage cells during dentin formation and regeneration Reviewed

    Akihiro Hosoya, Akira Yukita, Tadashi Ninomiya, Toru Hiraga, Kunihiko Yoshiba, Nagako Yoshiba, Etsuo Kasahara, Hiroaki Nakamura

    HISTOCHEMISTRY AND CELL BIOLOGY   140 ( 2 )   201 - 211   2013.8

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    Small ubiquitin-related modifier (SUMO) conjugation (SUMOylation) is a post-translational modification involved in various cellular processes including the regulation of transcription factors. In this study, to analyze the involvement of SUMOylation in odontoblast differentiation, we examined the immunohistochemical localization of SUMO-1, SUMO-2/3, and Osterix during rat tooth development. At the bud and cap stages, localization of SUMOs and Osterix was hardly detected in the dental mesenchyme. At the bell stage, odontoblasts just beginning dentin matrix secretion and preodontoblasts near these odontoblasts showed intense immunoreactivity for these molecules. However, after the root-formation stage, these immunoreactivities in the odontoblasts decreased in intensity. Next, to examine whether the SUMOylation participates in dentin regeneration, we evaluated the distribution of SUMOs and Osterix in the dental pulp after cavity preparation. In the coronal pulp chamber of an untreated rat molar, odontoblasts and pulp cells showed no immunoreactivity. At 4 days after cavity preparation, positive cells for SUMOs and Osterix appeared on the surface of the dentin beneath the cavity. Odontoblast-like cells forming reparative dentin were immunopositive for SUMOs and Osterix at 1 week, whereas these immunoreactivities disappeared after 8 weeks. Additionally, we further analyzed the capacity of SUMO-1 to bind Osterix by performing an immunoprecipitation assay using C2C12 cells, and showed that Osterix could undergo SUMOylation. These results suggest that SUMOylation might regulate the transcriptional activity of Osterix in odontoblast lineage cells, and thus play important roles in odontoblast differentiation and regeneration.

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  • Odontoblast response to cavity preparation with Er:YAG laser in rat molars: an immunohistochemical study Reviewed

    Yoshimi Shigetani, Hironobu Suzuki, Hayato Ohshima, Kunihiko Yoshiba, Nagako Yoshiba, Takashi Okiji

    ODONTOLOGY   101 ( 2 )   186 - 192   2013.7

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    This study aimed to examine the dynamics of odontoblast-lineage cells following cavity preparation with erbium:yttrium-aluminum-garnet (Er:YAG) laser in rat molars. Cavity preparation was made with Er:YAG laser in the mesial surface of the maxillary left first molar of 8-week-old Wistar rats. Contralateral first molar served as unirradiated control. Immediately, 6 and 12 h and 1, 2, 3, 5 and 7 days after the lasing (n = 5, each), specimens were collected and processed for immunohistochemistry for heat-shock protein (HSP)-25 and nestin as markers for odontoblast-lineage cells. Cell proliferation assay using bromodeoxyuridine (BrdU) labeling was also performed. Unirradiated teeth showed HSP-25- and nestin-immunoreactivity in odontoblasts. At 6-12 h after irradiation, the odontoblastic layer was disorganized and some of odontoblasts lost the immunoreactivity to HSP-25 and nestin. At 1-2 days, however, HSP-25- and nestin-immunoreactivities in the odontoblast layer showed a noticeable recovery, resulting in the rearrangement of odontoblast-like cells intensely immunoreactive to HSP-25 and nestin at 3-7 days. BrdU-positive cells showed a significant increase at 2 days (P &lt; 0.05 vs. immediate previous time point; one-way analysis of variance and Scheff, post hoc test), peaked at 3 days and then decreased significantly (P &lt; 0.05). It was concluded that under the present experimental condition in rat molars, cavity preparation with Er:YAG laser induced mild and reversible damage to odontoblasts. The reparative process was characterized by the rearrangement of HSP-25- and nestin-immunoreactive odontoblast-like cells, which took place subsequent to the odontoblastic layer disorganization with partial loss of these immunoreactivities.

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  • ヒト歯髄創傷治癒過程における細胞外基質の局在変化 Fibrillin-1基質の動的リモデリングに関する検索 Reviewed

    吉羽 永子, 吉羽 邦彦, 大倉 直人, 重谷 佳見, 武井 絵梨花, 細矢 明宏, 中村 浩彰, 興地 隆史

    日本歯科保存学雑誌   56 ( 3 )   161 - 168   2013.6

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    目的:細胞外基質fibrillin-1は,弾性線維の構成タンパクであると同時に,その分解によりtransforming growth factor-β(TGF-β)を遊離させることが知られている.また,著者らはfibrillin-1がヒト歯髄直接覆髄後の創傷治癒過程で発現を消失させることを見いだしている.そこで本研究では,TGF-β制御活性を有する細胞外基質decorinとfibronectin,および創傷治癒と組織線維化に関与するtenascin-Cが,ヒト歯髄直接覆髄後の創傷治癒過程でfibrillin-1様の局在変化を示すか否かを検索した.さらに,スライス培養したヒト歯髄を用いて,fibrillin-1の発現消失へのmatrix metalloproteinase(MMP)の関与の実態を検索した.材料と方法:矯正治療により抜歯予定のヒト健全歯をmineral trioxide aggregate(MTA)で直接覆髄した後,2週および6週後に抜歯し,fibrillin-1,fibronectin,decorinおよびtenascin-Cに対する免疫組織化学的染色を行った.また,健全歯髄を組織培養し,fibrillin-1 mRNA発現を定量RT-PCRで測定するとともに,MMP阻害剤であるNNGHを添加培養し,fibrillin-1タンパクの局在変化を免疫染色により解析した.結果:直接覆髄後2週および6週において,fibrillin-1に対する免疫陽性反応は,覆髄部直下の歯髄組織で局所的に消失していた.一方,fibronectin,decorinおよびtenascin-Cは同部で一様に陽性反応を示しており,明らかな局在変化は認められなかった.一方,培養歯髄組織ではfibrillin-1の染色性が減弱し,mRNAの発現も有意に低下したが,MMP-3 mRNA発現は有意に亢進した.NNGHを添加培養するとfibrillin-1の染色性が向上した.結論:検索対象とした4種のタンパクのなかでfibrillin-1のみ直接覆髄後の創傷治癒過程で局在変化を示した.Fibrillin-1免疫反応性の消失には,MMPsによるタンパクの分解に加えて遺伝子発現の下方制御も関与するものと考えられた.(著者抄録)

    DOI: 10.11471/shikahozon.56.161

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  • Mineral trioxide aggregateに対するラット皮下結合組織の応答 マクロファージ関連分子の免疫組織化学的・分子生物学的解析 Reviewed

    伊藤 崇史, 山中 裕介, 金子 友厚, 吉羽 邦彦, 吉羽 永子, 重谷 佳見, 興地 隆史

    日本歯科保存学雑誌   56 ( 1 )   9 - 16   2013.2

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    目的:Mineral trioxide aggregate(MTA)に対するマクロファージや樹状細胞の生体内での応答様式を追究するため,MTAを移植されたラット皮下結合組織における各種マクロファージ・樹状細胞関連分子の発現状況を,免疫組織化学的,もしくは定量的遺伝子発現解析により検討した.材料と方法:MTA(ProRoot MTA)および対照として水酸化カルシウム系覆髄材(Life)を被験材料とし,それぞれシリコンチューブへ充填後,Wistar系雄性ラットの背部皮下へ移植した.同一サイズのシリコンロッドをコントロール群とした.14日経過後に移植体周囲組織を採取し,ED1(CD68:抗マクロファージおよび樹状細胞)およびOX6(抗主要組織適合抗原クラス・分子)を用いて酵素抗体二重染色を施した後,ED1+/OX6+細胞およびED1+/OX6-細胞の密度を求めた.さらに,リアルタイムPCR法を用いてCD163(組織修復(M2)マクロファージのマーカー)およびCD34(血管内皮細胞,一部の真皮樹状細胞などが発現)のmRNA発現レベルを定量した.成績:MTA移植群では,ED1+/OX6+細胞はほかの全群と比較して,またED1+/OX6-細胞はLife移植群と比較して,有意に高い密度を示した(p&lt;0.05).また,MTA移植群におけるCD163およびCD34 mRNA発現は,他群より有意に高レベルであった(p&lt;0.05).結論:ラット皮下結合組織において,MTAはLifeと比較して有意に高密度のCD68陽性細胞亜群(ED1+/OX6+細胞およびED1+/OX6-細胞)の浸潤と,有意に高レベルのCD34,CD163 mRNA発現を誘導した.マクロファージの関与する創傷治癒・組織修復機構が,MTAに対する生体反応に関与する可能性が示唆された.(著者抄録)

    DOI: 10.11471/shikahozon.56.9

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  • Evaluation of the responses of MHC class II molecule-expressing cells and macrophages to epoxy resin-based and 4-META-containing, methacrylate resinbased root canal sealers in rat subcutaneous tissue Reviewed

    Yusuke Yamanaka, Yoshimi Shigetani, Kunihiko Yoshiba, Tomoatsu Kaneko, Nagako Yoshiba, Takashi Okiji

    Dental Materials Journal   32 ( 5 )   822 - 827   2013

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    Major histocompatibility complex (MHC) class II molecule-expressing cells and macrophages play a pivotal role in mediating the host tissue response to biomaterials. This study investigated the responses of these cells to epoxy resin-based and 4-META-containing, methacrylate resin-based endodontic sealers (AH Plus and MetaSEAL respectively) in rat connective tissue. Silicone tubes loaded with one of the sealers or solid silicone rods (control) were subcutaneously implanted in male Wistar rats for three time periods of 7, 14, or 28 days. Tissue specimens were immunoperoxidase-stained for MHC class II molecules and CD68 (a general macrophage marker). Results showed that AH Plus-implanted tissue displayed significantly more MHC class II-positive cells than the control at 14 and 28 days, whereas MetaSEAL-implanted tissue showed significantly more CD68-positive cells than both AH Plus-implanted tissue and the control at all time periods. It was concluded that the epoxy resin-based sealer induced the infiltration of MHC class II molecule-expressing cells, whereas 4-META-containing, methacrylate resin-based sealer elicited macrophage infiltration.

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  • Two Distinct Processes of Bone-like Tissue Formation by Dental Pulp Cells after Tooth Transplantation Reviewed

    Akihiro Hosoya, Akira Yukita, Kunihiko Yoshiba, Nagako Yoshiba, Masafumi Takahashi, Hiroaki Nakamura

    JOURNAL OF HISTOCHEMISTRY & CYTOCHEMISTRY   60 ( 11 )   861 - 873   2012.11

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    Dental pulp is involved in the formation of bone-like tissue in response to external stimuli. However, the origin of osteoblast-like cells constructing this tissue and the mechanism of their induction remain unknown. We therefore evaluated pulp mineralization induced by transplantation of a green fluorescent protein (GFP)-labeled tooth into a GFP-negative hypodermis of host rats. Five days after the transplantation, the upper pulp cavity became necrotic; however, cell-rich hard tissue was observed adjacent to dentin at the root apex. At 10 days, woven bone-like tissue was formed apart from the dentin in the upper pulp. After 20 days, these hard tissues expanded and became histologically similar to bone. GFP immunoreactivity was detected in the hard tissue-forming cells within the root apex as well as in the upper pulp. Furthermore, immunohistochemical observation of alpha-smooth muscle actin, a marker for undifferentiated cells, showed a positive reaction in cells surrounding this bone-like tissue within the upper pulp but not in those within the root apex. Immunoreactivities of Smad4, Runx2, and Osterix were detected in the hard tissue-forming cells within both areas. These results collectively suggest that the dental pulp contains various types of osteoblast progenitors and that these cells might thus induce bone-like tissue in severely injured pulp. (J Histochem Cytochem 60:861-873, 2012)

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  • Immunohistochemical analysis of two stem cell markers of alpha-smooth muscle actin and STRO-1 during wound healing of human dental pulp Reviewed

    Nagako Yoshiba, Kunihiko Yoshiba, Naoto Ohkura, Yoshimi Shigetani, Erika Takei, Akihiro Hosoya, Hiroaki Nakamura, Takashi Okiji

    HISTOCHEMISTRY AND CELL BIOLOGY   138 ( 4 )   583 - 592   2012.10

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    Recent studies have employed two markers, alpha-smooth muscle actin (alpha-SMA) and STRO-1, to detect cells with mesenchymal stem cell properties in dental pulp. The present study aimed to explore the expression profile of alpha-SMA and STRO-1 in intact dental pulp as well as during wound healing in adult dental pulp tissue. Healthy pulps were mechanically exposed and capped with the clinically used materials MTA (ProRoot White MTA) or Ca(OH)(2) to induce a mineralized barrier at the exposed surface. After 7-42 days, the teeth were extracted and processed for immunohistochemical analysis using antibodies against alpha-SMA, STRO-1 and nestin (a neurogenic cytoskeletal protein expressed in odontoblasts). In normal pulp, alpha-SMA was detected in vascular smooth muscle cells and pericytes. Double immunofluorescent staining with STRO-1 and alpha-SMA showed that STRO-1 was localized in vascular smooth muscle cells, pericytes and endothelial cells, in addition to nerve fibers. During the process of dental pulp healing, numerous alpha-SMA-positive cells emerged at the wound margin at 14 days, and the initially formed mineralized barrier was lined with alpha-SMA-positive cells similar in appearance to reparative odontoblasts, some of which co-expressed nestin. STRO-1 was abundant in nerve fibers. In the advanced stage of mineralized barrier formation at 42 days, cells lining the barrier were stained with nestin, and no staining of alpha-SMA was detected in those cells. These observations indicate that alpha-SMA-positive cells temporarily appear along the wound margin during the earlier phase of mineralized barrier formation and STRO-1 is confined in vascular and neuronal elements.

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  • Thy-1-positive cells in the subodontoblastic layer possess high potential to differentiate into hard tissue-forming cells Reviewed

    Akihiro Hosoya, Toru Hiraga, Tadashi Ninomiya, Akira Yukita, Kunihiko Yoshiba, Nagako Yoshiba, Masafumi Takahashi, Susumu Ito, Hiroaki Nakamura

    HISTOCHEMISTRY AND CELL BIOLOGY   137 ( 6 )   733 - 742   2012.6

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    The cells of the subodontoblastic cell-rich layer in dental pulp are speculated to contain odontoblast progenitor cells because of their positional relationship with odontoblasts as well as their high alkaline phosphatase (ALP) activity. However, it has yet to be determined whether these cells have the ability to differentiate into odontoblastic cells. In the present study, we firstly found that the majority of cells in the subodontoblastic layer expressed Thy-1, a cell-surface marker of stem and progenitor cells. Then, we evaluated the capacity of Thy-1 high- and low-expressing (Thy-1(high) and Thy-1(low)) cells separated from rat dental pulp cells by use of a fluorescence-activated cell sorter to differentiate into hard tissue-forming cells in vitro and in vivo. Following stimulation with bone morphogenetic protein-2, Thy-1(high) cells in vitro showed accelerated induction of ALP activity and formation of alizarin red-positive mineralized matrix compared with Thy-1(low) cells. Furthermore, subcutaneous implantation of Thy-1(high) cells efficiently induced the formation of bone-like matrix. These results collectively suggest that Thy-1-positive dental pulp cells localized in the subodontoblastic layer had the ability to differentiate into hard tissue-forming cells, and thus these cells may serve as a source of odontoblastic cells.

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  • Gene Expression Analysis of Membrane Transport Proteins in Normal and Lipopolysaccharide-inflamed Rat Dental Pulp Reviewed

    Naoto Ohkura, Yoshimi Shigetani, Nagako Yoshiba, Kunihiko Yoshiba, Takashi Okiji

    JOURNAL OF ENDODONTICS   38 ( 5 )   648 - 652   2012.5

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    Introduction: Membrane transport proteins (transporters) play a crucial role in the transmembrane uptake and/or efflux of various compounds such as inorganic ions, endogenous bioactive substances such as prostaglandins (PGs), and drugs such as nonsteroidal anti-inflammatory drugs. This study aimed to analyze mRNA expression of selected transporters related to drug disposition and PG transport in normal and lipopolysaccharide (LPS)-inflamed rat incisor pulp. Methods: Pulp tissues were subjected to reverse transcription-polymerase chain reaction (PCR) detection for transporter isoforms belonging to organic anion transporting polypeptide (Oatp), organic anion transporter (Oat), organic cation transporter (Oct), multidrug resistance-associated protein (Mrp), and multidrug resistance protein (Mdr) families. The levels of mRNA expression for PG transporters (Oatp1a5, Oatp1b2, Oatp2a1, Oatp2b1, and Oatp3a1) were compared in normal and LPS-inflamed pulps by using real-time PCR. Results: The pulp tissue expressed mRNAs for various transporters belonging to the Oatp, Oat, Oct, Mrp, and Mdr families. LPS inflammation caused significant up-regulation of Oatp2a1 (P &lt; .01) and significant down-regulation of Oatp1a5, Oatp2b1 (P &lt; .01), and Oatp3a1 (P &lt; .05). Conclusions: Rat incisor dental pulp expressed mRNAs for various transporter isoforms. The levels of mRNA expression for PG transporters were significantly up-regulated or down-regulated in LPS-inflamed dental pulp. (J Endod 2012;38:648-652)

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  • Expression of Angiogenic Factors in Rat Periapical Lesions Reviewed

    Yusuke Yamanaka, Tomoatsu Kaneko, Kunihiko Yoshiba, Reika Kaneko, Nagako Yoshiba, Yoshimi Shigetani, Jacques E. Noer, Takashi Okiji

    JOURNAL OF ENDODONTICS   38 ( 3 )   313 - 317   2012.3

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    Introduction: Angiogenic factors such. as VEGFR2 (vascular endothelial cell growth factor receptor 2), BcI-2 (a prosurvival and proangiogenic signaling molecule), and chernokine (C-X-C motif) ligand 1 (CXCL1) (a proangiogenic chemokine) may have critical roles in enhancing the establishment of apical periodontitis. To understand the role of these factors in the pathogenesis of apical periodontitis, we conducted immunohistochemical and molecular biological analysis. Methods: Apical periodontitis was induced in the lower first molars of Wistar rats by making unsealed pulp exposures. After, 14, 21, and 28 days, the molars were retrieved, embedded as frozen sample blocks, and cut in a cryostat. Normal lower first molars served as controls. lmmunostaining for CD31 (a marker for endothelial cells), BcI-2, and real-time polymerase chain reaction analysis of VEGFR2, BcI-2, CXCL1, and CXCR2 messenger RNA were performed. In the real-time polymerase chain reaction analysis, messenger RNA was extracted from CD31-stained endothelial cells that were retrieved with laser capture microdissection. For statistical analysis of immunohistochemistry, the immunostained area was plotted, and pixel counts were determined. Then, the percentage of the immunostained area in the total area was calculated. Results: The density of the CD31-stained area increased until 21 days after pulp exposure. On the other hand, BcI-2 stained area showed the highest density at 14 days (active lesion expanding phase) and then decreased until 28 days (lesion stability phase). VEGFR2, BcI-2, CXCL1, and CXCR2 messenger RNA expression in endothelial cells showed the highest levels at 14 days and then decreased until 28 days. Conclusions: The increase in microvascular density and the upregulation of VEGFR2, BcI-2, CXCL1, and CXCR2 messenger RNA expression in endothelial cells took place coincidently with the expanding phase of experi- mentally induced periapical lesions. These data suggest that these angiogenic factors play a role in the lesion development. (J Endod 2012;38:313-317)

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  • Expressional Alterations of Fibrillin-1 during Wound Healing of Human Dental Pulp Reviewed

    Nagako Yoshiba, Kunihiko Yoshiba, Naoto Ohkura, Akihiro Hosoya, Yoshimi Shigetani, Yusuke Yamanaka, Naoya Izumi, Hiroaki Nakamura, Takashi Okiji

    JOURNAL OF ENDODONTICS   38 ( 2 )   177 - 184   2012.2

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    Introduction: The degradation of fibrillins, the major constituents of microfibrils, is known to facilitate the release of active transforming growth factor-beta (TGF-beta), a signaling molecule contributing to mineralized tissue barrier formation in exposed dental pulps. To examine the involvement of fibrillins in the barrier formation, we examined the temporospatial expression of (1) genes and proteins of fibrillins and (2) factors possibly associated with fibrillin degradation and cyto- differentiation in exposed human pulps. Human pulp slice cultures were also examined for the role of fibrillins in mineralization. Methods: Clinically healthy pulps were mechanically exposed and capped with mineral trioxide aggregate. After 7 to 42 days, the teeth were processed for immunohistochemical and cytochemical staining of fibrillin-1, fibrillin-2, latent TGF-beta-binding protein (LTBP)-1, matrix metalloproteinase-3 (MMP-3), alkaline phosphatase (ALP), and in situ hybridization of fibrillin-1. Pulp tissue slices cultured with beta-glycero-phosphate were analyzed for fibrillin-1, fibrillin-2, and ALP with the immunohistochemical/cytochemical staining and quantitative reverse-transcriptase polymerase chain reaction. Results: Fibrillin-1-immunoreactivity was seen until 7 days but turned into undetectable since 14 days in the pulpal area just beneath the exposure site. MMP-3-immunoreaction was transiently detected at 14 days. At 42 days when the mineralized barrier was evident, fibrillin-1-immunoreactivity and fibrillin-1 expression remained down-regulated. Fibrillin-2, LTBP-1, and ALP were constantly detected in the fibrillin-1 undetectable area. Pulp slices cultured with beta-glycerophosphate showed mineralization with up-regulation of ALP and down-regulation of fibrillin-1. Conclusions: Degradation and down-regulation of fibrillin-1 expression took place during the mineralized tissue barrier formation in exposed pulps in vivo and beta-glycerophosphate-induced pulpal mineralization in vitro. (J Endod 2012;38:177-184)

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  • Mineral trioxide aggregates : Physicochemical and biological properties and their clinical implications

    Takashi OKIJI, Linlin HAN, Yoshimi SHIGETANI, Kunihiko YOSHIBA

    The Journal of Japan Endodontic Association   33 ( 1 )   3 - 13   2012

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    DOI: 10.20817/jeajournal.33.1_3

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  • Immunohistochemical analysis of subcutaneous tissue reactions to methacrylate resin-based root canal sealers Reviewed

    Y. Yamanaka, Y. Shigetani, K. Yoshiba, N. Yoshiba, T. Okiji

    INTERNATIONAL ENDODONTIC JOURNAL   44 ( 7 )   669 - 675   2011.7

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    P&gt;Aim
    To investigate subcutaneous tissue reactions to methacrylate resin-based root canal sealers by immunohistochemical assessment of inflammatory/immunocompetent cell infiltration.
    Methodology
    Silicone tubes containing freshly mixed Epiphany SE sealer, MetaSEAL, Super-Bond RC sealer, or a zinc oxide-eugenol sealer (Canals) were subcutaneously implanted into the backs of Wistar rats. Solid silicone rods implanted in different animals served as controls. After 7, 14 and 28 days, connective tissue surrounding the implants (n = 8, each) was processed for immunoperoxidase staining using OX6 (reactive to major histocompatibility complex class II molecules), ED1 (reactive to macrophages), and W3/13 (reactive primarily to neutrophils), and the number of positively stained cells within each field (1.2 x 0.8 mm) was enumerated. Statistical differences were analysed with Friedman&apos;s test and Scheffe&apos;s test (comparisons between test materials) or Mann-Whitney&apos;s U-test (test-control comparisons).
    Results
    Canals showed a significantly higher number of W3/13-positive cells (mostly neutrophils) than MetaSEAL at 28 days (P &lt; 0.05). There were no significant differences in the numbers of OX6- or ED1-positive cells between each test material at any time point. Test-control comparisons revealed several significant differences for each antibody. This was most notable for ED1, where all the test materials at each time point, except for Epiphany SE at 28 days, showed significantly larger values than the corresponding controls.
    Conclusions
    All the methacrylate resin-based sealers tested showed a similar level of inflammatory/immunocompetent cell infiltration. MetaSEAL induced less-intense neutrophil infiltration than Canals. Controls exhibited milder infiltration of inflammatory/immunocompetent cells compared with all the test materials.

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  • 半導体レーザー照射後のラット臼歯における非コラーゲンタンパクの遺伝子発現 Reviewed

    重谷 佳見, 大倉 直人, 吉羽 邦彦, 吉羽 永子, 大島 勇人, 興地 隆史

    日本歯科保存学雑誌   53 ( 5 )   495 - 501   2010.10

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    ラット臼歯へGaAlAs半導体レーザーを照射した後に,歯髄腔内に硬組織形成が生じることが知られている.そこで本研究では,RT-PCR法を用いて同レーザー照射後のラット臼歯における各種非コラーゲンタンパクのmRNA発現を検索した.生後8週齢Wistar系雄性ラットの上顎第一臼歯近心に,半導体レーザー装置(オサダライトサージ3000)を用いて,出力1.5W,60秒3回の条件でレーザー照射を行った.なお,非照射の上顎臼歯を対照とした.照射1,3,7日後に抜歯を行い,歯冠部よりmRNAを抽出した後,RT-PCR法にてosteo-pontin,osteonectin,osteocalcin,dentin sialophosphoprotein,dentin matrix protein 1,およびbone sialoproteinのmRNA発現解析を行った.また,歯髄の反応を組織学的に観察した.その結果,すべてのタンパクのmRNA発現が3日後までに増加を示すとともに,この発現亢進は7日後も持続していることが明らかになった.組織学的には,1〜3日後までは歯髄の局所的壊死が照射部近傍に観察されたが,7日後には象牙芽細胞様細胞の再配列と少量の新生硬組織が認められた.以上より,半導体レーザー照射されたラット臼歯では,新生硬組織形成に先立ち各種非コラーゲンタンパクのmRNA発現レベルの上昇が生じることが確認された.(著者抄録)

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  • Potential of periodontal ligament cells to regenerate alveolar bone Reviewed

    Akihiro Hosoya, Tadashi Ninomiya, Toru Hiraga, Kunihiko Yoshiba, Nagako Yoshiba, Etsuo Kasahara, Hidehiro Ozawa, Hiroaki Nakamura

    Journal of Oral Biosciences   52 ( 2 )   72 - 80   2010

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    Regeneration of periodontal tissues, lost as a result of periodontal disease, is a key objective of periodontal treatment. Although several periodontal regeneration therapies have been devised, the origin of the undifferentiated cells that regenerate periodontal tissues remains unknown. Therefore, in the present study, to clarify the existence of osteoblast progenitor cells in the periodontal ligament, as well as to investigate the mechanism of alveolar bone regeneration without any effects from the original bone, we evaluated osteoblast differentiation induced by transplantation of GFP-transgenic rat molars into the subcutaneous tissues of wild-type rats. Ten days after transplantation, initial alveolar bone was formed apart from the cementum in the bifurcation region. After 20 days, this bone tissue had expanded to almost all of the bifurcation. GFP localization showed that the osteoblasts were derived from the transplant. Alpha-SMA-and BMP4-positive cells were observed near the root surface at 5 days after transplantation. With the progress of alveolar bone regeneration, osteoblasts expressing Runx2 and Osterix appeared in the bone-forming region. These results indicate that periodontal ligament tissue remaining on the root surface after a tooth extraction contains undifferentiated cells that have the ability to regenerate alveolar bone. The process of osteoblast differentiation in this model might be similar to that for normal alveolar bone formation. Thus, periodontal ligament cells might be useful for the regeneration of alveolar bone in tissue engineering applications.

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  • Immunohistochemical analysis of nestin, osteopontin, and proliferating cells in the reparative process of exposed dental pulp capped with mineral trioxide aggregate Reviewed

    Momoko Kuratate, Kunibiko Yosbiba, Yosbimi Sbigetani, Nagako Yosbiba, Hayato Obsbima, Takashi Okiji

    JOURNAL OF ENDODONTICS   34 ( 8 )   970 - 974   2008.8

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    This study investigated the reparative process of mechanically exposed pulps capped with mineral trioxide aggregate (MTA). Maxillary first molars of 8-week-old rats were MTA-capped for 1-14 days, and 5-bromo-2'-deoxyuridine-labeled proliferating cells and immunoreactivity for nestin and osteopontin were analyzed. MTA capping caused mild necrotic changes followed by progressive new matrix formation and calcified bridging. Proliferating cells peaked at 3 days when matrix formation was inconspicuous. Nestin-expressing cells appeared at 3 days, were arranged beneath the newly formed matrix at 5 days, and showed odontoblast-like morphology by 14 days. Osteopontin immunoreactivity was detected just beneath the necrotic area after 1 day. These findings suggest that pulpal responses to MTA capping involve proliferation and migration of progenitors followed by their differentiation into odontoblast-like cells, a mechanism basically similar to those to calcium hydroxide. Osteopontin might play a triggering role in initiation of the pulpal reparative process.

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  • Alveolar bone regeneration of subcutaneously transplanted rat molar Reviewed

    Akihiro Hosoya, Tadashi Ninomiya, Toru Hiraga, Chen Zhao, Kunihiko Yoshiba, Nagako Yoshiba, Masafumi Takahashi, Takahiro Okabe, Shigeyuki Wakitani, Hirohito Yamada, Etsuo Kasahara, Hidehiro Ozawa, Hiroaki Nakamura

    BONE   42 ( 2 )   350 - 357   2008.2

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    Regeneration of alveolar bone is essential for periodontal treatment. Recently, cell replacement therapy has been focused on periodontal disease, but the source of the cells that regenerate alveolar bone is still uncertain. Therefore, to clarify the source of these bone-regenerating cells, we transplanted GFP-transgenic rat molars into the subcutaneous tissues of wild-type rats. Five days after transplantation, the tooth was surrounded by connective tissue containing many blood vessels. At 10 days, bone-like tissue was formed in the connective tissue between the branches of the bifurcated root. This hard tissue expanded to almost all of this bifurcation area without osseous ankylosis after 20 days. All ostcoblast-like cells in the newly formed matrix were immunopositive for GFP. In addition, these cells and the peripheral cells of the matrix showed intense immunoreactivity for BMP4, Runx2, BSP, and OPN. These results demonstrate that periodontal ligament tissue contains osteoprogenitor cells that have the ability to regenerate alveolar bone. Our model suggests that these regeneration processes might be similar to normal alveolar bone formation. (c) 2007 Elsevier Inc. All rights reserved.

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  • Association of TIMP-2 with extracellular matrix exposed to mechanical stress and its co-distribution with periostin during mouse mandible development Reviewed

    Nagako Yoshiba, Kunihiko Yoshiba, Akihiro Hosoya, Masahiro Saito, Takamasa Yokoi, Takashi Okiji, Norio Amizuka, Hidehiro Ozawa

    CELL AND TISSUE RESEARCH   330 ( 1 )   133 - 145   2007.10

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    Matrix remodeling is regulated by matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs). Periostin, originally identified in a mouse osteoblastic library, plays a role in cell adhesion and migration and in mechanical stress-induced matrix remodeling. In this study, we analyzed and compared the distribution patterns of TIMP-2 and periostin during mouse mandible development. Immunohistochemical staining for TIMP-2 and periostin was carried out on serial cryosections obtained from mice at embryonic days 13-16, postnatal day 2 (P2), P35, and 12 weeks of age. TIMP-2 and periostin exhibited a strikingly similar protein distribution during mandible development. From bud to early bell stages of molars, TIMP-2 and periostin were highly expressed on the lingual and anterior sides of the basement membrane and on the adjacent jaw mesenchyme. In pre- and postnatal incisors, the basement membrane of the apical loop and dental follicle was immunostained for TIMP-2 and periostin. At postnatal stages, TIMP-2 and periostin were prominently confined to the extracellular matrix (ECM) of gingival tissues, periodontal ligaments, and tendons (all recipients of mechanical strain). However, periostin was solely detected in the lower portion of the inner root sheath of hair follicles. Gingiva of P2 cultured in anti-TIMP-2 antibody-conditioned medium showed markedly reduced staining of periostin. We suggest that TIMP-2 and periostin are co-distributed on ECM exposed to mechanical forces and coordinately function as ECM modulators.

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  • Immunohistochemical study of hard tissue formation in the rat pulp cavity after tooth replantation Reviewed

    Chen Zhao, Akihiro Hosoya, Hiroshi Kurita, Tao Hu, Toru Hiraga, Tadashi Ninomiya, Kunihiko Yoshiba, Nagako Yoshiba, Masafumi Takahashi, Kenji Kurashina, Hidehiro Ozawa, Hiroaki Nakamura

    ARCHIVES OF ORAL BIOLOGY   52 ( 10 )   945 - 953   2007.10

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    While mineralized tissue is formed in the pulp cavity after tooth replantation or transplantation, little is known of this hard tissue formation. Therefore, we conducted histological and immunohistochemical evaluations of hard tissue formed in the pulp of rat maxillary molars after tooth replantation. At S days after replantation, degenerated odontoblasts were lining the pulp cavity. At 14 days, dentin- or bone-like tissue was present in the pulp cavity. Immunore activity for osteopontin (OPN) and bone sialoprotein (BSP) was strong in the bone-like tissue, but weak in the dentin-like tissue. Conversely, dentin sialoprotein (DSP) was localized in the dentin-like tissue, but not in the bone-like tissue. Cells positive for BMP4, Smad4, Runx2, and Osterix were found around the blood vessels of the root apex at 5 days. At 14 days, these cells were also localized around the bone-like tissue. Cells expressing alpha-smooth muscle actin (SMA) were seen around the newly formed bone-like tissue, whereas no such cells were found around the newly formed dentin-like tissue. In an experiment involving the transplantation of a green fluorescent protein (GFP)-transgenic rat tooth into a wild-type rat tooth socket, GFP-positive cells were detected on the surface of the bone-like tissue and over all dentin-like tissue. These results indicate that the original pulp cells had the ability to differentiate into osteoblast-like cells as well as into odontoblast-like cells. (C) 2007 Elsevier Ltd. All rights reserved.

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  • Hard tissue formation in subcutaneously transplanted rat dental pulp Reviewed

    A. Hosoya, H. Nakamura, T. Ninomiya, K. Hoshi, K. Yoshiba, N. Yoshiba, M. Takahashi, T. Okabe, N. Sahara, H. Yamada, E. Kasahara, H. Ozawa

    JOURNAL OF DENTAL RESEARCH   86 ( 5 )   469 - 474   2007.5

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    While dental pulp appears to be able to form mineralized matrices that do not always resemble dentin, the precise characteristics of the hard tissue and the mechanism of its induction remain unknown. Therefore, we evaluated hard tissue induced by transplantation of pulp into subcutaneous tissue. Seven days after transplantation, initial hard tissue was formed at the inner periphery of the pulp. After 14 days, this hard tissue expanded inwardly. Mineralized matrix was immunopositive for osteocalcin, osteopontin, and bone sialoprotein, but negative for dentin sialoprotein. Transplantation of GFP-labeled pulp into wild-type rats showed these formative cells to have been derived from the transplant. TEM observation revealed apatite crystals within necrotic cells and matrix vesicles at the initial stage of calcification. These results indicate that pulp cells possess the ability to form a bone- or cementum-like matrix. Calcification of the matrix may occur in necrotic cells and matrix vesicles, followed by collagenous calcification.

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  • Immunohistochemical localization of alpha-smooth muscle actin during rat molar tooth development Reviewed

    Akihiro Hosoya, Hiroaki Nakamura, Tadashi Ninomiya, Kunihiko Yoshiba, Nagako Yoshiba, Hiroyuki Nakaya, Shigeyuki Wakitani, Hirohito Yamada, Etsuo Kasahara, Hidehiro Ozawa

    JOURNAL OF HISTOCHEMISTRY & CYTOCHEMISTRY   54 ( 12 )   1371 - 1378   2006.12

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    The dental follicle contains mesenchymal cells that differentiate into osteoblasts, cementoblasts, and fibroblasts. However, the characteristics of these mesenchymal cells are still unknown. alpha-Smooth muscle actin (alpha-SMA) is known to localize in stem cells, and precursor cells of various tissues. In the present study, to characterize the undifferentiated cells in the dental follicle, immunohistochemical localization of a-SMA was examined during rat molar tooth development. Rat mandibles were collected at embryonic days (E) 15-20 and postnatal days (P) 7-28. Immunohistochemical stainings for alpha-SMA, periostin, Runt-related transcription factor-2 (Runx2), tissue nonspecific alkaline phosphatase (TNAP), and bone sialoprotein (BSP) were carried out using paraffin-embedded sections. a-SMA localization was hardly detected in the bud and cap stages. At the early bell stage, alpha-SMA-positive cells were visible in the dental follicle around the cervical loop. At the late bell to early root formation stage (P14), these cells were detected throughout the dental follicle, but they were confined to the apical root area at P28. Double immunostaining for alpha-SMA and periostin demonstrated that alpha-SMA-positive cells localized to the outer side of periostin-positive area. Runx2-positive cells were visible in the a-SMA-positive region. TNAP-positive cells in the dental follicle localized nearer to alveolar bone than Runx2-positive cells. BSP was detected in osteoblasts as well as in alveolar bone matrix. These results demonstrate that a-SMA-positive cells localize on the alveolar bone side of the dental follicle and may play a role in alveolar bone formation.

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  • Differential regulation of TIMP-1,-2, and-3 mRNA and protein expressions during mouse incisor development Reviewed

    N Yoshiba, K Yoshiba, C Stoetzel, F Perrin-Schmitt, Y Cam, JV Ruch, A Hosoya, H Ozawa, H Lesot

    CELL AND TISSUE RESEARCH   324 ( 1 )   97 - 104   2006.4

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    Tissue inhibitors of metalloproteinases (TIMPs) possess multiple functions, in addition to their matrix metalloproteinase (MMP) inhibitory activity. The continuously growing incisor of mouse possesses a stem cell compartment at the apical end of the epithelium (the apical loop) and thus provides an excellent tool to analyze the mechanisms of organogenesis and cytodifferentiation. To understand the functions of TIMPs in tooth development, we have analyzed the gene expression and protein localization of TIMP-1, -2, and -3 during mouse incisor development, from embryonic day 13 (E13) to postnatal day 3 (P3). TIMP-1 was present on the basement membrane during early developmental stages. At P2, TIMP-1 was strongly detected along the apical loop, transiently disappeared from the basement membrane in the cytodifferentiation zone, and later reappeared at the distal end of functional ameloblasts. Expression of TIMP-2 protein was restricted to the outer part of the apical loop throughout the examined stages. At P2, TIMP-2 was present on the basement membrane at the outer part of the apical loop. The dental follicle also expressed Timp-2, and the corresponding protein was abundant within the extracellular matrix. Timp-3 mRNA was highly expressed in the mesenchyme surrounding the apical loop. During matrix formation, Timp-3 was expressed by subodontoblasts, and the protein was detected in this layer and between odontoblasts. Distinct temporal and spatial expression patterns of TIMPs suggest divergent functions of these factors in incisor organogenesis.

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  • Odontoblast responses to GaAlAs laser irradiation in rat molars: an experimental study using heat-shock protein-25 immunohistochemistry Reviewed

    Y Tate, K Yoshiba, N Yoshiba, M Iwaku, T Okiji, H Ohshima

    EUROPEAN JOURNAL OF ORAL SCIENCES   114 ( 1 )   50 - 57   2006.2

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    Pulpal responses to gallium-aluminum-arsenide (GaAlAs) laser irradiation applied to the tooth remains to be elucidated. This study aimed to evaluate the effect of the GaAlAs laser on odontoblasts using immunohistochemistry for heat-shock protein (HSP)-25, which labels mature and newly differentiated odontoblasts. The mesial surface of the upper right first molar of 8-wk-old Wistar rats was lased at an output power of 0.5-1.5 W for 180 s. The animals were perfusion-fixed at intervals of 6 h to 30 d after irradiation. At 6 h to 7 d, the intensity of HSP-25-immunoreactivity was found to be disturbed in the coronal odontoblast-layer in an energy-dependent manner. At 30 d, tertiary dentin with/without bone-like tissue was formed abundantly in the dental pulp. Statistical analysis revealed that the area occupied by the new hard tissues was significantly wider in 1.5 W-lased specimens than in 0.5 W-lased specimens. An intense HSP-25 immunoreactivity was seen in the odontoblasts underlying the tertiary dentin, whereas immunoreactivity was weak around the bone-like tissue. It was concluded that the GaAlAs laser may induce the formation of tertiary dentin by influencing the secretory activity of odontoblasts. However, higher energies may cause irreversible changes to the pulp, often leading to the formation of an intrapulpal bone-like tissue.

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  • Immunohistochemical Study of Osteodentin in the Unerupted Rat Incisor Reviewed

    Akihiro Hosoya, Hiroaki Nakamura, Shoji Akahane, Kunihiko Yoshiba, Nagako Yoshiba, Tadashi Ninomiya, Kazuto Hoshi, Noriyuki Sahara, Etsuo Kasahara, Hidehiro Ozawa

    journal of oral biosciences   48 ( 2 )   132 - 137   2005

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    To characterize osteodentin in the pre-functional rat incisor, we performed histological and histochemical evaluation of the anterior apex of the incisor in 3-day-old rats. The unerupted incisor was composed of osteodentin, with numerous cells present in the anterior apex. The osteodentin was immunopositive for osteocalcin, bone sialoprotein, and dentin sialoprotein, with an immunolocalization pattern similar to that of dentin. Back-scattered electron microanalysis (BSE) and electron probe microanalysis (EPMA) showed that osteodentin was not uniformly calcified. These results indicate that osteodentin in the rat incisor possesses dentin-like characteristics, and may be fragile in structure. © 2006, Japanese Association for Oral Biology. All rights reserved.

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  • Temporospatial gene expression and protein localization of matrix metalloproteinases and their inhibitors during mouse molar tooth development Reviewed

    N Yoshiba, K Yoshiba, C Stoetzel, F Perrin-Schmitt, Y Cam, JV Ruch, H Lesot

    DEVELOPMENTAL DYNAMICS   228 ( 1 )   105 - 112   2003.9

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    The gene expression and protein distribution of matrix metalloproteinase (MMP) -2, -9, membrane type-1 MMP (MT1-MMP), as well as of TIMP-1, -2, and -3 were analyzed during mouse molar development. Immunohistochemical data demonstrated that all the MMPs investigated were expressed in the dental epithelium and mesenchyme. In contrast, gene and protein expression analysis for TIMPs showed that they were differentially expressed. TIMP-1 was expressed in the dental epithelium and mesenchyme between E13 and E16 and was transiently up-regulated at E14, the cap stage. TIMP-1 expression was also detected in differentiating odontoblasts. TIMP-2 RNA transcripts were found in the peridental and dental mesenchyme, odontoblasts, and ameloblasts. Protein analysis revealed high expression on the lingual side of the dental epithelium and underlying mesenchyme together with transient expression in the enamel knot at E14 and expression in the gingival tissue and enamel matrix postnatally. TIMP-3 RNA transcripts were found in discrete regions of the dental epithelium, including at high levels in the cervical loop at E16. Expression was also detected in preodontoblasts at E16 and transiently during ameloblast differentiation. Analysis of the protein distribution revealed a lower level of TIMP-3 on the lingual side of the dental epithelium at E14. MT1-MMP was expressed in the dental mesenchyme between E13 and E16, at relatively high levels in the cervical loop at E14, and in the odontoblasts and ameloblasts. The distinct temporospatial distribution patterns of the TIMPs suggest that these inhibitors play several intrinsic roles during tooth development. Developmental Dynamics 228.105-112, 2003. (C) 2003 Wiley-Liss, Inc.

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  • Class II antigen-presenting dendritic cell and nerve fiber responses to cavities, caries, or caries treatment in human teeth Reviewed

    K Yoshiba, N Yoshiba, M Iwaku

    JOURNAL OF DENTAL RESEARCH   82 ( 6 )   422 - 427   2003.6

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    Major histocompatibility complex (MHC) class 11 molecule-expressing cells are distributed in human dental pulp, and have been shown to accumulate beneath caries lesions. The responses of these cells and nerve fibers were analyzed under 5 different clinical conditions: shallow and deep experimental cavities, active and slow untreated caries, and treated caries. Under deep cavities, class 11 molecule-expressing dendritic cells displaced the injured odontoblasts during a period of one month, while such a response was not observed in shallow cavities and untreated or treated carious teeth. The class 11 molecules seen in the neural elements under active caries were no longer detectable in treated carious teeth. However, six months after treatment, clusters consisting of dendritic cells, Tlymphocytes, and nerve fibers still remained locally in the subodontoblastic area. These results indicate that dental pulps respond differently to cavity preparation and restoration between normal and caries conditions, and that immunoresponses persist for many months, even after caries treatment.

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  • An immunohistochemical study on hard tissue formation in a subcutaneously transplanted rat molar Reviewed

    A Hosoya, K Yoshiba, N Yoshiba, KT Hoshi, M Iwaku, H Ozawa

    HISTOCHEMISTRY AND CELL BIOLOGY   119 ( 1 )   27 - 35   2003.1

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    While dental pulp undergoes calcification following tooth replantation or transplantation, we actually know little about these mechanisms. We therefore conducted histological and immunohistochemical evaluations of mineralized tissue that formed in the pulp of rat maxillary molar transplanted into abdominal subcutaneous tissue. One, 2, 3, and 4 weeks post-transplantation, the teeth were investigated immunohistochemically using antibodies to osteocalcin (OCN), osteopontin (OPN), bone sialoprotein (BSP), dentin sialoprotein (DSP), and tissue non-specific alkaline phosphatase (TNAP). In the 1st week after transplantation, cell-rich hard tissue was formed at the root apex. At 2 weeks, formations of hard tissue, with few cells in the root canals and bone-like tissue in the coronal pulp chamber, were noted. After 3 and 4 weeks, the amounts of these hard tissues were increased. The immunolocalization of OCN, OPN, and BSP was seen strongly in coronal and apical hard tissues, but weakly in the root hard tissue. Conversely, DSP localized in the root hard tissue, but not in other newly formed hard tissues. At I week, TNAP localized along the periphery of the apical hard tissue and the lower surfaces of root predentin. These results demonstrate that the newly formed hard tissues in the pulp cavity of subcutaneously transplanted molars could be classified into three types, suggesting that these might be formed by type-specific cells.

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  • Odontoblast processes in human dentin revealed by fluorescence labeling and transmission electron microscopy Reviewed

    K Yoshiba, N Yoshiba, S Ejiri, M Iwaku, H Ozawa

    HISTOCHEMISTRY AND CELL BIOLOGY   118 ( 3 )   205 - 212   2002.9

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    In the present undertaking, the distribution of odontoblast processes in human dentin was determined through the DiI carbocyanine dye fluorescent staining of the cell membrane, while F-actin was identified by rhodamine-phalloidin. Confocal laser scanning microscopy revealed intense labeling for both agents in inner dentin, while transmission electron microscopy (TEM) identified dentinal tubules including odontoblast processes in this area, each process being surrounded by a cell membrane and containing an abundance of filamentous structures. Electron-dense "lamina limitans" lined the dentinal tubules. Individual cell processes became narrower toward the middle area, and their overall numbers decreased as well under TEM. Labeling for F-actin was absent in both middle and outer dentin, while faint labeling for DiI was visible along the dentinal tubules as far as the dentino-enamel junction (DEJ), where it was also recognized within the tubules themselves. Under TEM, the dentinal tubules lined with electron-dense structures were, in fact, empty in the middle and outer dentin. Immediately below the DEJ, however, the tubules manifested dense concentrations of fine granular material. Our study, therefore, appears to suggest that odontoblast processes do not extend beyond the inner dentin of fully erupted human premolars.

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  • Immunohistochemical study on pulpal response in rat molars after cavity preparation by Er : YAG laser Reviewed

    K Tanabe, K Yoshiba, N Yoshiba, M Iwaku, H Ozawa

    EUROPEAN JOURNAL OF ORAL SCIENCES   110 ( 3 )   237 - 245   2002.6

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    While Er:YAG laser systems are in extensive use for caries removal and cavity preparation, the effects of such treatment on pulp tissue remain unclear. This study evaluates these systems using immunohistochemical methods and compares the results with information gained from treatment using conventional burs. Cervical cavities were prepared in the upper first molars of rats, using either an Er:YAG laser or a conventional tungsten-carbide bur. At intervals of 5 min, 6 h, 12 h, 1 d, 3 d and 7 d after cavity preparation, the teeth were processed for immunohistochemical analyses of tissue non-specific alkaline phosphatase, OX6-positive major histocompatibility complex class II antigen-expressing cells and PGP 9.5-immunoreactive nerve fibers. DNA fragmentation was detected by the terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) method. Tissue non-specific alkaline phosphatase was observed mainly in the subodontoblastic layer under the cavity lesion, from 5 min, in both groups. The inumunoreactivity was more pronounced in the laser group, but by 7 d no significant differences were recognizable. At 12 h, TUNEL-positive cells were detected around the odontoblastic layer in both groups. From 3 d to 7 d, a limited number of positive cells were still visible in the group that underwent standard treatment. Clear similarities in the distribution patterns of OX6-immunopositive cells and PGP 9.5-immunoreactive nerve fibers were also noted. From 12 h to 1 d, OX6-positive cells accumulated along the pulp-dentin border, extending their processes into the dentinal tubules. Numerous bead-like PGP 9.5-immunoreactive nerve fibers were observed under the odontoblastic layer at 7 d. These results demonstrated that there was no appreciable difference in the manner in which pulp tissue responded to treatment with either Er:YAG laser or a conventional drill. This would seem to indicate the usefulness of the Er:YAG laser system in the removal of caries and cavity preparation.

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  • Distribution of noncollagenous proteins during reparative dentinogenesis in human teeth Reviewed

    K Yoshiba, N Yoshiba, M Iwaku, H Ozawa

    DENTIN/PULP COMPLEX   157 - 158   2002

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    It has been well-documented that the exposed dental pulp possesses the ability to form a hard tissue barrier (dentin bridge). However, the exact mechanisms by which pulp cells differentiate into odontoblasts in this process are still unknown. During primary dentinogenesis, odontoblasts express a number of mineral-related noncollagenous proteins, including bone sialoprotein (BSP), osteopontin (OPN), and dentin sialoprotein (DSP), a dentin-specific protein. The purpose of this study was to analyze immunohistochemically the expression of these molecules during reparative dentinogenesis in human teeth after pulp was capped with calcium hydroxide [Ca(OH)(2)]. At 14 days, a thin layer of fibrous matrix barrier was formed at the exposure site. An alignment of cells was observed adjacent to the barrier, and some cells were weakly immunopositive for BSP, OPN, and DSP. At 28 days, the hard tissue barrier was thickened and pulpally lined with elongated odontoblast-like cells that expressed BSP, OPN, and DSP. A number of globular structures intensely stained for BSP and OPN were observed between the cells. The superficial layer of the barrier was immunopositive for BSP and DSP, whereas OPN was found entirely in the matrix. These results suggest that BSP, OPN, and DSP are involved in reparative dentinogenesis after pulp capping with Ca(OH)(2) and that cells that form reparative dentin express an odontoblastic phenotype.

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  • Hard tissue formation in subcutaneously transplanted rat molars Reviewed

    A Hosoya, K Yoshiba, N Yoshiba, M Iwaku, K Hoshi, H Ozawa

    DENTIN/PULP COMPLEX   166 - 167   2002

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    The purpose of this study was to elucidate the pulpal calcification mechanism in the pulp cavity of subcutaneously transplanted rat molars. First and second maxillary molars of 4-week-old Wistar rats were extracted and immediately transplanted into abdominal subcutaneous connective tissue. At 1, 2, 3, and 4 weeks after transplantation, transplanted teeth were excised with the surrounding tissue, fixed, decalcified, embedded in paraffin, and sectioned. They were investigated by staining with hematoxylin-eosin and immunohistochemistry using antibodies to osteocalcin (OCN), osteopontin (OPN), bone sialoprotein (BSP), dentin sialoprotein (DSP), and tissue-nonspecific alkaline phosphatase (TNAP). One week after transplantation, coronal pulp tissue underwent degeneration, but blood vessels were noted to have entered the root canal from the apex. Some flat cells were found adjacent to the root predentin. Hard tissue including numerous cells was formed adjacent to the apical predentin toward the pulp cavity. At 2 weeks, blood vessels entered the pulp chamber, and bone-like tissue was formed apart from the predentin. In the root canal, hard tissue having few cells was formed adjacent to the predentin. After 3 weeks, the amounts of these hard tissues were significantly increased. In these hard tissues, OCN, OPN, and BSP were localized strongly in the coronal and apical hard tissues but weakly in the root hard tissue. On the other hand, DSP was localized in the root hard tissue but did not localize in the other newly formed hard tissues. TNAP localized strongly in the periphery of the apical hard tissue and the surface of the lower root predentin at I week, but uniform localization was observed in the soft tissue in the whole pulp cavity after 3 weeks. These findings demonstrate that the newly formed hard tissues in the pulp cavity of rat molars after transplantation into subcutaneous tissue could be classified into three types by histologic and immunohistochemical characteristics, and suggest that these might be formed by cells of three different types.

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  • Differential expression of laminin-5 subunits during incisor and molar development in the mouse Reviewed

    K Yoshiba, N Yoshiba, D Aberdam, G Meneguzzi, F Perrin-Schmitt, C Stoetzel, JV Ruch, H Lesot

    INTERNATIONAL JOURNAL OF DEVELOPMENTAL BIOLOGY   44 ( 3 )   337 - 340   2000.4

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    Rodent incisors are continuously growing teeth and enamel deposition is restricted to the labial side. In the present study, the expression of laminin-5 subunits (alpha 3, beta 3 and gamma 2) has been analyzed by in site hybridization in developing mouse lower incisors and compared to that reported in the molar. At the bud stage (E12), mRNAs for all subunits were detected in the whole epithelial thickening. At E14,when histogenesis had started,transcripts for a alpha 3 and gamma 2 subunits were restricted to the outer dental epithelium (ODE), whereas the beta 3 subunit was intensely expressed in the inner dental epithelium (IDE). A transient expression for alpha 3 subunit was seen in the enamel knot area and disappeared at E15. Subsequently, all laminin-5 subunit genes were re-expressed in differentiating ameloblasts on the labial side. Similar patterns of transcription were observed in incisor and molar, suggesting that the differential expression of laminin-5 subunits in the IDE might be involved in the histogenesis of the IDE and ameloblast differentiation. At E16.5, cells of the IDE at the anterior extremity of the incisor and in the anterior part of the lingual IDE expressed transcripts for a3 and beta 3 but not for gamma 2 subunit. Similar expression patterns were observed in the enamel-free areas of the E18 molar. This specific expression might thus be related to cells that do not differentiate as functional ameloblasts. Throughout incisor development, intense expression for all laminin-5 subunits was restricted to the labial side of the cervical loop. The asymmetrical expression of laminin-5 might be related to incisor morphogenesis and to the differences in histogenesis and cytodifferentiation of the IDE that exist in the labial versus lingual aspect of the cervical loop.

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  • Immunohistochemical localization of laminin-5 and integrin A6 beta 4 during cytodifferentiation of ameloblasts in the mouse incisor Reviewed

    K Yoshiba, N Yoshiba, M Iwaku, H Ozawa

    JOURNAL OF INVESTIGATIVE DERMATOLOGY   113 ( 6 )   1143 - 1143   1999.12

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  • Immunohistochemical localizations of class II antigens and nerve fibers in human carious teeth: HLA-DR immunoreactivity in Schwann cells Reviewed

    N Yoshiba, K Yoshiba, M Iwaku, H Ozawa

    ARCHIVES OF HISTOLOGY AND CYTOLOGY   61 ( 4 )   343 - 352   1998.10

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    Nerve fibers and class II major histocompatibility complex (MHC) antigen-expressing dendritic cells have been known to gather in the dental pulp beneath carious lesions. Significant functional interactions presumably occur between the neural and immune elements. The present study analyzed the morphological relationship between class II-expressing cells and nerve fibers in fuman carious teeth, visualized by a HLA-DR monoclonal antibody and a protein gene product 9.5 (PGP 9.5) polyclonal antibody; a confocal laser scanning microscope (CLSM) and an electron microscope were used. In pulps affected by early caries, HLA-DR-positive dendritic cells aggregated mainly in the cell-free zone associated with bundles of PGP 9.5-immunoreactive nerve fibers, In pulps affected by advanced caries, the accumulated HLA-DR-positive cells and PGP 9.5-immunoreactive nerve fibers showed close association with each other especially beneath the odontoblast layer: the cells even embraced the nerve fibers. Intriguingly, class II molecules were recognized not only in dendritic cells but also in the Schwann cells of non-myelinated nerves in the pulp. Using immunoelectron microscopy, class II molecules were localized on the surface of the non-myelinating Schwann cells and also within some vesicles, whereas myelinating Schwann cells lacked this immunoreactivity. PGP 9.5-immunoreactive nerve fibers were also observed densely in the odontoblast layer, and CLSM revealed an intimate association of the nerve fibers and dendritic cells. The immunoreactivity for HLA-DR in Schwann cells depended upon the severity of the carious lesion. Class II-expressing Schwann cells are suggested to function as antigen-presenting cells in addition to dendritic cells.

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  • Expression and localization of laminin-5 subunits in the mouse incisor Reviewed

    N Yoshiba, K Yoshiba, D Aberdam, G Meneguzzi, F Perrin-Schmitt, C Stoetzel, JV Ruch, H Lesot

    CELL AND TISSUE RESEARCH   292 ( 1 )   143 - 149   1998.4

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    Laminin-5 is associated with several epithelial tissues and forms part of the anchoring filaments of hemidesmosomes. Recent data have shown that the expression of laminin-5 subunits is impaired in junctional epidermolysis bullosa (JEB), and, in these patients, enamel hypoplasia is commonly observed. Rodent incisors are continuously growing teeth with an asymmetry between their labial and lingual sides. Enamel matrix formation is restricted to the labial side. We have analyzed the changes in the expression and localization of laminin-5 subunits (alpha 3, beta 3, and gamma 2) in lower incisors of the mouse. The apical loop located at the end of the labial side contained stem cells and showed expression for all laminin-5 subunits. In the anterior direction, the inner dental epithelial cells (LDE) transiently lost the immunoreactivity for all subunits, whereas the transcripts for the beta 3 subunit remained in the IDE. All subunit mRNAs and proteins were expressed in ameloblasts facing predentine and also in secretory and maturation stage ameloblasts. Enamel matrix contained laminin-5. On the lingual side, the expression of laminin-5 subunits was continuous from the epithelial root sheath to the epithelial rests of Malassez in the periodontal ligament. These results suggest that spatial and temporal regulation of laminin-5 subunits correlates with the histogenesis of the dental organ, ameloblast differentiation, and enamel formation and also that laminin-5 plays a role in the adhesion between dental epithelial cells and the extracellular matrix (enamel or dentine) in areas where the dental basement membrane is absent.

    DOI: 10.1007/s004410051044

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  • Expression and localization of laminin-5 subunits during mouse tooth development Reviewed

    K Yoshiba, N Yoshiba, D Aberdam, G Meneguzzi, F Perrin-Schmitt, C Stoetzel, JV Ruch, H Lesot

    DEVELOPMENTAL DYNAMICS   211 ( 2 )   164 - 176   1998.2

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    Tooth morphogenesis is regulated by epithelial-mesenchymal interactions mediated by the easement membrane (BM). Laminins are major glycoprotein components of the BMs, which are involved in several cellular activities. The expression and localization of the alpha 3, beta 3, and gamma 2 laminin-5 subunits have been analyzed by in situ hybridization and immunohistochemistry during mouse molar development. Initially (E12), mRNAs of all subunits were detected in the entire dental epithelium and the corresponding proteins were located in the BM. During cap formation (E13-14), transcripts for the alpha 3 and gamma 2 subunits were localized in the outer dental epithelium (ODE), whereas the beta 3 subunit mRNA was present in the inner dental epithelium (IDE). During the early bell stage (E16), immunoreactivity for all subunits disappeared from the BM along the IDE, although intense signals for beta 3 mRNA were detectable in cells of the IDE, Subsequently, when the dentinal matrix was secreted by odontoblasts (E18-19.5), mRNAs of all three subunits were re-expressed by ameloblasts, and the corresponding proteins were detected in ameloblasts and in the enamel matrix. Tissue recombination experiments demonstrated that when E16 IDE or ODE was associated with E18 dental papilla mesenchyme, immunostaining for all laminin-5 subunits disappeared from the BM, whereas when cultured with non-dental limb bud mesenchyme, they remained positive after 48 hr of culture. These results suggest that the temporospatial expression of laminin-5 subunits in tooth development, which appears to be differentially controlled by the dental mesenchyme, might be related to the enamel organ histo-morphogenesis and the ameloblast differentiation. (C) 1998 Wiley-Liss, Inc.

    DOI: 10.1002/(SICI)1097-0177(199802)211:2<164::AID-AJA5>3.0.CO;2-F

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  • Immunohistochemical localization of HLA-DR-positive cells in unerupted and erupted normal and carious human teeth Reviewed

    N Yoshiba, K Yoshiba, H Nakamura, M Iwaku, H Ozawa

    JOURNAL OF DENTAL RESEARCH   75 ( 8 )   1585 - 1589   1996.8

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    Class II major histocompatibility complex (MHC) antigen-expressing cells are generally associated with the early phase of the immune response. We have studied the distribution of class II-expressing cells in developing, normal, and carious human teeth to clarify when human pulp acquires an immunologic defense potential and how this reacts to dental caries. Antigen-expressing cells were identified immunohistochemically by means of HLA-DR monoclonal antibody. In the pulp of unerupted developing teeth, numerous HLA-DR-positive cells were distributed mainly in and around the odontoblast layer. In erupted teeth, HLA-DR-positive cells were located, for the most part, just beneath the odontoblast layer, with slender cytoplasmic processes extending into the layer. Superficial caries lesions caused an aggregation of HLA-DR-positive cells in dental pulp corresponding to the lesion. In teeth with deeper caries lesions, this aggregation of cells expanded to include the odontoblast layer. Also noted were HLA-DR-positive cells lying along the pulp-dentin border, with cytoplasmic processes projecting deep into the dentinal tubules, where they co-localized with odontoblast processes. These findings suggest that: (1) human dental pulp is equipped with immunologic defense potential prior to eruption; (2) in the initial stage of caries infection, an immunoresponse mediated by class-II-expressing cells is initiated in human dental pulp; and (3) HLA-DR-positive cells trespass deep into dentinal tubules as the caries lesion advances.

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  • Immunolocalization of fibronectin during reparative dentinogenesis in human teeth after pulp capping with calcium hydroxide Reviewed

    K Yoshiba, N Yoshiba, H Nakamura, M Iwaku, H Ozawa

    JOURNAL OF DENTAL RESEARCH   75 ( 8 )   1590 - 1597   1996.8

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    Exposed dental pulp is known to possess the ability to form a hard-tissue barrier (dentin bridge). The exact mechanisms by which pulp cells differentiate into odontoblasts in this process are unknown. Fibronectin has been demonstrated to play a crucial role in odontoblast differentiation during tooth development. This study tested the hypothesis that fibronectin is involved in the initial stages of replacement odontoblast differentiation and reparative dentin formation. We observed its immunohistochemical localization during dentin bridge formation in human teeth, after pulp was capped with calcium hydroxide [Ca(OH)(2)]. One day after the capping, precipitation of crystalline structures was observed at the TEM level in association with cell debris at the interface between the superficial necrotic zone and underlying pulp tissue. This layer of dystrophic calcification showed positive reaction for fibronectin, and pulp cells appeared to be closely associated with this layer, seven to ten days post-operatively. At 14 days, an alignment of cells, some of which were elongated and odontoblast-like, was observed adjacent to the fibronectin-positive irregular matrix. Between the cells, corkscrew fiber-like fluorescence was visible. At 28 days, the irregular fibrous matrix was followed by the formation of tubular dentin-like matrix lined with odontoblast-like cells. Therefore, it would seem that fibronectin associated with the initially formed calcified layer might play a mediating role in the differentiation of pulp cells into odontoblasts during reparative dentinogenesis, after pulp was capped with Ca(OH)(2).

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  • Immunolocalization of the small proteoglycan decorin in human teeth Reviewed

    N Yoshiba, K Yoshiba, M Iwaku, H Ozawa

    ARCHIVES OF ORAL BIOLOGY   41 ( 4 )   351 - 357   1996.4

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    The immunolocalization of decorin was studied by confocal laser scanning microscopy and transmission electron microscopy. In the apical area of developing teeth, labelling for decorin was found in the dental papilla cells, preodontoblasts and also in the Hertwig's epithelial cells. Mantle dentine and the initial predentine were negative. In circumpulpal dentine, intense reactivity extended along the calcification front and dentinal tubules. Fluorescence was also evident in odontoblast cell bodies and their processes in predentine. None was perceived, however, in the predentinal matrix. Faint staining was observed on the calcified dentinal matrix. Immunoelectron microscopy revealed staining for decorin in collagen fibrils lining the predentine-dentine junction, and where arrays of labelled filaments were noted orthogonal to the collagen fibrils. Staining extending from the calcification front was observed in the matrix adjacent to the dentinal tubule. The decorin observed at the calcification front might regulate the mineralization of dentinal matrix. Copyright (C) 1996 Elsevier Science Ltd.

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  • Distribution of the small proteoglycan decorin in human teeth Reviewed

    N Yoshiba, K Yoshiba, M Iwaku, H Ozawa

    DENTIN/PULP COMPLEX   304 - 305   1996

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    The immune-localization of decorin in both normal and carious human teeth was studied using confocal laser scanning microscope and transmission electron microscope. In normal teeth, labeling for decorin was observed in odontoblast cell bodies and its processes located in predentin. No reaction was observed in predentinal matrix, however, fluorescence was intense along the calcification front and dentinal tubules. Odontoblasts under carious lesions reacted positively to decorin, while, the dentinal tubules affected by caries did not. Immunoelectron microscopy revealed staining for decorin in collagen fibrils lining the predentin-dentin junction. Decorin might regulate the mineralization of dentinal matrix.

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  • IMMUNOELECTRONMICROSCOPIC STUDY OF THE LOCALIZATION OF FIBRONECTIN IN THE ODONTOBLAST LAYER OF HUMAN TEETH Reviewed

    N YOSHIBA, K YOSHIBA, H NAKAMURA, M IWAKU, H OZAWA

    ARCHIVES OF ORAL BIOLOGY   40 ( 2 )   83 - 89   1995.2

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    Indirect immunofluorescence-based studies have shown similarities in the distribution patterns of fibronectin-positive fibrous structures and so-called von Korff fibres. The aim of the present study was to analyse the reactivity of fibronectin in the odontoblast layer of fully developed human teeth by means of immunoelectron microscopy. Between the odontoblasts, discrete and undulatory fibrillar fascicles with peroxidase labelling were observed. They seemed to be in contact with odontoblasts in some areas, while in others they appeared to be intervening between two neighbouring odontoblasts. Higher magnifications of the fibrillar material demonstrated axial periodic staining of about 70 nm. Peroxidase reaction of fibronectin was also recognized along the cell membrane of odontoblasts facing predentine. The fibronectin in fibrillar fascicles observed between odontoblasts would be held in place by the direct molecular interaction with collagen fibrils and contribute to the pulpward migration of these cells and maintenance of their specific morphology. At the distal end of odontoblasts, a tight seal would be maintained by means of odontoblast-fibronectin adhesion.

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  • EFFECTS OF ANTIBACTERIAL CAPPING AGENTS ON DENTAL PULPS OF MONKEYS MECHANICALLY EXPOSED TO ORAL MICROFLORA Reviewed

    K YOSHIBA, N YOSHIBA, M IWAKU

    JOURNAL OF ENDODONTICS   21 ( 1 )   16 - 20   1995.1

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    The effects of antibacterial drugs on bacterially contaminated dental pulps were investigated in monkeys. Class V buccal cavities with pulpal exposures were prepared and then left open to the oral environment for 24 h. The exposed pulps were capped with a-tricalcium phosphate (alpha-TCP) containing a mixture of antibacterial drugs. Either alpha-TCP or Ca(OH)(2) was used as a control. Pulpal responses were histologically evaluated after 4 wk. Those teeth capped with alpha-TCP alone showed total pulp necrosis and bacterial growth within the pulp chamber. By contrast, the pulps capped with alpha-TCP containing mixed antibacterial drugs remained almost normal without any necrotic layer, but showed persistent absorbing response to capping materials and no signs of hard tissue barrier formation. In teeth capped with Ca(OH)(2), a hard tissue barrier was formed below the exposure site, with a wide loss of pulp tissue. No inflammation was seen under the barrier. These results indicate that mixed antibacterial drugs added to alpha-TCP effectively disinfected pulpal lesions, without destroying any of the sound pulp tissue. However, hard tissue barrier formation was delayed by this mixture as compared with Ca(OH)(2).

    DOI: 10.1016/S0099-2399(06)80551-0

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  • HISTOLOGICAL OBSERVATIONS OF HARD-TISSUE BARRIER FORMATION IN AMPUTATED DENTAL-PULP CAPPED WITH ALPHA-TRICALCIUM PHOSPHATE-CONTAINING CALCIUM HYDROXIDE Reviewed

    K YOSHIBA, N YOSHIBA, M IWAKU

    ENDODONTICS & DENTAL TRAUMATOLOGY   10 ( 3 )   113 - 120   1994.6

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    This study was performed to evaluate the pulpal response to alpha-tricalcium phosphate (alphaTCP) containing calcium hydroxide (Ca(OH)2). The dental pulps of monkeys were amputated and dressed with four agents: alphaTCP, alphaTCP containing 1% Ca(OH)2, alphaTCP containing 5% Ca(OH)2, and Ca(OH)2 being used as a control. The pulpal responses were histologically evaluated after 4 and 8 weeks. The pulp tissue treated with alphaTCP proliferated above the level of the original wound surface, and a thin layer of hard tissue barrier was formed directly against the capping agent. The barrier demonstrated atubular matrix lined with flattened or cuboidal cells, but occasionally appeared irregular in form. Ca(OH), dressing resulted in destruction of pulp tissue, with a thick hard tissue barrier being formed below the level of the exposure site. The barrier consisted coronally of osteodentin and pulpally of tubular dentin lined with odontoblast-like cells. By contrast, 1% Ca(OH), added to alphaTCP produced a slight proliferation of pulp tissue. An atubular matrix barrier, pulpally lined with cuboidal cells, formed above the exposure site. It was later followed by the formation of tubular matrix lined with columnar cells. Teeth treated with 5% Ca(OH), showed a thin necrotic layer and a thick barrier formation. The barrier was composed of tubular dentin-like tissue lined with odontoblast-like cells. It would appear that alphaTCP containing a small amount of Ca(OH)2 may be clinically useful as capping agent, as it induced consistent hard tissue formation, without excessive destruction of underlying pulp tissue.

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  • A CONFOCAL LASER-SCANNING MICROSCOPIC STUDY OF THE IMMUNOFLUORESCENT LOCALIZATION OF FIBRONECTIN IN THE ODONTOBLAST LAYER OF HUMAN TEETH Reviewed

    N YOSHIBA, K YOSHIBA, M IWAKU, H NAKAMURA, H OZAWA

    ARCHIVES OF ORAL BIOLOGY   39 ( 5 )   395 - 400   1994.5

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    The distribution of fibronectin in dental pulp was studied in developing and developed human teeth by indirect immunofluorescence using a confocal laser scanning microscope. In the apical region of developing teeth, intense fluorescence was found along the basement membrane facing the mesenchyme of Hertwig's epithelial sheath and first-formed (mantle) predentine. With further elongation of odontoblasts, fibronectin was observed between the cells, appearing as corkscrew fibres passing from the pulp into predentine parallel to the long axis of the odontoblasts. In the coronal region of developing and developed teeth a similar distribution of fibronectin was observed in the odontoblast layer. At the border zone between odontoblasts and predentine the reaction was intense, but was weak in the predentine itself. In the calcified dentinal matrix it had disappeared completely, except for the area along the dentinal tubules. The results demonstrate that fibronectin is present in the odontoblast layer during all stages of dentinogenesis. Fibronectin-positive fibrous structures between odontoblasts probably correspond to von Korff fibres, and are closely related to odontoblast differentiation and dentinogenesis.

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  • Histological observations of hard tissue barrier formation in amputated dental pulp capped with α‐tricalcium phosphate containing calcium hydroxide Reviewed

    K. Yoshiba, N. Yoshiba, M. Iwaku

    Dental Traumatology   10 ( 3 )   113 - 120   1994

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    Abstract This study was performed to evaluate the pulpal response to α‐tricalcium phosphate (αTCP) containing calcium hydroxide (Ca(OH)2). The dental pulps of monkeys were amputated and dressed with four agents: αTCP, αTCP containing 1% Ca(OH)2, αTCP containing 5% Ca(OH)2, and Ca(OH)2 being used as a control. The pulpal responses were histologically evaluated after 4 and 8 weeks. The pulp tissue treated with αTCP proliferated above the level of the original wound surface, and a thin layer of hard tissue barrier was formed directly against the capping agent. The barrier demonstrated atubular matrix lined with flattened or cuboidal cells, but occasionally appeared irregular in form. Ca(OH)2 dressing resulted in destruction of pulp tissue, with a thick hard tissue barrier being formed below the level of the exposure site. The barrier consisted coronally of osteodentin and pulpally of tubular dentin lined with odontoblast‐like cells. By contrast, 1% Ca(OH)2 added to αTCP produced a slight proliferation of pulp tissue. An atubular matrix barrier, pulpally lined with cuboidal cells, formed above the exposure site. It was later followed by the formation of tubular matrix lined with columnar cells. Teeth treated with 5% Ca(OH)2, showed a thin necrotic layer and a thick barrier formation. The barrier was composed of tubular dentin‐like tissue lined with odontoblast‐like cells. It would appear that αTCP containing a small amount of Ca(OH)2 may be clinically useful as a capping agent, as it induced consistent hard tissue formation, without excessive destruction of underlying pulp tissue. Copyright © 1994, Wiley Blackwell. All rights reserved

    DOI: 10.1111/j.1600-9657.1994.tb00535.x

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Books

  • Distribution of noncollagenous proteins during reparative dentinogenesis in human teeth.

    Dentin/Pulp Comples:Proceeding of the International Conference on Dentin/Pulp Complex 2001.  2002 

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  • Distribution of concollagenous proteins during reparative dentinogensis in human teeth.

    Dentin/Pulp Comples:Proceeding of the International Conference on Dentin/Pulp Complex 2001.  2002 

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  • Distribution of odontoblast processes in human coronal dentin. (共著)

    Dentin/Pulp Complex : Proceedings of the International Confernce on Dentin/Pulp Complex 1995. Quintessence Publishing Co.  1996 

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  • Distribution of the small proteoglycan decorin in human teeth. (共著)

    Dentin/Pulp Complex : Proceedings of the International Conference on Dentin/Pulp Complex 1995. Quintessence Publishing Co.  1996 

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  • Hard tissue formation in subcutaneously transplanted rat molars.

    Dentin/Pulp Complex:Proceeding of the International Conference on Dentin/Pulp Complex 2001. Quintessence Publishing Co. 

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  • Hard tissue formation in subcutaneously transplanted rat molars.

    Dentin/Pulp Comples:Proceeding of the International Conference on Dentin/Pulp Complex 2001.Quintessence Publishing Co. 

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MISC

  • Impact of the tissue nonspecific alkaline phosphatase during the root formation

    大倉直人, 大倉直人, 吉羽永子, 高原信太郎, GUTIERREZ Rosa Edith Baldeon, GOMEZ-KASIMOTO Susan, 井田貴子, 枝並直樹, 竹中彰治, 吉羽邦彦, 野杁由一郎

    日本歯科保存学会学術大会プログラムおよび講演抄録集(Web)   159th   2023

  • The comparison analyses of wound-healing mechanism according to the root development stage using the regeneration model rats

    高原信太郎, 大倉直人, 吉羽邦彦, 吉羽永子, 竹中彰治, 枝並直樹, 庭野和明, 野杁由一郎

    日本歯科保存学会学術大会プログラムおよび講演抄録集(Web)   158th   2023

  • The localization of various mesenchymal stem cell marker-positive cells in early healing stage after regenerative endodontic procedure

    高原信太郎, 大倉直人, 吉羽永子, 竹中彰治, 枝並直樹, 吉羽邦彦, 野杁由一郎

    日本歯科保存学会学術大会プログラムおよび講演抄録集(Web)   159th   2023

  • Interactions between Apically-extruded Bioceramic Sealers and periodontal tissues

    高原信太郎, 枝並直樹, 竹中彰治, 吉羽邦彦, 大倉直人, 吉羽永子, 野杁由一郎

    日本歯科保存学会学術大会プログラムおよび講演抄録集(Web)   157th   2022

  • Comparison of Biomineralization Ability of Calcium Silicate-based and Calcium Hydroxide-based Intracanal Medicaments

    枝並直樹, 竹中彰治, 吉羽邦彦, 大倉直人, 吉羽永子, 高原信太郎, 野杁由一郎

    日本歯科保存学会学術大会プログラムおよび講演抄録集(Web)   157th   2022

  • ヒト歯髄においてシュワン細胞はマクロファージをM2型へ転換する

    吉羽永子, 大倉直人, 前川知樹, 泉健次, 細矢明宏, 中村浩彰, 前田健康, 野杁由一郎, 吉羽邦彦

    Journal of Oral Biosciences Supplement (Web)   2019   2019

  • 歯髄創傷治癒モデルラットを用いたグルコース輸送担体Glut2とGlut4の局在および遺伝子発現の解析

    遠間 愛子, 大倉 直人, 枝並 直樹, 竹内 亮祐, 吉羽 永子, 吉羽 邦彦, 野杁 由一郎

    新潟歯学会雑誌   48 ( 2 )   114 - 114   2018.12

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  • 歯髄創傷モデルラットを用いた修復象牙質形成時におけるGlut1-Runx2関連の解析

    竹内 亮祐, 大倉 直人, 枝並 直樹, 遠間 愛子, 吉羽 永子, 吉羽 邦彦, 野杁 由一郎

    新潟歯学会雑誌   48 ( 2 )   113 - 114   2018.12

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  • マイクロスコープを用いた再歯根尖切除術の1例

    大倉 直人, 山本 信一, 阿部 達也, 竹内 亮祐, 遠間 愛子, 枝並 直樹, 吉羽 永子, 吉羽 邦彦, 野杁 由一郎

    新潟歯学会雑誌   48 ( 1 )   29 - 35   2018.6

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    根尖性歯周組織病変の難治化は、根尖部での複雑な解剖学的問題に加え、残存バイオフィルムによる細菌学的な影響もあるとされている。こうした様々な要因の改善策として外科的歯内療法が選択される。しかし、補綴処置を行う上で考慮する歯冠・歯根長比の問題で複数回に渡り歯根切断が行われることは稀である。今回は、歯根尖切除術を過去3度に渡って施行しても治癒しなかった症例に対して、マイクロスコープの使用下で再歯根尖切除術と根尖部の緊密な封鎖を目的としたmineral trioxide aggregate(以下MTA)による逆根管充填を含めた再外科的歯内療法が奏効した症例を報告する。患者は59歳の女性で、上顎右側中切歯および側切歯に対して打診痛、根尖部圧痛および瘻孔を認める慢性根尖膿瘍と診断し、最初に通常の感染根管治療を行った。その後、病変部の治癒が期待できなかったため歯根尖切除を行い、逆根管形成後にMTAで逆根管充填を行った。手術から1ヵ月後で瘻孔は消失し、さらに6ヵ月後のレントゲン写真では根尖部の透過像が消失しており、治癒傾向を認めた。(著者抄録)

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  • マイクロスコープを用いた再歯根尖切除術の1例

    大倉 直人, 山本 信一, 阿部 達也, 竹内 亮祐, 遠間 愛子, 枝並 直樹, 吉羽 永子, 吉羽 邦彦, 野杁 由一郎

    新潟歯学会雑誌   48 ( 1 )   29 - 35   2018.6

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    根尖性歯周組織病変の難治化は、根尖部での複雑な解剖学的問題に加え、残存バイオフィルムによる細菌学的な影響もあるとされている。こうした様々な要因の改善策として外科的歯内療法が選択される。しかし、補綴処置を行う上で考慮する歯冠・歯根長比の問題で複数回に渡り歯根切断が行われることは稀である。今回は、歯根尖切除術を過去3度に渡って施行しても治癒しなかった症例に対して、マイクロスコープの使用下で再歯根尖切除術と根尖部の緊密な封鎖を目的としたmineral trioxide aggregate(以下MTA)による逆根管充填を含めた再外科的歯内療法が奏効した症例を報告する。患者は59歳の女性で、上顎右側中切歯および側切歯に対して打診痛、根尖部圧痛および瘻孔を認める慢性根尖膿瘍と診断し、最初に通常の感染根管治療を行った。その後、病変部の治癒が期待できなかったため歯根尖切除を行い、逆根管形成後にMTAで逆根管充填を行った。手術から1ヵ月後で瘻孔は消失し、さらに6ヵ月後のレントゲン写真では根尖部の透過像が消失しており、治癒傾向を認めた。(著者抄録)

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  • 歯髄創傷治癒モデルラットを用いたGlucose Transporter-4の局在および遺伝子発現の解析

    遠間 愛子, 大倉 直人, 枝並 直樹, 竹内 亮祐, 吉羽 永子, 吉羽 邦彦, 野杁 由一郎

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   147回   46 - 46   2017.10

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  • 歯髄創傷治癒過程におけるマクロファージの集積とmyofibroblast様細胞の分化

    枝並 直樹, 吉羽 邦彦, 吉羽 永子, 大倉 直人, 竹内 亮祐, 遠間 愛子, 野杁 由一郎

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   147回   182 - 182   2017.10

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  • 歯髄創傷治癒モデルラットを用いたGlucose Transporter-1およびrunt-related transcription factor 2の局在と遺伝子発現解析

    竹内 亮祐, 大倉 直人, 枝並 直樹, 遠間 愛子, 吉羽 永子, 吉羽 邦彦, 野杁 由一郎

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   147回   86 - 86   2017.10

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  • ヒト歯髄組織創傷治癒過程における骨髄由来間葉系前駆細胞fibrocyteの動態検索

    吉羽 永子, 大倉 直人, 細矢 明宏, 中村 浩彰, 野杁 由一郎, 吉羽 邦彦, 枝並 直樹, 遠間 愛子, 竹内 亮介

    Journal of Oral Biosciences Supplement   2017   427 - 427   2017.9

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  • ラット臼歯断髄後におけるmyofibroblastの動態解析

    枝並 直樹, 吉羽 永子, 大倉 直人, 野杁 由一郎, 吉羽 邦彦

    新潟歯学会雑誌   47 ( 1 )   52 - 52   2017.6

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  • ヒト歯髄組織創傷治癒過程における骨髄由来間葉系前駆細胞fibrocyteの動態検索

    吉羽永子, 大倉直人, 細矢明宏, 中村浩彰, 野杁由一郎, 吉羽邦彦

    Journal of Oral Biosciences Supplement (Web)   2017   2017

  • 各種ケイ酸カルシウム系セメントの生体機能性と直接覆髄後の歯髄反応

    吉羽 邦彦, 枝並 直樹, 日向 剛, 韓 臨麟, 竹内 亮祐, 遠間 愛子, 大倉 直人, 武井 絵梨花, 吉羽 永子, 興地 隆史

    日本歯科医師会雑誌   69 ( 5 )   454 - 454   2016.8

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  • 培養ヒト歯髄に対するprostaglandin EP4レセプターアゴニストの影響

    大倉 直人, 吉羽 永子, 吉羽 邦彦, 小田 陽平, 興地 隆史

    日本歯科医師会雑誌   69 ( 5 )   454 - 454   2016.8

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  • 培養ヒト歯髄に対するprostaglandin EP2レセプターアゴニストの影響

    大倉 直人, 枝並 直樹, 竹内 亮祐, 遠間 愛子, 吉羽 永子, 吉羽 邦彦, 小田 陽平, 興地 隆史

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   144回   124 - 124   2016.6

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  • ラット臼歯におけるMTAによる直接覆髄後のGlucose Transporter-2の免疫組織化学および遺伝子発現の解析

    遠間 愛子, 大倉 直人, 枝並 直樹, 竹内 亮祐, 吉羽 永子, 吉羽 邦彦

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   144回   111 - 111   2016.6

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  • マイクロスコープを用いた再歯根尖切除術の1症例

    大倉 直人, 山本 信一, 枝並 直樹, 吉羽 永子, 吉羽 邦彦, 野杁 由一郎

    日本歯内療法学会学術大会プログラム・抄録集   37回   60 - 60   2016.6

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  • ラット臼歯におけるMineral trioxide aggregateによる直接覆髄後のGlucose Transporter-1の免疫局在および遺伝子発現解析

    竹内 亮祐, 大倉 直人, 枝並 直樹, 遠間 愛子, 吉羽 永子, 吉羽 邦彦

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   144回   112 - 112   2016.6

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  • ラット皮下組織における4-META含有レジン系シーラーの生体親和性 Reviewed

    枝並 直樹, 重谷 佳見, 吉羽 邦彦, 日向 剛, 吉羽 永子, 興地 隆史

    日本歯科保存学雑誌   59 ( 1 )   65 - 73   2016.2

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    目的:近年注目されている接着性レジンシーラーは根管壁象牙質への接着性が謳われる一方で,再根管治療時の除去が困難となることが懸念されている.そこで,比較的容易に除去可能な4-META含有レジンシーラー(メタシールSoft)が開発されたが,その生体親和性についての知見は十分とはいえない.本研究では,同シーラーをラット背部皮下に埋入後,局所に浸潤する炎症性細胞の分布密度を免疫組織化学的に定量することにより,生体親和性を評価した.材料と方法:被験レジン系シーラーとしてメタシールSoft,対照材料としてエポキシレジン系シーラー(AHプラス)および酸化亜鉛ユージノール系シーラー(キャナルス)を使用した.滅菌PTFEチューブに練和したシーラーを填入し,ただちに4週齢Wistar系雄性ラット(各観察期間ともn=5)の背部皮下組織内に埋入した.コントロールとしてシーラーを填入していない滅菌PTFEチューブを用いた.7,14,28日後に皮下組織を切り出し,4%パラホルムアルデヒド溶液で24時間浸漬固定し,凍結切片を作成後,H-E染色による組織学的観察を行うとともに,マクロファージ,好中球の局在についてそれぞれCD68,CD43を一次抗体として酵素抗体染色を行い観察した.さらに,チューブ開口部と接する組織内における陽性細胞の密度を求め,統計学的に分析した(一元配置分散分析およびBonferroni Dunn検定,α=0.05).結果:H-E染色では,すべての実験群において,7日経過例でチューブ開口部を中心とした炎症性細胞の浸潤が認められたが,時間の経過とともに炎症性細胞は減少し,線維組織による被包が観察された.メタシールSoft群ではシーラーの溢出がしばしば組織内に観察され,28日経過例においてもその残存が認められた.定量解析の結果,CD68陽性細胞については7日経過例でキャナルス群がコントロール群より有意に高い値を示したが,14,28日経過例においては各群間で有意差は認めなかった.CD43陽性細胞については,7,14日経過例でメタシールSoft群がキャナルス群より有意に低い値を示した.28日経過例では,各群間で有意差は認めなかった.結論:ラット皮下結合組織においては,4-META含有レジン系シーラーは酸化亜鉛ユージノール系シーラーと比較して,軽微な好中球浸潤を誘発することが示された.(著者抄録)

    DOI: 10.11471/shikahozon.59.65

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    Other Link: https://search.jamas.or.jp/index.php?module=Default&action=Link&pub_year=2016&ichushi_jid=J01092&link_issn=&doc_id=20160314250008&doc_link_id=%2Fdo1conde%2F2016%2F005901%2F008%2F0065-0074%26dl%3D0&url=http%3A%2F%2Fwww.medicalonline.jp%2Fjamas.php%3FGoodsID%3D%2Fdo1conde%2F2016%2F005901%2F008%2F0065-0074%26dl%3D0&type=MedicalOnline&icon=https%3A%2F%2Fjk04.jamas.or.jp%2Ficon%2F00004_2.gif

  • 実験的歯の移動におけるラット臼歯歯髄内prostaglandin I2合成酵素と受容体の発現解析

    大倉 麻里子, 大倉 直人, 吉羽 永子, 吉羽 邦彦, 依田 浩子, 大島 勇人, 齋藤 功, 興地 隆史

    新潟歯学会雑誌   45 ( 2 )   97 - 98   2015.12

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  • ケイ酸カルシウム系覆髄材の生体機能性 ラット皮下組織内埋入によるin vivoの検討

    日向 剛, 吉羽 邦彦, 韓 臨麟, 枝並 直樹, 吉羽 永子, 興地 隆史

    新潟歯学会雑誌   45 ( 2 )   109 - 109   2015.12

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  • 培養ヒト歯髄に対するprostaglandin EP4レセプターアゴニストの影響

    大倉 直人, 吉羽 永子, 吉羽 邦彦, 小田 陽平, 興地 隆史

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   143回   75 - 75   2015.11

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  • 各種ケイ酸カルシウム系セメントによるラット臼歯直接覆髄後の被蓋硬組織形成

    枝並 直樹, 吉羽 邦彦, 武井 絵梨花, 日向 剛, 竹内 亮祐, 遠間 愛子, 重谷 佳見, 吉羽 永子, 興地 隆史

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   143回   148 - 148   2015.11

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  • ラット臼歯歯髄断髄後のProstaglandin Transporterに対する免疫組織学的局在解析

    大倉 直人, 枝並 直樹, 吉羽 永子, 吉羽 邦彦, 依田 浩子, 大島 勇人, 興地 隆史

    Journal of Oral Biosciences Supplement   2015   367 - 367   2015.9

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  • 培養ヒト歯髄の各種遺伝子発現に対するprostaglandin EP4レセプターアゴニストの影響

    大倉 直人, 吉羽 永子, 吉羽 邦彦, 興地 隆史

    日本歯内療法学会学術大会プログラム・抄録集   36回   71 - 71   2015.7

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  • ラット臼歯歯髄断髄後のProstaglandin Transporterに対する免疫組織学的局在解析

    大倉直人, 枝並直樹, 吉羽永子, 吉羽邦彦, 依田浩子, 大島勇人, 興地隆史

    Journal of Oral Biosciences Supplement (Web)   2015   2015

  • ヒト歯髄におけるプロスタグランジンE2輸送担体および特異的レセプターの免疫組織化学的局在解析

    大倉 直人, 大倉 麻里子, 吉羽 永子, 吉羽 邦彦, 小田 陽平, 依田 浩子, 大島 勇人, 興地 隆史

    Journal of Oral Biosciences Supplement   2014   183 - 183   2014.9

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  • ヒト歯髄におけるProstaglandinレセプターの免疫組織学的局在解析

    大倉 直人, 重谷 佳見, 吉羽 永子, 吉羽 邦彦, 興地 隆史

    日本歯内療法学会学術大会プログラム・抄録集   35回   101 - 101   2014.7

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  • ラット臼歯におけるMineral Trioxide Aggregateによる直接覆髄後のM2マクロファージの動態

    武井 絵梨花, 重谷 佳見, 吉羽 邦彦, 日向 剛, 吉羽 永子, 興地 隆史

    新潟歯学会雑誌   44 ( 1 )   55 - 56   2014.6

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  • ヒト歯髄におけるプロスタグランジンE<sub>2</sub>輸送担体および特異的レセプターの免疫組織化学的局在解析

    大倉直人, 大倉麻里子, 吉羽永子, 吉羽邦彦, 小田陽平, 依田浩子, 大島勇人, 興地隆史

    Journal of Oral Biosciences Supplement (Web)   2014   2014

  • ラット炎症歯髄に対する薬物輸送担体を介したProstaglandin E2輸送経路解析

    大倉 直人, 重谷 佳見, 吉羽 永子, 吉羽 邦彦, 興地 隆史

    新潟歯学会雑誌   43 ( 2 )   155 - 155   2013.12

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  • ラット臼歯矯正移動時における歯髄内prostanoid receptorの遺伝子発現と免疫組織学的局在解析

    大倉 直人, 大倉 麻里子, 重谷 佳見, 吉羽 永子, 吉羽 邦彦, 齋藤 功, 興地 隆史

    Journal of Oral Biosciences Supplement   2013   189 - 189   2013.9

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  • ヒト歯髄組織からoutgrowthする細胞による組織構築に関する研究

    吉羽 永子, 吉羽 邦彦, 大倉 直人, 細矢 明宏, 中村 浩彰, 興地 隆史

    Journal of Oral Biosciences Supplement   2013   193 - 193   2013.9

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  • ラット臼歯矯正移動時における歯髄内prostanoid receptorの遺伝子発現と免疫組織学的局在解析

    大倉直人, 大倉麻里子, 重谷佳見, 吉羽永子, 吉羽邦彦, 齋藤功, 興地隆史

    Journal of Oral Biosciences Supplement (Web)   2013   ROMBUNNO.P2‐24 (WEB ONLY)   2013

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    J-GLOBAL

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  • ヒト歯髄組織からoutgrowthする細胞による組織構築に関する研究

    吉羽永子, 吉羽邦彦, 大倉直人, 細矢明宏, 中村浩彰, 興地隆史

    Journal of Oral Biosciences Supplement (Web)   2013   2013

  • ラット炎症歯髄に対する薬物輸送担体の遺伝子発現解析

    大倉 直人, 重谷 佳見, 細矢 明宏, 吉羽 永子, 吉羽 邦彦, 興地 隆史

    Journal of Oral Biosciences Supplement   2012   160 - 160   2012.9

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  • ラット炎症歯髄に対する薬物輸送担体の遺伝子発現解析

    大倉 直人, 重谷 佳見, 細矢 明宏, 吉羽 永子, 吉羽 邦彦, 興地 隆史

    日本歯科医師会雑誌   65 ( 5 )   628 - 628   2012.8

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  • 半導体レーザー照射後のラット臼歯における硬組織形成誘導機構の解明

    重谷 佳見, 大倉 直人, 細矢 明宏, 鈴木 啓展, 吉羽 邦彦, 吉羽 永子, 大島 勇人, 興地 隆史

    日本歯科医師会雑誌   65 ( 5 )   629 - 629   2012.8

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  • ラット実験的根尖歯周炎の成立過程における血管新生関連遺伝子の発現

    山中 裕介, 金子 友厚, 吉羽 邦彦, 伊藤 崇史, 吉羽 永子, 重谷 佳見, 興地 隆史

    新潟歯学会雑誌   42 ( 1 )   62 - 62   2012.6

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  • ラット炎症歯髄に対する薬物輸送担体の遺伝子発現解析

    大倉直人, 重谷佳見, 細矢明宏, 吉羽永子, 吉羽邦彦, 興地隆史

    Journal of Oral Biosciences Supplement (Web)   2012   2012

  • ラット実験的炎症歯髄におけるプロスタノイド受容体遺伝子発現の経時的解析

    大倉 直人, 重谷 佳見, 細矢 明宏, 吉羽 永子, 吉羽 邦彦, 興地 隆史

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   134回   104 - 104   2011.5

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  • ラット炎症歯髄に対する薬物輸送担体の遺伝子発現解析

    大倉 直人, 重谷 佳見, 細矢 明宏, 吉羽 永子, 吉羽 邦彦, 興地 隆史

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   133回   96 - 96   2010.10

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  • ヒト歯髄創傷治癒過程で生じるFibrillin-1の分解は細胞分化と石灰化を誘導する

    吉羽 永子, 吉羽 邦彦, 大倉 直人, 細矢 明宏, 重谷 佳見, 興地 隆史

    特定非営利活動法人日本歯科保存学会学術大会プログラムおよび講演抄録集   132回   117 - 117   2010.5

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  • 半導体レーザー照射後のラット臼歯における硬組織関連タンパクの遺伝子発現

    重谷 佳見, 大倉 直人, 吉羽 邦彦, 吉羽 永子, 興地 隆史

    日本歯内療法学会学術大会プログラム・抄録集   31回   110 - 110   2010.5

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  • 半導体レーザー照射後のラット臼歯における硬組織関連タンパクの遺伝子発現

    重谷佳見, 大倉直人, 吉羽邦彦, 吉羽永子, 興地隆史

    日本歯内療法学会学術大会プログラム・抄録集   31st   2010

  • Mineral trioxide aggregate(MTA)の物理化学的特性と直接覆髄後の歯髄反応

    吉羽 邦彦, 鞍立 桃子, 重谷 佳見, 韓 臨麟, 吉羽 永子, 興地 隆史

    新潟歯学会雑誌   39 ( 2 )   181 - 182   2009.12

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  • 半導体レーザー照射に対するラット臼歯歯髄初期反応

    笹 なつき, 重谷 佳見, 吉羽 邦彦, 吉羽 永子, 監物 新一, 大島 勇人, 興地 隆史

    Journal of Oral Biosciences   51 ( Suppl. )   100 - 100   2009.8

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  • 早期臨床実習への窩洞形成修復実習の導入

    HAN Linlin, 興地隆史, 吉羽永子, 小林哲夫, 藤井規孝, 小野和宏, 前田健康

    日本歯科医学教育学会総会・学術大会プログラム・抄録集   28th   128   2009

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  • Bone-like tissue formation in the pulp cavity after tooth replantation

    HOSOYA Akihiro, CHEN Zhao, YOSHIBA Kunihiko, YOSHIBA Nagako, YAMADA Hirohito, KASAHARA Etsuo, OZAWA Hidehiro, NAKAMURA Hiroaki

    51   88 - 88   2008.5

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  • 歯根膜組織の歯槽骨再生能

    細矢 明宏, 平賀 徹, 中村 浩彰, 吉羽 邦彦, 吉羽 永子, Zhao Chen, 高橋 将文, 岡部 高弘, 脇谷 滋之, 山田 博仁, 笠原 悦夫, 二宮 禎, 小澤 英浩

    THE BONE   22 ( 1 )   3 - 7   2008.1

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  • MTAによるラット臼歯覆髄モデルにおける歯髄反応の免疫組織化学的解析

    鞍立 桃子, 吉羽 邦彦, 重谷 佳見, 吉羽 永子, 大島 勇人, 興地 隆史

    新潟歯学会雑誌   37 ( 2 )   249 - 250   2007.12

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  • Association of TIMP-2 with extracellular matrix exposed to mechanical stress and its co-distribution with periostin during mouse tooth development Reviewed

    N. Yoshiba, K. Yoshiba, A. Hosoya, M. Saito, T. Yokoi, T. Okiji, N. Amizuka, H. Ozawa

    European Cells and Materials   14 ( SUPPL.2 )   142   2007.11

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  • MTAによるラット臼歯覆髄モデルにおける歯髄反応

    鞍立 桃子, 吉羽 邦彦, 重谷 佳見, 吉羽 永子, 大島 勇人, 興地 隆史

    Journal of Oral Biosciences   49 ( Suppl. )   114 - 114   2007.8

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  • ラット歯髄皮下移植後の硬組織形成

    細矢 明宏, 中村 浩彰, 星 和人, 吉羽 邦彦, 吉羽 永子, 高橋 将文, 岡部 高弘, 山田 博仁, 笠原 悦男, 二宮 禎, 佐原 紀行, 小澤 英浩

    THE BONE   21 ( 4 )   399 - 403   2007.7

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  • 歯の形態形成と細胞分化におけるTIMP-1,-2,-3のmRNAとタンパクの発現

    吉羽 永子, 吉羽 邦彦, 興地 隆史

    新潟歯学会雑誌   37 ( 1 )   47 - 48   2007.7

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    tissue inhibitors of metalloproteinase(TIMP)-1、-2、-3の役割を解明する目的で、マウス切歯をモデルとして、その発生過程、細胞分化及び基質形成期におけるこれらの発現を検索した。マウス切歯形態形成、細胞分化、及び基質形成過程において、TIMP-1、-2、-3は時間的空間的にそれぞれ全く異なった発現パターンを示し、臼歯発生過程とも異なるものであった。歯胚上皮-間葉接合部に局在するTIMP-1はエナメル器のアポトーシスを回避している可能性が示唆された。基底膜に観察されるTIMP-2は、同様な機構で歯性上皮細胞の増殖を制御している可能性が示唆された。歯小嚢で発現されるTIMP-2はその組織化に関与していると思われた。現在のところ、象牙芽細胞下層におけるTIMP-3の役割は不明である。

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  • An immunohistochemical study on pulpal responses to mineral trioxide aggregate in human teeth

    YOSHIBA Kunihiko, YOSHIBA Nagako, SHIGETANI Yoshimi, HOSOYA Akihiro, OKIJI Takashi

    50   138 - 138   2007.5

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  • 歯周組織発生過程におけるα-平滑筋アクチンの局在と歯槽骨形成との関連

    細矢 明宏, 中村 浩彰, 吉羽 邦彦, 吉羽 永子, 中谷 宏幸, 脇谷 滋之, 山田 博仁, 笠原 悦男, 二宮 禎, 小澤 英浩

    THE BONE   21 ( 3 )   281 - 285   2007.5

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  • 歯髄の潜在的硬組織形成能

    細矢明宏, 中村浩彰, 二宮 禎, 星 和人, 吉羽邦彦, 吉羽永子, 高橋将文, 岡部高弘, 佐原紀行, 山田博仁, 笠原悦男, 小澤英浩

    The BONE   21(4)   399 - 403   2007

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  • An immunohistochemical study on pulpal responses to mineral trioxide aggregate in rat molars

    KURATATE Momoko, YOSHIBA Kunihiko, SHIGETANI Yoshimi, YOSHIBA Nagako, OKIJI Takashi

    49   150 - 150   2006.10

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  • Alveolar bone regeneration after tooth transplantation into subcutaneous tissue

    HOSOYA Akihiro, NAKAMURA Hiroaki, YOSHIBA Kunihiko, YOSHIBA Nagako, YAMADA Hirohito, KASAHARA Etsuo, OZAWA Hidehiro

    49   42 - 42   2006.10

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  • マウス歯の発生過程におけるTIMP-2とPeriostinは類似の時間的空間的発現パターンを示す

    吉羽 永子, 吉羽 邦彦, 興地 隆史, 斎藤 正寛, 横井 隆政, 細矢 明宏, 網塚 憲生, 小澤 英浩

    Journal of Oral Biosciences   48 ( Suppl. )   153 - 153   2006.9

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  • Expression of TIMP-2 and Periostin during Mouse Tooth Development

    YOSHIBA Nagako, YOSHIBA Kunihiko, OKIJI Takashi, SAITO Masahiro, YOKOI Takamasa, HOSOYA Akihiro, OZAWA Hidehiro

    49   98 - 98   2006.4

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  • Immunohistochemical Localization of α-SMA during rat molar tooth development

    HOSOYA Akihiro, NAKAMURA Hiroaki, YOSHIBA Kunihiko, YOSHIBA Nagako, YAMADA Hirohito, KASAHARA Etsuo, OZAWA Hidehiro

    48   59 - 59   2005.10

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  • 歯科用レーザーのう蝕治療への応用に関する研究 レーザー照射に対する歯髄反応について

    吉羽 邦彦, 楯 泰昌, 吉羽 永子, 岩久 正明, 興地 隆史, 大島 勇人

    歯界展望   特別号 ( 健康な心と身体は口腔から-発ヨコハマ2004- )   274 - 274   2005.6

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  • ラット臼歯への半導体レーザー照射に対する歯髄反応

    楯 泰昌, 吉羽 邦彦, 吉羽 永子, 岩久 正明, 興地 隆史, 大島 勇人

    新潟歯学会雑誌   34 ( 2 )   288 - 288   2005.1

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  • Gene Expression and Protein Localization of MMP-2, -9, MT1-MMP and TIMP-1, -2, -3 During Mouse Molar Tooth Development

    YOSHIBA Nagako, YOSHIBA Kunihiko, OKIJI Takashi, HOSOYA Akihiro, OZAWA Hidehiro

    47   195 - 195   2004.10

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  • Immunohistochemical Study of Hard Tissue Formation in a Subcutaneously-transplanted Rat Dental Pulp

    HOSOYA Akihiro, YOSHIBA Kunihiko, YOSHIBA Nagako, YAMADA Hirohito, KASAHARA Etsuo, OZAWA Hidehiro

    47   27 - 27   2004.10

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  • マウス切歯発生過程におけるTIMPsの発現

    吉羽 永子, 吉羽 邦彦, 興地 隆史, 細矢 明宏, 小澤 英浩

    Journal of oral biosciences   46 ( 5 )   451 - 451   2004.9

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  • 歯髄の硬組織形成機構に関する微細構造学的研究 ラット切歯歯髄皮下移植実験モデルを用いた検討

    細矢 明宏, 中村 浩彰, 星 和人, 佐原 紀行, 二宮 禎, 吉羽 邦彦, 吉羽 永子, 小澤 英浩

    Journal of Oral Biosciences   46 ( 5 )   377 - 377   2004.9

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  • 歯科用レーザーのう蝕治療への応用に関する研究 レーザー照射に対する歯髄反応について

    吉羽 邦彦, 楯 泰昌, 吉羽 永子, 岩久 正明, 興地 隆史, 大島 勇人

    日本歯科医師会雑誌   57 ( 4 )   401 - 401   2004.7

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  • Immunohistochemical study on pulpal response of rat molars to irradiation by semiconductor laser

    TATE Yasuaki, YOSHIBA Kunihiko, YOSHIBA Nagako, SHIGETANI Yoshimi, OKIJI Takashi, IWAKU Masaaki

    46   117 - 117   2003.10

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  • マウス臼歯発生過程におけるMMPsおよびTIMPsの発現

    吉羽 邦彦, 吉羽 永子, 小澤 英浩

    歯科基礎医学会雑誌   45 ( 5 )   293 - 293   2003.9

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  • ラット皮下移植歯髄の硬組織誘導能に関する免疫組織化学的研究

    細矢 明宏, 星 和人, 吉羽 邦彦, 吉羽 永子, 笠原 悦男, 小澤 英浩

    歯科基礎医学会雑誌   45 ( 5 )   295 - 295   2003.9

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  • ラット臼歯における半導体レーザー照射に対する歯髄反応

    楯 泰昌, 吉羽 邦彦, 吉羽 永子, 岩久 正明, 大島 勇人

    歯科基礎医学会雑誌   45 ( 5 )   295 - 295   2003.9

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  • An immunohistochemical study of hard tissue formation in the pulp cavity of rat molar after transplantation into subcutaneous tissue. Reviewed

    A. Hosoya, K. Yoshiba, N. Yoshiba, K. Hoshi, M. Iwaku, H. Ozawa

    JOURNAL OF DENTAL RESEARCH   82   B252 - B252   2003.6

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  • Responses of MHC class II molecule-expressing cells to cavity preparation and caries treatment.

    K. Yoshiba, N. Yoshiba, M. Iwaku

    JOURNAL OF DENTAL RESEARCH   82   B187 - B187   2003.6

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  • Immunohistochemical Distribution of Antigen Presenting Cells After Cavity Preparation and Caries Treatment

    YOSHIBA Kunihiko, YOSHIBA Nagako, IWAKU Masaaki

    46   102 - 102   2003.5

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  • ラット臼歯の皮下移植後に観察される歯髄腔内硬組織形成

    細矢 明宏, 小澤 英浩, 吉羽 邦彦, 吉羽 永子, 岩久 正明, 星 和人

    THE BONE   17 ( 2 )   3 - 6   2003.3

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  • ラット臼歯の皮下移植後に形成される歯髄腔内硬組織形成

    細矢明宏, 吉羽邦彦, 吉羽永子, 星和人, 岩久正明, 小澤英浩

    The BONE   17(2)   107 - 110   2003

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  • ラット切歯先端部硬組織の免疫組織化学的研究

    細矢 明宏, 星 和人, 吉羽 邦彦, 吉羽 永子, 笠原 悦男, 小澤 英浩

    歯科基礎医学会雑誌   44 ( 5 )   417 - 417   2002.9

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  • う蝕治療へのEr:YAG laserの応用

    重谷 佳見, 田辺 啓太, 楯 泰昌, 吉羽 邦彦, 吉羽 永子, 岡本 明, 子田 晃一, 岩久 正明

    新潟歯学会雑誌   32 ( 1 )   83 - 84   2002.7

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  • Expression of non-collagenous proteins and TGF-bs during reparative dentinogenesis.

    K Yoshiba, N Yoshiba, M Iwaku

    JOURNAL OF DENTAL RESEARCH   81   A155 - A155   2002.3

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  • マウス切歯歯胚における半接着斑関連タンパクの局在

    吉羽 邦彦, 吉羽 永子, 岩久 正明, 小澤 英浩

    歯科基礎医学会雑誌   43 ( 5 )   566 - 566   2001.8

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  • ラット臼歯皮下移植実験モデルにおける歯髄腔内硬組織形成に関する免疫組織化学的研究

    細矢 明宏, 吉羽 邦彦, 吉羽 永子, 星 和人, 岩久 正明, 小澤 英浩

    歯科基礎医学会雑誌   43 ( 5 )   569 - 569   2001.8

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  • 歯髄の石灰化に関する免疫組織化学的研究 ラット臼歯皮下移植実験モデルを用いた検討

    細矢 明宏, 吉羽 邦彦, 吉羽 永子, 星 和人, 岩久 正明, 小澤 英浩

    日本歯科保存学雑誌   44 ( 春季特別 )   21 - 21   2001.4

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  • ラット臼歯の皮下移植後に形成される歯髄腔内硬組織に関する免疫組織化学的研究

    細矢 明宏, 吉羽 邦彦, 吉羽 永子, 星 和人, 岩久 正明, 小澤 英浩

    新潟歯学会雑誌   30 ( 2 )   268 - 268   2000.12

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  • マウス歯胚発生過程におけるラミニンー5の特異的発現

    吉羽 邦彦, 吉羽 永子, 岩久 正明

    歯科基礎医学会雑誌   42 ( 5 )   460 - 460   2000.8

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  • Er YAGレーザーを用いたラット臼歯窩洞形成後の歯髄反応に関する免疫組織化学的研究

    田辺 啓太, 吉羽 邦彦, 吉羽 永子, 岩久 正明, 小澤 英浩

    歯科基礎医学会雑誌   42 ( 5 )   456 - 456   2000.8

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    Language:Japanese   Publisher:歯科基礎医学会  

    CiNii Article

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  • Er:YAGレーザーによる窩洞形成後の歯髄反応に関する免疫組織化学的研究 ラット臼歯を用いた基礎的研究

    田辺 啓太, 吉羽 邦彦, 吉羽 永子, 岩久 正明, 小澤 英浩

    新潟歯学会雑誌   29 ( 2 )   208 - 209   1999.12

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    Language:Japanese   Publisher:新潟歯学会  

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  • Immunohistochemical study on pulpal response in rat molars after cavity preparation by Er:YAG Laser

    TANABE Keita, YOSHIBA Kunihiko, YOSHIBA Nagako, IWAKU Masaaki, OZAWA Hidehiro

    42   66 - 66   1999.10

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  • エナメル芽細胞分化過程におけるラミニン-5とα6β4インテグリンの局在

    吉羽 邦彦, 吉羽 永子, 岩久 正明, 小澤 英浩

    歯科基礎医学会雑誌   41 ( 5 )   442 - 442   1999.8

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    Language:Japanese   Publisher:歯科基礎医学会  

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  • Expression and localization of Laminin-5 and Integrins in Mouse Tooth.

    YOSHIBA Kunihiko, YOSHIBA Nagako, IWAKU Masaaki, OZAWA Hidehiro

    41   167 - 167   1998.10

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  • Immunohistochemical Localizations of Class II Antigens and Nerve Fibers in Human Carious Teeth.

    YOSHIBA Nagako, YOSHIBA Kunihiko, IWAKU Masaaki, OZAWA Hidehiro

    41   165 - 165   1998.10

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  • Distribution of Odontoblast Processes in Human Dentin

    YOSHIBA Kunihiko, YOSHIBA Nagako, EJIRI Sadakazu, IWAKU Masaaki, OZAWA Hidehiro

    日本歯科保存学雑誌   40   117 - 117   1997.5

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  • Immunolocalization of the Small Proteoglycan Decorin in Human Teeth

    YOSHIBA Nagako, YOSHIBA Kunihiko, IWAKU Masaaki, OZAWA Hidehiro

    40   118 - 118   1997.5

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  • DISTRIBUTION OF FIBRONECTIN DURING DENTIN BRIDGE FORMATION AFTER PULP CAPPING Reviewed

    K YOSHIBA, N YOSHIBA, M IWAKU, H NAKAMURA, H OZAWA

    JOURNAL OF DENTAL RESEARCH   73   318 - 318   1994

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    Language:English   Publishing type:Research paper, summary (international conference)   Publisher:AMER ASSOC DENTAL RESEARCH  

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  • DISTRIBUTION OF FIBRONECTIN IN ODONTOBLAST LAYER OF HUMAN TEETH

    N YOSHIBA, K YOSHIBA, M IWAKU, H NAKAMURA, H OZAWA

    JOURNAL OF DENTAL RESEARCH   73   121 - 121   1994

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    Web of Science

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  • ORAL HEALTH OF A WORKING-AGED POPULATION IN AN AGING AREA

    M FUKUSHIMA, K YOSHIBA, N YOSHIBA, M IWAKU

    JOURNAL OF DENTAL RESEARCH   73   235 - 235   1994

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    Web of Science

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  • Epidemiological Survey of Oral Disease in an Adult Population : Prevalence of Periodontal Disease and Root Surface Caries.

    Japanese Journal of Conservative Dentistry   36 ( 1 )   295 - 302   1993

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  • Evaluation of Various Twbed Media for Efficient Bacteriological Examination in Endodontics

    Niigata Dental Journal   22 ( 1 )   1992

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Awards

  • 令和3年度新潟大学優秀論文賞

    2021.12   新潟大学  

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  • 学術賞

    2014   日本歯科保存学会  

    吉羽 永子

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  • 奨励賞

    1998   日本歯科保存学会  

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    Country:Japan

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  • 海外派遣助成賞

    1995   財団法人山下太郎  

    吉羽 永子

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Research Projects

  • Functional analysis of extracellular matrix laminin affecting macrophage phenotype

    Grant number:22H03259

    2022.4 - 2026.3

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

    Research category:Grant-in-Aid for Scientific Research (B)

    Awarding organization:Japan Society for the Promotion of Science

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    Grant amount:\17160000 ( Direct Cost: \13200000 、 Indirect Cost:\3960000 )

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  • 歯髄の創傷治癒・再生過程におけるGli1陽性幹細胞の動態と分化誘導機構の解明

    Grant number:21K09914

    2021.4 - 2024.3

    System name:科学研究費助成事業 基盤研究(C)

    Research category:基盤研究(C)

    Awarding organization:日本学術振興会

    吉羽 邦彦, 吉羽 永子, 枝並 直樹, 細矢 明宏, 入江 一元

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    Grant amount:\4160000 ( Direct Cost: \3200000 、 Indirect Cost:\960000 )

    歯の保存には歯髄の保護・保存が重要であり、直接覆髄や一部生活歯髄切断など歯髄保存療法が、また失われた歯髄に対しては再生歯内療法が試みられている。しかし根管内に形成される硬組織はセメント質あるいは骨様組織であり、理想的な「象牙質・歯髄複合体の再生」には至っていない。また、より迅速かつ確実な処置法の開発が望まれている。本研究では、歯髄の創傷治癒・再生過程において中心的役割を果たすと考えられている間葉系幹細胞の動態について、転写因子Gli1発現をマーカーとして細胞系譜解析により検索するとともに、覆髄材・根管充填材として注目されているバイオセラミックス配合材料の歯髄創傷治癒に与える影響について検討した。
    遺伝子改変(Gli1-CreERT2/Tomato)マウスにおいて、Gli1陽性細胞は歯根膜および歯髄の血管周囲に散在性に認められること、また、臼歯歯根膜から分取されたGli1陽性細胞はコロニー形成能、多分化能を示し幹細胞特性を持つことが明らかにされた。これらの結果から、再生歯内療法応用後の治癒過程、特に硬組織形成におけるGli1陽性細胞の関与が示唆された。一方、バイオセラミックス配合覆髄材・根管充填材のin vitro・in vivoにおけるアパタイト析出能を検討した結果、すべての被験材料は疑似体液中でその表面にCa-Pを含むアパタイト様構造物を析出させたが、ラット皮下移植実験では、材料間にその析出能の差異が認められた。また、レジン含有ケイ酸カルシウム(MTA)系セメントは断髄後、CD68陽性マクロファージの持続的な浸潤と修復象牙質形成の遅延が観察され、材料選択の重要性が示唆された。

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  • シュワン細胞がマクロファージを神経保護作用を有する表現型に誘導する機序の解明

    2021.4 - 2022.3

    System name:科学技術人材育成費補助金「ダイバーシティ研究環境実現イニシアティブ(先端型)」

    Awarding organization:文部科学省

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  • ヒト歯髄の創傷治癒過程におけるM2マクロファージとシュワン細胞の相互作用の解明

    Grant number:19K10146

    2019.4 - 2022.3

    System name:科学研究費助成事業 基盤研究(C)

    Research category:基盤研究(C)

    Awarding organization:日本学術振興会

    吉羽 永子, 吉羽 邦彦, 大倉 直人, 枝並 直樹

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    Grant amount:\4290000 ( Direct Cost: \3300000 、 Indirect Cost:\990000 )

    免疫機能の中心的役割を担っているマクロファージは、その誘導因子により、異なる性質を持つ2つの分化型に分化することが知られている。すなわちM1マクロファージと称される炎症を促進するものと、その反対にM2マクロファージという炎症を抑制し組織の修復に働くタイプのものである。本研究では、ヒト歯髄組織におけるマクロファージの特にM2型の動態について、in vivoおよびin vitroにおいて解析している。
    令和元年度では先ずin vivoすなわち、(1)ヒトの健全歯髄組織、(2)覆髄剤であるMTAを用いた直接覆髄後の創傷治癒過程、さらに(3)様々な病態のう蝕歯髄におけるM2マクロファージの動態を検索した。その結果、これら全ての状態において、CD163をマーカーとするM2マクロファーは神経線維保護機能を有するシュワン細胞と共在していることが明らかとなった。シュワン細胞はp75 nerve growth factor receptor (NGFR)を発現していることから、そのリガンドの1つであるbrain-derived neurotrophic factor (BDNF)の局在を調べたところ、それらCD163陽性のM2マクロファージはBDNFを発現していることが確認された。シュワン細胞とM2マクロファージの相互作用をさらに解析するために、ヒト歯髄組織からシュワン細胞を取り出し、in vitroにおいてTHP-1細胞由来マクロファージ(M0)との共培養を試みた。その結果、シュワン細胞に接触しとどまっているTHP-1マクロファージは、その形を球状から紡錘形に変化させ、CD163陽性のM2マクロファージに分化することが明らかとなった。歯髄は神経線維が非常に豊富な組織である。シュワン細胞はその神経線維を物理的にあるいは様々なサイトカインを分泌することで直接的に保護している。一方で、シュワン細胞は、神経線維を破壊するM1型マクロファージをM2型に変化させることで神経線維を保護していることが示唆された。

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  • The clarification of the pulp generation mechanism to improve the odontoblast differentiation by the transporter of ascorbic acid.

    Grant number:19K10147

    2019.4 - 2020.3

    System name:Grants-in-Aid for Scientific Research

    Research category:Grant-in-Aid for Scientific Research (C)

    Awarding organization:Japan Society for the Promotion of Science

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    Grant amount:\4290000 ( Direct Cost: \3300000 、 Indirect Cost:\990000 )

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  • A study on the mechanisms of induction and differentiation of stem cells during wound healing and regeneration of dentin-pulp complex

    Grant number:16H05516

    2016.4 - 2020.3

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

    Research category:Grant-in-Aid for Scientific Research (B)

    Awarding organization:Japan Society for the Promotion of Science

    YOSHIBA Kunihiko

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    Grant amount:\17030000 ( Direct Cost: \13100000 、 Indirect Cost:\3930000 )

    This study aimed to elucidate the mechanism of dental pulp wound healing and reparative dentinogenesis. We investigated the cellular events and expression of related factors after direct pulp capping or pulpotomy. The results suggested that α-SMA-positive myofibroblasts are progenitors of odontoblast-like cells and that TGF-β1 and EDA-fibronectin are involved in their differentiation. In addition, bone marrow-derived mesenchymal cells, fibrocytes, appeared transiently in the early stages and M2 phenotype macrophages colocalized with Schwann cells in healthy as well as inflamed pulp. It was suggested that various types of cells are involved in pulp wound healing and repair process.

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  • Role of alpha-SMA expressing cells in dental pulp wound healing and regeneration

    Grant number:16K11546

    2016.4 - 2019.3

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

    Research category:Grant-in-Aid for Scientific Research (C)

    Awarding organization:Japan Society for the Promotion of Science

    Yoshiba Nagako

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    Grant amount:\4680000 ( Direct Cost: \3600000 、 Indirect Cost:\1080000 )

    Dental pulp is a soft connective tissue encased in mineralized dentine. When dentine continuity is disrupted by caries or traumatic injury, the resident odontoblasts are also destroyed at that site. Pulp capping agents can help to cover the site through the production of a mineralized matrix through the activation of newly differentiated odontoblast-like cells, however, the exact mechanisms of the process have not been fully revealed. The present study has demonstrated that α-SMA expressing cells play a crucial role in the wound healing.

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  • Regeneration of rat molar pulp tissue by the implantation of stem cells from incisor pulp: development of a rat model of autologous pulp tissue engineering

    Grant number:26293405

    2014.4 - 2017.3

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

    Research category:Grant-in-Aid for Scientific Research (B)

    Awarding organization:Japan Society for the Promotion of Science

    OKIJI Takashi

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    Grant amount:\16250000 ( Direct Cost: \12500000 、 Indirect Cost:\3750000 )

    This study aimed to establish an experimental autologous coronal pulp regeneration model using rat molars and examine whether co-implantation of endothelial cells with stem cells accelerates pulp tissue regeneration. Rat bone marrow mesenchymal stem cells (MSCs) with PLLA/peptide hydrogel constructs were implanted into the coronal pulp chamber of pulpotomized maxillary first molars of Wistar rats. One week after the implantation, a pulp-like tissue was generated in the pulp chamber. In teeth in which rat endothelial cells were co-implanted with MSCs, gene expression levels of pro-angiogenic factors such as Cxcl1 and dentin sialophosphoprotein were elevated compared with MSC-implanted teeth. The co-implanted teeth also showed accelerated scaffold absorption and complete dentin bridge formation. A similar analysis is in progress using stem cells isolated from rat incisor pulp tissues.

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  • Contribution of Myofibroblasts in Dental Pulp Healing and Regeneration

    Grant number:25462952

    2013.4 - 2016.3

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

    Research category:Grant-in-Aid for Scientific Research (C)

    Awarding organization:Japan Society for the Promotion of Science

    YOSHIBA NAGAKO, YOSHIBA KUNIHIKO, OHKURA NAOTO

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    Grant amount:\4810000 ( Direct Cost: \3700000 、 Indirect Cost:\1110000 )

    Alpha-smooth muscle actin (α-SMA) is a marker of myofibroblasts, which play a central role in wound healing. Fibrillin-1 is a major constituent of microfibrils and an extracellular-regulator of TGF-β1, an important cytokine for myofibroblasts differentiation. We examined alterations of α-SMA and fibrillin-1 expression in organotypic culture of dental pulp. After 7 days of culture, most fibroblasts and odontoblasts were immunoreactive for α-SMA with a significant increase of α-SMA mRNA expression. Furthermore, immunostaining of fibrillin-1 became faint with mRNA downregulation. Administration of inhibitors for extracellular matrix proteases resulted in the recovery of fibrillin-1 immunostaining, and fibroblasts lost their immunoreactivity for α-SMA with mRNA downregulation. These findings suggest that fibrillin-1 degradation and downregulation might be implicated in the myofibroblasts differentiation in dental pulp wound healing.

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  • Application of activated macrophages derived from dental pulp stem cells to dental pulp tissue engineering

    Grant number:25670808

    2013.4 - 2015.3

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Challenging Exploratory Research

    Research category:Grant-in-Aid for Challenging Exploratory Research

    Awarding organization:Japan Society for the Promotion of Science

    OKIJI Takashi, YOSHIBA Nagako, YOSHIBA Kunihiko, KANEKO Tomoatsu, SHIGETANI Yoshimi, OHKURA Naoto

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    Grant amount:\3640000 ( Direct Cost: \2800000 、 Indirect Cost:\840000 )

    This study aimed to examine the role of macrophages in the regeneration of dental pulp tissues. After co-implantation of ED2-positive macrophages isolated from rat dental pulp with stem cells into the coronal pulp chamber of pulpotomized rat molars, expression of macrophage markers in the engineered pulp tissue was analyzed. When IL-4 treated ED2-positive macrophages were co-implanted with stem cells, the engineered tissues showed a significant increase in the number of ED2-positive cells and the expression of CD34 and CD163 mRNAs. These findings suggest that M2 macrophages show proliferation and act as wound healing macrophages after co-implantation of macrophages with stem cells.

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  • A study on the mechanisms of tissue wound healing and regeneration of dental pulp

    Grant number:24592863

    2012.4 - 2015.3

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

    Research category:Grant-in-Aid for Scientific Research (C)

    Awarding organization:Japan Society for the Promotion of Science

    YOSHIBA KUNIHIKO, YOSHIBA Nagako

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    Grant amount:\5200000 ( Direct Cost: \4000000 、 Indirect Cost:\1200000 )

    The aim of this study was to elucidate cellular events during pulp tissue wound healing and reparative dentin formation after direct pulp capping. M2 macrophage-associated molecule-expressing cells transiently accumulated beneath the exposure site. The deposition of osteopontin and dentin matrix protein 1 (DMP1) at exposed pulp sites preceded the appearance of nestin-immunoreactive cells, active cell proliferation and new matrix formation. These suggest that M2 macrophages participate in the initial phases of the healing and that osteopontin and DMP1 act as a trigger of pulp repair. In addition, numerous α-smooth muscle actin (SMA)-positive cells emerged at the wound margin, and the initially formed mineralized barrier was lined with α-SMA-positive cells similar in appearance to reparative odontoblasts, some of which co-expressed nestin. Fibrillin-1 degradation and down-regulation might be implicated in the differentiation of α-SMA-positive cells in this process.

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  • Dental pulp regeneration by stem cell transplantation: development of scaffolds and establishment of an animal experimental model

    Grant number:23390433

    2011.4 - 2014.3

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

    Research category:Grant-in-Aid for Scientific Research (B)

    Awarding organization:Japan Society for the Promotion of Science

    OKIJI Takashi, YOSHIBA Nagako, YOSHIBA Kunihiko, OHSHIMA Hayato, KANEKO Tomoatsu, SHIGETANI Yoshimi

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    Grant amount:\18720000 ( Direct Cost: \14400000 、 Indirect Cost:\4320000 )

    This study was designed with the ultimate purpose of establishing a tooth pulp tissue engineering using transplantation of dental pulp stem cells, and conducted to select appropriate scaffolds and develop an experimental engineering system using rat molars. It was demonstrated that rat dental pulp is equipped with stem-like cells that coexpress CD146 and MAP1B and are distributed predominantly in theperivascular area. rat dental pulp by means of immunohistochemistry. Moreover, transplantation of rat mesenchymal stem cells into pulpotomized rat molars with a PLLA/Puramatrix scaffold resulted in the formation of new mineralized tissues at 4 weeks. These results suggest that the experimental model used in this study is useful for investigating the pulp tissue regeneratiopn using stem cell transplantation.

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  • Elucidation of the mechanism of post-repantation replacement resorption as a basis for the development of periodontal ligament regeneration therapy

    Grant number:23659887

    2011 - 2012

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Challenging Exploratory Research

    Research category:Grant-in-Aid for Challenging Exploratory Research

    Awarding organization:Japan Society for the Promotion of Science

    OKIJI Takashi, YOSHIBA Kunihiko, YOSHIBA Nagako, SHIGETANI Yoshimi, KANEKO Tomoatsu, YAMANAKA Yusuke, ITO Takafumi, HINATA Go

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    Grant amount:\3640000 ( Direct Cost: \2800000 、 Indirect Cost:\840000 )

    External replacement resorption, characterized by the fusion of the bone and tooth root (ankylosis), is known as an uncontrollable complication after tooth replantation. This study was conducted as an initial exploration of the pathogenic mechanisms of post-replantation replacement resorption. Experimental replantation of rat molars were carried out in two different conditions (immediate replantation and replantation after sodium hypochlorite immersion), and pathological changes of the pulpal and periodontalligament (PDL) tissues were examined at 1 to 3 weeks postooperatively by means of histology and immunohistochemistry. Results demonstrated that (i) ankylotic changes in the PDL, (ii) bone-like tissue formation in the pulp, and (iii) CD68-expressing macrophage infiltration in the pulp and PDL were more prominent in the NaClO-immersed group than in the immediately replanted group. These findings suggest that severe damage to the PDL cells remaining on the root surface triggers the development of the above-mentioned pathological changes.

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  • Degradation and down-regulation of fibrillin-1 during wound healing of human dental pulp tissue

    Grant number:22592119

    2010 - 2012

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

    Research category:Grant-in-Aid for Scientific Research (C)

    Awarding organization:Japan Society for the Promotion of Science

    YOSHIBA Nagako, YOSHIBA Kunihiko

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    Grant amount:\4550000 ( Direct Cost: \3500000 、 Indirect Cost:\1050000 )

    Degradation of fibrillins, the major constituents of microfibrils, is known to facilitate therelease of active transforming growth factor-β (TGF-β), a signaling molecule contributing tomineralized tissue barrier formation in exposed dental pulps. To examine the involvement offibrillins in the barrier formation, temporospatial expression of (i) genes and proteins offibrillins, and (ii) factors possibly associated with fibrillin degradation and cytodifferentiationwere examined in exposed human pulps. Human pulp slice cultures were also examined for therole of fibrillins in mineralization. Degradation and downregulation of fibrillin-1 expressiontook place during the mineralized tissue barrier formation in exposed pulps in vivo as well as in pulp slice cultures, which should be correlated with the temporary appearance ofα-SMA-positive fibroblasts and pulpal tissue mineralization.

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  • A study on the mechanisms of tissue repair and regeneration of dentin-pulp complex

    Grant number:21592417

    2009 - 2011

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

    Research category:Grant-in-Aid for Scientific Research (C)

    Awarding organization:Japan Society for the Promotion of Science

    YOSHIBA Kunihiko, YOSHIBA Nagako, SHIGETANI Yoshimi

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    Grant amount:\4680000 ( Direct Cost: \3600000 、 Indirect Cost:\1080000 )

    The aim of this study was to elucidate cellular events during pulp tissue repair and reparative dentin formation after tooth injury. The up-regulation ofα-smooth muscle actin(SMA) was characteristically observed in fibroblastic cells in an organ culture system of rat molar. Thy-1-positive dental pulp cells localized in the subodontoblastic layer had the ability to differentiate into hard tissue-forming cells. Theα-SMA-positive and/or Thy-1-positive cells may play a role in tissue repair and regeneration of dental pulp.

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  • Pathogenic mechanisms in apical periodontitis : innate immunity, acquired immunity and dendritic cell maturation

    Grant number:20390483

    2008 - 2010

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

    Research category:Grant-in-Aid for Scientific Research (B)

    Awarding organization:Japan Society for the Promotion of Science

    OKIJI Takashi, YOSHIBA Kunihiko, YOSHIBA Nagako, OHSHIMA Hayato, SHIGETANI Yoshimi, KANEKO Tomoatsu

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    Grant amount:\16770000 ( Direct Cost: \12900000 、 Indirect Cost:\3870000 )

    This study aimed to advance the understanding of the involvement of dendritic cells and immunoregulatory molecules in the pathogenic mechanisms in apical periodontitis. Results demonstrated that unsealed pulp exposure caused the upregulation of MHC class II molecules, CD86, CD83, TLR2 and TLR4 mRNAs in the periodontal ligament of rat molars, as revealed by laser microdissection and real time PCR. Moreover, by employing a whole tooth culture system of the rat molar, it was demonstrated that resident macrophages and dendritic cells upregulated the expression of CD14, TLR4 and CX3CR1 following LPS stimulation.

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  • A study of pulpal responses to laser irradiation

    Grant number:19592197

    2007 - 2009

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

    Research category:Grant-in-Aid for Scientific Research (C)

    Awarding organization:Japan Society for the Promotion of Science

    YOSHIBA Nagako, YOSHIBA Kunihiko, TAKENAKA Shohji

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    Grant amount:\4550000 ( Direct Cost: \3500000 、 Indirect Cost:\1050000 )

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  • 歯内歯周疾患の病態形成機序-新規疾患モデルを用いた樹状細胞の動態解析

    Grant number:19659496

    2007 - 2008

    System name:科学研究費助成事業 萌芽研究

    Research category:萌芽研究

    Awarding organization:日本学術振興会

    興地 隆史, 吉羽 邦彦, 吉羽 永子

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    Grant amount:\3200000 ( Direct Cost: \3200000 )

    樹状細胞は生体内で最も有力な抗原提示細胞であり、この機能を通じて外来抗原刺激に対する免疫応答の発動と質の決定という重要な役割を演じる。これらの細胞が根分岐部における生体防御機構の発動様式の特異性に密接に関連すると推定されるが、根分岐部病変に対する適切な動物モデルは現在まで開発されておらず、根分岐部歯根膜で営まれる免疫防御機構の実態は不明のままである。本研究ではこの点に着目し、新たな実験的根分岐部病変誘発法を採用し、樹状細胞を中心とする各種免疫担当細胞の動態の検索を通じて根分岐部病変の成立・持続過程で営まれる免疫機構の解明をはかることを目的とするものである。
    本年度は先ず、ラット臼歯歯根膜の種々の部位(根分岐部,根尖部,近心側,遠心側)における抗原提示細胞の成熟度,活性化度の相違を検索することを目的として,major histocompatibility complex(MHC)クラスII分子,CD86,CD83およびToll-like receptor(TLR)4のmRNA発現をリアルタイムPCR法を用いて定量解析した.その結果,全ての検索対象遺伝子とも、最も口腔環境に近接した部位である根分岐部において発現レベルが著明に高く,根尖部においては低レベルであった.従って、ラット正常歯根膜の各部位には,細菌刺激の多寡などの微小環境の相違に応じて、成熟度,活性化度の異なる抗原提示細胞が存在することが示唆された.さらに、ラット臼歯を1日間露髄開放し,正常時と露髄開放後の分岐部歯根膜における上記mRNA発現をリアルタイムPCR法を用いて定量,比較したところ,各遺伝子とも開放1日経過後で有意に増加した.従って、ラット臼歯の根分岐部歯根膜に存在する抗原提示細胞が,歯髄の感染に起因する細菌刺激侵襲に応じて,速やかに成熟度や活性化度を増加させることが示唆された.

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  • A study on mechanisms of tissue repair and regeneration of the dentin-pulp complex

    Grant number:18592085

    2006 - 2008

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

    Research category:Grant-in-Aid for Scientific Research (C)

    Awarding organization:Japan Society for the Promotion of Science

    YOSHIBA Kunihiko, YOSHIBA Nagako, HOSOYA Akihiro

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    Grant amount:\4020000 ( Direct Cost: \3600000 、 Indirect Cost:\420000 )

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  • Immune defense mechanisms of the dentin/pulp complex: Immunohistochemical analysis on the heterogeneity and kinetics of dendritic cells

    Grant number:17390508

    2005 - 2007

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (B)

    Research category:Grant-in-Aid for Scientific Research (B)

    Awarding organization:Japan Society for the Promotion of Science

    OKIJI Takashi, YOSHIBA Kunihiko, YOSHIBA Nagako, OHSHIMA Hayato

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    Grant amount:\14340000 ( Direct Cost: \13500000 、 Indirect Cost:\840000 )

    This study investigated dendritic cell (DC) subpopulations and co-stimulatory molecule-expressing cells in pulpitis and apical periodontitis by means of immunohistochemistry and transmission electron microscopy.
    (1) Pulpitis and apical periodontitis were induced in rat molars by making unsealed pulp exposures, and kinetics of DCs was investigated by means of immunoelectron microscopy for MHC class II molecules, CD11c, CD86 and OX62 (a marker for rat DC subpopulation). Results demonstrated that DCs in normal periodontal ligament and periapical lesions consisted of two subpopulations: the subpopulations differently expressed CD11c and OX62 and might differ in lineage, state of maturation and function. In the induced periapical lesions, CD86-expressing cells, comprising approximately 10% of MHC class II molecule-expressing cells, were frequently distributed in the vicinity of nerve fibers, suggesting the involvement of mature DC-nerve interaction in the development of periapical lesions. Moreover, kinetics of DCs during wound healing process of exposed rat molar pulps was investigated after 1VITA-capping, which constantly induced pulp healing with dentin bridge formation. It was demonstrated that DC-like cells showed an accumulation subjacent to the wound surface during initial healing process of the exposed pulps.
    (2) Immunohistochemistry and electron microscopy for human apical periodontitis revealed that DCs expressing BLA-DR were mainly distributed in lymphocyte-rich areas. By means of laser microdisection and RT-PCR analysis, it was demonstrated that upregulation of CD83 and CD86 on DCs and CD28 on T lymphocytes occurred in the lymphocyte-rich areas. These findings may indicate that DCs act as antigen presenting cells that stimulate T lymphocyte activation.

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  • Evaluation of pulpal responses to GaAIAs laser irradiation.

    Grant number:16591909

    2004 - 2006

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

    Research category:Grant-in-Aid for Scientific Research (C)

    Awarding organization:Japan Society for the Promotion of Science

    YOSHIBA Nagako, YOSHIBA Kunihiko, TAKENAKA Shoji

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    Grant amount:\3500000 ( Direct Cost: \3500000 )

    This study aimed to evaluate the effect of the GaAIAs laser on dentin-pulp complex, in order to establish the potential applications of the laser for clinical practice in caries treatment.
    1. Immunohistochemical analysis of caries teeth.
    Major histocompatibility complex (MHC) class II molecule-expressing cells are distributed in human dental pulp, and have been show to accumulate beneath caries lesions. After caries treatment, clusters consisting of class II molecule-expressing dendritic cells, T-lymphocytes, and nerve fibers still remained locally in the subodontoblastic area. Under active and deep caries, a number of STRO-1-positive stem cells have been recognized along blood vessels.
    2. Responses of GaAIAs laser irradiation to dentin-pulp complex in rat molars.
    The medial surface of the molar was lased at an output power of 0.5-1.5 W for 180s. At 7days after irradiation, tertiary dentin has been formed. At 30 days, tertiary dentin with/without bone-like tissue was formed abundantly in the dental pulp. Statistical analysis revealed that the area occupied by the new hard tissues was significantly wider in 1.5W-lased speciments than in 0.5W-lased speciments.
    3. Responses of GaAIAs laser irradiation to dentin-pulp complex in human teeth.
    The volunteers, who had been scheduled to undergo extraction for various therapeutic reasons, were enrolled in the study. Informed consent was obtained from subjects after the proposed study was fully explained. Class V circular preparations were cut into the buccal surfaces, and the cavity was irradiated by GaAIAs laser, and then filled with composite resin. As a control, the cavity was filled without the laser irradiation. Under deep cavities, class II molecule-expressing dendritic cells displaced the injured odontoblasts in control, while few dendritic cells displaced the injured odontoblasts in the lased teeth.

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  • A study on mechanisms of tissue formation and regeneration of the dentin-pulp complex

    Grant number:16591910

    2004 - 2005

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

    Research category:Grant-in-Aid for Scientific Research (C)

    Awarding organization:Japan Society for the Promotion of Science

    YOSHIBA Kunihiko, YOSHIBA Nagako, HOSOYA Akihiro, OZAWA Hidehiro

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    Grant amount:\3600000 ( Direct Cost: \3600000 )

    This study aims to elucidate mechanisms of tissue formation and regeneration of the dentin-pulp complex. The results were as follows :
    1. During mouse tooth development, the expression of an intermediate filament nestin was intense in differentiating odontoblasts, and became restricted to the secretory pole of functional odontoblasts, suggesting that nestin plays a role in terminal differentiation, especially in cell elongation and/or polarization of odontoblasts.
    2. Distinct temporal and spatial expression patterns of TIMPs suggest divergent functions of these factors in incisor organogenesis. Intense expression of Timp-1 was detected in differentiating odontoblasts, while Timp-3 was expressed by subodontoblasts, suggesting their involvement in odontoblast differentiation.
    3. The unerupted rat incisor was composed of osteodentin, with numerous cells present in the anterior apex. The osteodentin was immunopositive for osteocalcin, bone sialoprotein, and dentin sialoprotein, with an immunolocalization pattern similar to that of dentin. These results indicate that osteodentin in the rat incisor possesses a dentin-like characteristics.
    4. We performed a histological and immunohistochemical evaluation of mineralized tissue induced by pulp transplantation into subcutaneous tissue. The results indicate that pulp cells are able to form hard tissue with bone- or cementum-like characteristics without dentin influence. Calcification may start in necrotic cells and matrix vesicles, followed by collagenous calcification.
    5. Pulpal responses to GaAlAs laser irradiation were elucidated. The results indicated that the GaAlAs laser may induce the formation of tertiary dentin by influencing the secretory activity of odontoblasts. However, higher energies may cause irreversible changes to the pulp, often leading to the formation of an intrapulpal bone-like tissue.

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  • A study on mechanisms of tissue repair and regeneration of the dentin-pulp complex

    Grant number:14571811

    2002 - 2003

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

    Research category:Grant-in-Aid for Scientific Research (C)

    Awarding organization:Japan Society for the Promotion of Science

    YOSHIBA Kunihiko, YOSHIBA Nagako, IWAKU Masaaki

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    Grant amount:\3900000 ( Direct Cost: \3900000 )

    This study aims to elucidate mechanisms of tissue repair and/or regeneration of the dentin-pulp complex. Several factors involved in cell differentiation during tooth development and reparative dentinogenesis were analyzed.
    The results were as follows :
    1.During mouse molar tooth development, MMP-2,-9,and MT1-MMP were expressed in the dental epithelium and mesenchyme. In contrast, TIMPs(1-3) were differentially expressed. The distinct temporospatial distribution patterns of the TIMPs suggest that these inhibitors play several intrinsic roles during tooth development.
    2.The odontoblast-like cells forming reparative dentin expressed TGF-βs, especially β2 and β3, and also TGF-β type II receptor, suggesting that cells that form reparative dentin express odontoblast phenotype and that TGF-βs may be involved in the differentiation of replacement odontoblasts during reparative dentinogenesis after pulp capping.
    3.Cavity preparation in normal teeth caused alteration in the distribution of MHC class II molecule-expressing cells, although this change appears to be reversible. By contrast, in the caries-removed and restored teeth, antigen presentation and cellular and/or humoral immunoresponses would seem to persist, even after treatment.
    4.The pulpal responses to the irradiation of tooth surfaces with an 810nm-GaAlAs semiconductor laser were evaluated in the rat molars. This laser at certain power settings could induce the formation of tertiary dentin without hard tissue injury nor evoking inflammatory reactions in the pulp.

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  • う蝕修復処置後の歯髄反応に関する免疫組織化学的研究

    Grant number:13771127

    2001 - 2002

    System name:科学研究費助成事業 若手研究(B)

    Research category:若手研究(B)

    Awarding organization:日本学術振興会

    吉羽 永子

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    Grant amount:\2100000 ( Direct Cost: \2100000 )

    本研究は、より生物学的な歯髄保護材開発のための基礎的研究の環として、免疫担当細胞、特に抗原提示細胞、炎症性細胞ならびに神経線維の分布、局在が歯髄疾患の病態を把握する指標となり得るかどうかを検討するとともに、う蝕修復処置後の歯髄治癒過程におけるこれらの動態を免疫組織化学的、微細構造学的に検索するものである。研究要旨の同意を得られた抜去適応の健全あるいはう蝕歯を用い、従来の臨床術式に則し、う蝕検知液下にてう蝕を可久的に除去し、また細菌感染のある歯髄組織の修復過程と比較するために健全歯を用い、様々な深さの窩洞を形成し、レジン充填処置を施し、一定期間経過後抜去して試料を得て検討し、以下の結果が得られた。
    1)健全歯:窩洞の深さによる段階的な象牙芽細胞の変化、それに伴う樹枝状細胞の局在変化が観察された。深さ二分の一までは、著しい変化は見られないものの、それを越えると急激に変化が認められ、三分の二をこえると、象牙芽細胞が消失し代わりに樹枝状細胞が配列していた。
    2)う蝕歯:表層が再石灰化していると思われた平滑面エナメルう蝕下においても、樹枝状細胞ならび神経線維の集積がすでに観察され、象牙芽細胞の変化もみとめられ七他の免疫細胞の局在には変化は認められなかった。
    3)う蝕歯処置後:う蝕を除去した後でも局所的な樹枝状細胞、T細胞、神経線維の集積は認められ、深部う蝕ではB細胞も観察され、樹枝状細胞とリンパ球の頻繁な接合も認められた。画像解析による統計学的検討により、樹枝状細胞のう蝕前後の変化に有意差は認められなかった。しかしながら窩洞の深さによらず象牙芽細胞は常に認められ、健全歯で観察されたような樹枝状細胞による置換は認められなかった。
    全症例において、窩縁相当部には樹枝状細胞の集積は認められないことから、マイクロリーケージはおこっていないと考えられる。さらに、長期例、修復材料による組織反応の違いについて、検討中である。

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  • A study on mechanisms of odontoblast differentiation during reparative dentinogenesis.

    Grant number:10671789

    1998 - 2000

    System name:Grants-in-Aid for Scientific Research Grant-in-Aid for Scientific Research (C)

    Research category:Grant-in-Aid for Scientific Research (C)

    Awarding organization:Japan Society for the Promotion of Science

    YOSHIBA Kunihiko, YOSHIBA Nagako, YAMAGA Masahiro

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    Grant amount:\3200000 ( Direct Cost: \3200000 )

    In order to elucidate mechanisms by which pulp cells differentiate into odontoblasts during reparative dentinogenesis, immunohistochemical analyses for several factors related to cell differentiation were performed. The results were as follows :
    1. During tooth development, differential expression of laminin-5 and integrin α6 and β4 subunits in the inner dental epithelium may be involved in ameloblast differentiation. These changes in basement membrane composition might be also related to odontoblast differentiation.
    2. In pulps affected by early caries, MHC class II antigen-expressing cells (HLA-DR-positive dendritic cells) aggregated mainly in the cell-free zone associated with PGP 9.5-immuno-reactive nerve fibers. In advanced caries, the accumulated HLA-DR-positive cells and PGP 9.5-immunoreactive nerve fibers showed close association with each other especially beneath the odontoblast layer. Class II molecules were recognized not only in dendritic cells but also in the Schwann cells of non-myelinated nerves in the pulp. These suggested a synergistic action between immune and nervous systems in pulpal inflammation and reparative process.
    3. Pulpal responses after cavity preparation with either Er : YAG laser or a conventional drill were evaluated. The immunoreactivity for tissue non-specific alkaline phosphatase was more pronounced in the laser group. Clear similarities in the distribution patterns of OX6-immunopositive cells and PGP 9.5- immunoreactive nerve fibers were noted. OX6-positive MHC class II antigen-expressing cells accumulated along the pulp-dentin border and numerous bead-like PGP 9.5-immunoreactive nerve fibers were observed under the odontoblastic layer, suggesting their involvement in pulp tissue repair and reparative dentin formation.
    4. During hard tissue formation in the pulp cavity of rat molar after transplantation into subcutaneous tissue, non-collagenous proteins, such as osteocalcin, osteopontin, bone sialoprotein and dentin sialoprotein showed different distribution patterns in newly-formed hard tissues in the coronal, root canal and apex areas, respectively. These suggested that each hard tissue was formed by differently-derived cells.

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  • 生体親和性歯髄保護材の開発に関する基礎的研究

    Grant number:97J08780

    1998 - 1999

    System name:科学研究費助成事業 特別研究員奨励費

    Research category:特別研究員奨励費

    Awarding organization:日本学術振興会

    吉羽 永子

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    Grant amount:\2400000 ( Direct Cost: \2400000 )

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  • 生体親和性材料に対する歯髄反応に関する研究

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    Grant type:Competitive

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  • 歯牙形成ならびに被蓋硬組織(dentin bridge)形成に関する研究

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    Grant type:Competitive

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  • 歯牙形成における上皮-間葉相互作用に関する研究

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    Grant type:Competitive

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  • Study on Pulpal Response to Biomaterials

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    Grant type:Competitive

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  • Study on tooth development and dentin bridge formation

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    Grant type:Competitive

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  • Epithelial-mesenchymal interactions during tooth development

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    Grant type:Competitive

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Teaching Experience

  • う蝕制御管理学演習IIA

    2022
    Institution name:新潟大学

  • う蝕制御管理学演習IIB

    2022
    Institution name:新潟大学

  • 歯科衛生士臨床実習II

    2021
    Institution name:新潟大学

  • 齲蝕学

    2020
    Institution name:新潟大学

  • 生涯にわたる歯と咬合

    2019
    Institution name:新潟大学

  • う蝕制御管理学演習ⅠA

    2017
    -
    2018
    Institution name:新潟大学

  • う蝕制御管理学演習ⅠB

    2017
    Institution name:新潟大学

  • エンドドンティックス

    2012
    -
    2013
    Institution name:新潟大学

  • 歯冠修復学

    2011
    -
    2014
    Institution name:新潟大学

  • 歯科診療補助Ⅱ

    2010
    Institution name:新潟大学

  • 統合科目Ⅱ

    2008
    -
    2019
    Institution name:新潟大学

  • 保存修復学実習

    2007
    Institution name:新潟大学

  • う蝕学

    2007
    -
    2018
    Institution name:新潟大学

  • 保存修復学

    2007
    -
    2018
    Institution name:新潟大学

  • 歯内療法学実習

    2007
    -
    2009
    Institution name:新潟大学

  • 統合科目II

    2007
    Institution name:新潟大学

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