Pub Date : 2025-01-30DOI: 10.1016/j.job.2025.100622
Naoki Miyasaka , Daisuke Torii , Takafumi Satomi , Kenichi Sakurai , Taka Nakahara , Takeo W. Tsutsui
Objectives
This study aimed to investigate the effects of aspirin on the early stages of odontogenic differentiation. The roles of FOXC1, RUNX2, and MCAM gene expression in the mechanism of odontogenic differentiation were evaluated by examining the effects of downregulated FOXC1 or RUNX2 expression using small interfering RNAs (siRNAs).
Methods
Dental pulp cells were treated with aspirin (0, 2.5, 50, 100 μg/ml) to assess its impact on mineralization. The gene expression levels of FOXC1, RUNX2, and MCAM were measured using digital polymerase chain reaction, and the effects of siRNA-mediated knockdown of FOXC1 and RUNX2 were analyzed. The mineralization potential was quantitatively assessed using Alizarin Red S staining and a calcium assay.
Results
Analysis of cell growth curves and doubling times indicated that aspirin did not affect cell proliferation at 2.5 μg/ml and 50 μg/ml; however, 50 μg/ml aspirin promoted mineralization. In the FOXC1 and RUNX2 knockdown experiments, fluctuations in FOXC1, RUNX2, and MCAM gene expression were observed in the aspirin-treated group, suggesting the involvement of these genes in mineralization. Alizarin red S staining and calcium assays further demonstrated that aspirin enhanced mineralization.
Conclusions
These findings indicate that aspirin promotes odontogenic differentiation and regulates the expression of FOXC1, RUNX2, and MCAM. This suggests that aspirin may serve as a promising new therapeutic agent in dental pulp regenerative medicine.
{"title":"Aspirin promotes odontogenic differentiation via a mechanism involving FOXC1, RUNX2, and MCAM expression","authors":"Naoki Miyasaka , Daisuke Torii , Takafumi Satomi , Kenichi Sakurai , Taka Nakahara , Takeo W. Tsutsui","doi":"10.1016/j.job.2025.100622","DOIUrl":"10.1016/j.job.2025.100622","url":null,"abstract":"<div><h3>Objectives</h3><div>This study aimed to investigate the effects of aspirin on the early stages of odontogenic differentiation. The roles of <em>FOXC1</em>, <em>RUNX2</em>, and <em>MCAM</em> gene expression in the mechanism of odontogenic differentiation were evaluated by examining the effects of downregulated <em>FOXC1</em> or <em>RUNX2</em> expression using small interfering RNAs (siRNAs).</div></div><div><h3>Methods</h3><div>Dental pulp cells were treated with aspirin (0, 2.5, 50, 100 μg/ml) to assess its impact on mineralization. The gene expression levels of <em>FOXC1</em>, <em>RUNX2</em>, and <em>MCAM</em> were measured using digital polymerase chain reaction, and the effects of siRNA-mediated knockdown of <em>FOXC1</em> and <em>RUNX2</em> were analyzed. The mineralization potential was quantitatively assessed using Alizarin Red S staining and a calcium assay.</div></div><div><h3>Results</h3><div>Analysis of cell growth curves and doubling times indicated that aspirin did not affect cell proliferation at 2.5 μg/ml and 50 μg/ml; however, 50 μg/ml aspirin promoted mineralization. In the <em>FOXC1</em> and <em>RUNX2</em> knockdown experiments, fluctuations in <em>FOXC1</em>, <em>RUNX2</em>, and <em>MCAM</em> gene expression were observed in the aspirin-treated group, suggesting the involvement of these genes in mineralization. Alizarin red S staining and calcium assays further demonstrated that aspirin enhanced mineralization.</div></div><div><h3>Conclusions</h3><div>These findings indicate that aspirin promotes odontogenic differentiation and regulates the expression of <em>FOXC1</em>, <em>RUNX2</em>, and <em>MCAM</em>. This suggests that aspirin may serve as a promising new therapeutic agent in dental pulp regenerative medicine.</div></div>","PeriodicalId":45851,"journal":{"name":"Journal of Oral Biosciences","volume":"67 1","pages":"Article 100622"},"PeriodicalIF":2.6,"publicationDate":"2025-01-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143075455","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-01-30DOI: 10.1016/j.job.2025.100621
Tingting Xu , Yi Peng , Yanan Xu , Jing Zhu , Qiao Yang , Yali Liu , Hefeng Yang
Objectives
To investigate the role of small extracellular vesicles derived from induced pluripotent stem cells (iPSC-sEVs) in periodontal tissue regeneration, elucidate their potential molecular mechanisms, and provide theoretical guidance for the clinical application of iPSC-sEVs as a cell-free therapeutic strategy for periodontal tissue regeneration.
Methods
We investigated the effects of iPSC-sEVs on the proliferation, migration, and osteogenic differentiation of periodontal ligament stem cells (PDLSCs) in vitro. The regenerative potential of iPSC-sEVs was evaluated in vivo, using a periodontal defect model. Bulk RNA sequencing was performed to elucidate the underlying molecular mechanisms.
Results
iPSC-sEVs were isolated, characterized, and systemically evaluated for regenerative potential. The results revealed that treatment with iPSC-sEVs significantly enhanced the proliferation, migration, and osteogenic differentiation of PDLSCs. In situ treatment with iPSC-sEVs loaded onto collagen sponges was performed in a rat model of periodontal defects. Micro-CT and histological analyses indicated that iPSC-sEV treatment markedly promoted alveolar bone repair and periodontal ligament regeneration. Mechanistically, the analysis of bulk RNA sequencing data coupled with experimental validation revealed that iPSC-sEV treatment significantly activated the mitogen-activated protein kinase (MAPK) signaling pathway in PDLSCs. Further investigation showed that the inhibition of this pathway completely abolished the proliferative effects of iPSC-sEVs on PDLSCs.
Conclusions
iPSC-sEVs promote PDLSC proliferation through MAPK signaling pathway activation, while also enhancing PDLSC migratory and osteogenic differentiation capacities, facilitates the repair and regeneration of damaged periodontal tissue and presents a potential novel therapeutic strategy for clinical periodontal tissue regeneration.
{"title":"Exploring the therapeutic potential of small extracellular vesicles derived from induced pluripotent stem cell in periodontal regeneration","authors":"Tingting Xu , Yi Peng , Yanan Xu , Jing Zhu , Qiao Yang , Yali Liu , Hefeng Yang","doi":"10.1016/j.job.2025.100621","DOIUrl":"10.1016/j.job.2025.100621","url":null,"abstract":"<div><h3>Objectives</h3><div>To investigate the role of small extracellular vesicles derived from induced pluripotent stem cells (iPSC-sEVs) in periodontal tissue regeneration, elucidate their potential molecular mechanisms, and provide theoretical guidance for the clinical application of iPSC-sEVs as a cell-free therapeutic strategy for periodontal tissue regeneration.</div></div><div><h3>Methods</h3><div>We investigated the effects of iPSC-sEVs on the proliferation, migration, and osteogenic differentiation of periodontal ligament stem cells (PDLSCs) in vitro. The regenerative potential of iPSC-sEVs was evaluated in vivo, using a periodontal defect model. Bulk RNA sequencing was performed to elucidate the underlying molecular mechanisms.</div></div><div><h3>Results</h3><div>iPSC-sEVs were isolated, characterized, and systemically evaluated for regenerative potential. The results revealed that treatment with iPSC-sEVs significantly enhanced the proliferation, migration, and osteogenic differentiation of PDLSCs. In situ treatment with iPSC-sEVs loaded onto collagen sponges was performed in a rat model of periodontal defects. Micro-CT and histological analyses indicated that iPSC-sEV treatment markedly promoted alveolar bone repair and periodontal ligament regeneration. Mechanistically, the analysis of bulk RNA sequencing data coupled with experimental validation revealed that iPSC-sEV treatment significantly activated the mitogen-activated protein kinase (MAPK) signaling pathway in PDLSCs. Further investigation showed that the inhibition of this pathway completely abolished the proliferative effects of iPSC-sEVs on PDLSCs.</div></div><div><h3>Conclusions</h3><div>iPSC-sEVs promote PDLSC proliferation through MAPK signaling pathway activation, while also enhancing PDLSC migratory and osteogenic differentiation capacities, facilitates the repair and regeneration of damaged periodontal tissue and presents a potential novel therapeutic strategy for clinical periodontal tissue regeneration.</div></div>","PeriodicalId":45851,"journal":{"name":"Journal of Oral Biosciences","volume":"67 1","pages":"Article 100621"},"PeriodicalIF":2.6,"publicationDate":"2025-01-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143075629","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
This study investigated the effects of thread design on the soft and hard tissues around implants in rat maxillary peri-implantitis-like lesions.
Methods
Fourteen, 9-week-old, female Wistar rats were used in this study. Two types of grade IV titanium tissue-level implants with a standard V-shape and buttress threads were prepared (control and test implants, respectively). The control and test implants were randomly placed into healed left or right sides four weeks after first molar extraction. Daily administration of lipopolysaccharide (LPS) into the peri-implant mucosal sulcus was performed in combination with Freund's incomplete and complete adjuvants. The maxillae were harvested 16 days after LPS administration for quantitative and qualitative analyses.
Results
LPS administration induced significant marginal bone loss, with increases in osteoclasts and polymorphonuclear cells around control implants. LPS administration did not change cell numbers around nor alter bone quality inside the buttress threads of the test implants, but resulted in a significant deterioration of bone quality, defined as the preferential alignment of collagen fibers inside the V-shaped threads of the control implant. LPS administration also significantly increased calprotectin production in the epithelium around the test implants and significantly increased calprotectin production in the connective tissue around both the control and test implants.
Conclusions
Buttress threads at specific angles provided resistance to LPS-induced inflammation in rats with LPS-induced peri-implantitis-like lesions. The upregulated production of calprotectin induced by LPS administration in the epithelium and connective tissues around the test implants may facilitate inflammation control around implants.
{"title":"Effects of thread design on soft and hard tissue healing around implants in lipopolysaccharide-induced peri-implantitis-like lesions in rat maxillae","authors":"Tomohiro Ishizaki , Yusuke Uto , Nao Inaba , Fumika Tsuda , Shinichiro Kuroshima , Takashi Sawase","doi":"10.1016/j.job.2025.100620","DOIUrl":"10.1016/j.job.2025.100620","url":null,"abstract":"<div><h3>Objectives</h3><div>This study investigated the effects of thread design on the soft and hard tissues around implants in rat maxillary peri-implantitis-like lesions.</div></div><div><h3>Methods</h3><div>Fourteen, 9-week-old, female Wistar rats were used in this study. Two types of grade IV titanium tissue-level implants with a standard V-shape and buttress threads were prepared (control and test implants, respectively). The control and test implants were randomly placed into healed left or right sides four weeks after first molar extraction. Daily administration of lipopolysaccharide (LPS) into the peri-implant mucosal sulcus was performed in combination with Freund's incomplete and complete adjuvants. The maxillae were harvested 16 days after LPS administration for quantitative and qualitative analyses.</div></div><div><h3>Results</h3><div>LPS administration induced significant marginal bone loss, with increases in osteoclasts and polymorphonuclear cells around control implants. LPS administration did not change cell numbers around nor alter bone quality inside the buttress threads of the test implants, but resulted in a significant deterioration of bone quality, defined as the preferential alignment of collagen fibers inside the V-shaped threads of the control implant. LPS administration also significantly increased calprotectin production in the epithelium around the test implants and significantly increased calprotectin production in the connective tissue around both the control and test implants.</div></div><div><h3>Conclusions</h3><div>Buttress threads at specific angles provided resistance to LPS-induced inflammation in rats with LPS-induced peri-implantitis-like lesions. The upregulated production of calprotectin induced by LPS administration in the epithelium and connective tissues around the test implants may facilitate inflammation control around implants.</div></div>","PeriodicalId":45851,"journal":{"name":"Journal of Oral Biosciences","volume":"67 1","pages":"Article 100620"},"PeriodicalIF":2.6,"publicationDate":"2025-01-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143041431","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Systemic administration of conditioned medium (CM) from stem cells derived from human exfoliated deciduous teeth (SHED-CM) in mouse models of rheumatoid arthritis, osteoporosis, and osteoarthritis suppresses excessive osteoclast activity and restores bone integrity. However, the mechanism through which SHED-CM regulates osteoclastogenesis remains largely unknown. In the present study, we examined the anti-osteoclastogenic mechanism of SHED-CM in vitro.
Methods
Bone marrow macrophages and RAW264.7 cells were treated with receptor activator of nuclear factor kappa-Β ligand (RANKL) in the presence of SHED-CM or CM from bone marrow mesenchymal stem cells (BMSC-CM). Osteoclast differentiation was assessed using tartrate-resistant acid phosphatase staining, actin ring formation, and expression of osteoclast-specific markers. RANKL-induced reactive oxygen species (ROS) production was analyzed as a critical mediator of osteoclastogenesis. The activation of endogenous antioxidant gene expression was examined using reverse transcription quantitative PCR. Liquid chromatography with tandem mass spectrometry (LC-MS) was used to identify proteins enriched in SHED-CM, and neutralizing antibodies were used to evaluate their functional roles.
Results
Compared to BMSC-CM, SHED-CM effectively inhibited RANKL-induced early osteoclast differentiation and late maturation. Notably, SHED-CM but not BMSC-CM suppressed RANKL-induced ROS production. SHED-CM increased the expression of genes encoding antioxidant enzymes. The LC-MS analysis identified seven proteins uniquely enriched in SHED-CM that activated the endogenous antioxidant system. Neutralizing antibodies against some of these proteins restore RANKL-induced ROS production and osteoclast differentiation.
Conclusions
SHED-CM inhibited osteoclastogenesis, partially through the activation of multiple antioxidant enzymes in osteoclast precursors, highlighting its potential for treating bone-destructive diseases.
{"title":"Factors secreted from the stem cells of human exfoliated deciduous teeth inhibit osteoclastogenesis through the activation of the endogenous antioxidant system","authors":"Cheng Ding , Noboru Hashimoto , Fumiya Kano , Hirofumi Tenshin , Takahiro Arai , Linze Xia , Yang Xu , Houjun Lao , Yifei Wang , Tomonori Iwasaki , Hideharu Hibi , Akihito Yamamoto","doi":"10.1016/j.job.2025.100618","DOIUrl":"10.1016/j.job.2025.100618","url":null,"abstract":"<div><h3>Objectives</h3><div>Systemic administration of conditioned medium (CM) from stem cells derived from human exfoliated deciduous teeth (SHED-CM) in mouse models of rheumatoid arthritis, osteoporosis, and osteoarthritis suppresses excessive osteoclast activity and restores bone integrity. However, the mechanism through which SHED-CM regulates osteoclastogenesis remains largely unknown. In the present study, we examined the anti-osteoclastogenic mechanism of SHED-CM in vitro.</div></div><div><h3>Methods</h3><div>Bone marrow macrophages and RAW264.7 cells were treated with receptor activator of nuclear factor kappa-Β ligand (RANKL) in the presence of SHED-CM or CM from bone marrow mesenchymal stem cells (BMSC-CM). Osteoclast differentiation was assessed using tartrate-resistant acid phosphatase staining, actin ring formation, and expression of osteoclast-specific markers. RANKL-induced reactive oxygen species (ROS) production was analyzed as a critical mediator of osteoclastogenesis. The activation of endogenous antioxidant gene expression was examined using reverse transcription quantitative PCR. Liquid chromatography with tandem mass spectrometry (LC-MS) was used to identify proteins enriched in SHED-CM, and neutralizing antibodies were used to evaluate their functional roles.</div></div><div><h3>Results</h3><div>Compared to BMSC-CM, SHED-CM effectively inhibited RANKL-induced early osteoclast differentiation and late maturation. Notably, SHED-CM but not BMSC-CM suppressed RANKL-induced ROS production. SHED-CM increased the expression of genes encoding antioxidant enzymes. The LC-MS analysis identified seven proteins uniquely enriched in SHED-CM that activated the endogenous antioxidant system. Neutralizing antibodies against some of these proteins restore RANKL-induced ROS production and osteoclast differentiation.</div></div><div><h3>Conclusions</h3><div>SHED-CM inhibited osteoclastogenesis, partially through the activation of multiple antioxidant enzymes in osteoclast precursors, highlighting its potential for treating bone-destructive diseases.</div></div>","PeriodicalId":45851,"journal":{"name":"Journal of Oral Biosciences","volume":"67 1","pages":"Article 100618"},"PeriodicalIF":2.6,"publicationDate":"2025-01-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143041438","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Temporomandibular joint osteoarthritis (TMJ-OA) with condylar resorption is a multifactorial condition involving hormonal imbalance and articular disk dysfunction, often leading to severe TMJ degeneration. This study investigated the combined effects of estrogen deficiency and anterior articular disk derangement (ADD) on condylar bone resorption in a mouse model.
Methods
Female C57BL/6J mice underwent ovariectomy (OVX) to induce estrogen deficiency and ADD was surgically induced for stress. The animals were divided into the control, OVX, ADD, and OVX + ADD groups. Microcomputed tomography and histological analyses were conducted to evaluate condylar bone structure, trabecular architecture, and osteoclast activity.
Results
OVX and ADD caused significant condylar bone loss, characterized by reduced bone volume per tissue volume (BV/TV) and abnormal trabecular architecture. The OVX + ADD group exhibited exacerbated bone resorption, with decreased BV/TV and increased trabecular separation compared to OVX or ADD alone. Histological analyses revealed increased osteoclast activity in the OVX + ADD group, suggesting a synergistic effect of estrogen deficiency and ADD on condylar degradation.
Conclusion
Estrogen deficiency amplifies the bone-resorptive and inflammatory effects of ADD, accelerates temporomandibular joint (TMJ) degeneration, and underscores the interplay between hormone imbalance and articular disk dysfunction in the pathophysiology of TMJ-OA. There is a need for integrated treatment strategies, such as effective hormone replacement therapy and articular disk repositioning, to effectively manage temporomandibular joint disorders, particularly in postmenopausal women or those with hormonal imbalances. Further research is required to elucidate these molecular pathways and evaluate long-term therapeutic interventions.
{"title":"Synergistic effects of estrogen deficiency and articular disk derangement on condylar bone loss","authors":"Kazuhiro Shibusaka , Soichiro Negishi , Nobuhiro Sakai , Youngkwan Kim , Hiroyuki Okada , Fumiko Yano","doi":"10.1016/j.job.2025.100616","DOIUrl":"10.1016/j.job.2025.100616","url":null,"abstract":"<div><h3>Objectives</h3><div>Temporomandibular joint osteoarthritis (TMJ-OA) with condylar resorption is a multifactorial condition involving hormonal imbalance and articular disk dysfunction, often leading to severe TMJ degeneration. This study investigated the combined effects of estrogen deficiency and anterior articular disk derangement (ADD) on condylar bone resorption in a mouse model.</div></div><div><h3>Methods</h3><div>Female C57BL/6J mice underwent ovariectomy (OVX) to induce estrogen deficiency and ADD was surgically induced for stress. The animals were divided into the control, OVX, ADD, and OVX + ADD groups. Microcomputed tomography and histological analyses were conducted to evaluate condylar bone structure, trabecular architecture, and osteoclast activity.</div></div><div><h3>Results</h3><div>OVX and ADD caused significant condylar bone loss, characterized by reduced bone volume per tissue volume (BV/TV) and abnormal trabecular architecture. The OVX + ADD group exhibited exacerbated bone resorption, with decreased BV/TV and increased trabecular separation compared to OVX or ADD alone. Histological analyses revealed increased osteoclast activity in the OVX + ADD group, suggesting a synergistic effect of estrogen deficiency and ADD on condylar degradation.</div></div><div><h3>Conclusion</h3><div>Estrogen deficiency amplifies the bone-resorptive and inflammatory effects of ADD, accelerates temporomandibular joint (TMJ) degeneration, and underscores the interplay between hormone imbalance and articular disk dysfunction in the pathophysiology of TMJ-OA. There is a need for integrated treatment strategies, such as effective hormone replacement therapy and articular disk repositioning, to effectively manage temporomandibular joint disorders, particularly in postmenopausal women or those with hormonal imbalances. Further research is required to elucidate these molecular pathways and evaluate long-term therapeutic interventions.</div></div>","PeriodicalId":45851,"journal":{"name":"Journal of Oral Biosciences","volume":"67 1","pages":"Article 100616"},"PeriodicalIF":2.6,"publicationDate":"2025-01-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143041514","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Low-level laser therapy (LLLT) using an erbium-doped yttrium aluminum garnet (Er:YAG) laser provides a non-invasive approach applicable to various dental treatments. Here, we investigated the effects of Er:YAG laser irradiation on human dental pulp stem cells (hDPSCs) in an in vitro experiment.
Methods
The hDPSCs were categorized into four groups: laser-irradiated with activators (VLT: activated vitamin D3, bone morphogenetic protein receptor inhibitor, and transforming growth factor-beta (TGF-β)) (LLLT(+)VLT), laser-irradiated without activators (LLLT(+)-only), non-irradiated with activators (LLLT(−)VLT), and non-irradiated without activators (control). Cell proliferation, hard tissue differentiation, TGF-β signaling pathway activity, mineralization induction, and gene expression levels were assessed using several approaches, including cell proliferation assays, ALP assays, western blotting, Alizarin Red S staining, X-ray diffraction, and quantitative polymerase chain reaction.
Results
Cell proliferation was similar between the LLLT(+)-only and control groups. The ALP activity was significantly higher in LLLT(+)VLT group than in LLLT(−)VLT group (p < 0.05); however, it was suppressed by TGF-β signaling inhibitors. Western blotting showed enhanced SMAD3 phosphorylation in the LLLT(+)VLT group. The mineralization nodules and mRNA levels of matrix vesicle marker genes were significantly higher in LLLT(+)VLT group, and the nodules were partially composed of hydroxyapatite. The hard tissue formation marker gene expression in LLLT(+)VLT group was significantly higher (p < 0.05) than that in the LLLT(+)-only and control groups; however, it was unchanged or suppressed compared with that in LLLT(−)VLT group.
Conclusions
LLLT using an Er:YAG laser, combined with VLT, may promote the differentiation of hDPSCs into hard tissue-forming cells and enhance mineralization.
{"title":"Enhancement of differentiation and mineralization of human dental pulp stem cells via TGF-β signaling in low-level laser therapy using Er:YAG lasers","authors":"Ryo Yoshida , Kazuyuki Kobayashi , Kazuo Onuma , Ryuji Yamamoto , Risako Chiba-Ohkuma , Takeo Karakida , Shunjiro Yamakawa , Noriyasu Hosoya , Yasushi Yamazaki , Yasuo Yamakoshi","doi":"10.1016/j.job.2025.100617","DOIUrl":"10.1016/j.job.2025.100617","url":null,"abstract":"<div><h3>Objectives</h3><div>Low-level laser therapy (LLLT) using an erbium-doped yttrium aluminum garnet (Er:YAG) laser provides a non-invasive approach applicable to various dental treatments. Here, we investigated the effects of Er:YAG laser irradiation on human dental pulp stem cells (hDPSCs) in an <em>in vitro</em> experiment.</div></div><div><h3>Methods</h3><div>The hDPSCs were categorized into four groups: laser-irradiated with activators (VLT: activated vitamin D<sub>3</sub>, bone morphogenetic protein receptor inhibitor, and transforming growth factor-beta (TGF-β)) (LLLT(+)VLT), laser-irradiated without activators (LLLT(+)-only), non-irradiated with activators (LLLT(−)VLT), and non-irradiated without activators (control). Cell proliferation, hard tissue differentiation, TGF-β signaling pathway activity, mineralization induction, and gene expression levels were assessed using several approaches, including cell proliferation assays, ALP assays, western blotting, Alizarin Red S staining, X-ray diffraction, and quantitative polymerase chain reaction.</div></div><div><h3>Results</h3><div>Cell proliferation was similar between the LLLT(+)-only and control groups. The ALP activity was significantly higher in LLLT(+)VLT group than in LLLT(−)VLT group (<em>p</em> < 0.05); however, it was suppressed by TGF-β signaling inhibitors. Western blotting showed enhanced SMAD3 phosphorylation in the LLLT(+)VLT group. The mineralization nodules and mRNA levels of matrix vesicle marker genes were significantly higher in LLLT(+)VLT group, and the nodules were partially composed of hydroxyapatite. The hard tissue formation marker gene expression in LLLT(+)VLT group was significantly higher (<em>p</em> < 0.05) than that in the LLLT(+)-only and control groups; however, it was unchanged or suppressed compared with that in LLLT(−)VLT group.</div></div><div><h3>Conclusions</h3><div>LLLT using an Er:YAG laser, combined with VLT, may promote the differentiation of hDPSCs into hard tissue-forming cells and enhance mineralization.</div></div>","PeriodicalId":45851,"journal":{"name":"Journal of Oral Biosciences","volume":"67 1","pages":"Article 100617"},"PeriodicalIF":2.6,"publicationDate":"2025-01-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143013839","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-01-16DOI: 10.1016/j.job.2025.100615
Meili Mu , Hiroshi Inoue , Dan Mao , Nagako Sougawa , Seiji Goda
Objectives
Interleukin-8 (IL-8), a proinflammatory factor in human tissues, plays an important role in inflammation. Type IV collagen, a key component of the basement membrane, interacts with integrins, which are primary receptors in the extracellular matrix (ECM). Integrins are essential for the regulation of various cellular behaviors and signal transduction pathways. However, the relationship between type IV collagen, β1 integrin, and gingival epithelial cells is poorly understood. The aim in this study was to elucidate the effect of the interaction between type IV collagen and β1 integrin on IL-8 secretion in human gingival epithelial cells (Ca9-22).
Methods
Ca9-22 cells were treated with or without type IV collagen, and IL-8 production was assessed using an enzyme-linked immunosorbent assay (ELISA). The role of β1 integrin was investigated using a β1 integrin-neutralizing antibody. Western blotting was performed to measure the phosphorylation levels of the relevant proteins. The effects of the focal adhesion kinase (FAK) inhibitor Y15 and the MEK inhibitor U0126 on β1 integrin/FAK and Erk1/2 MAPK pathways in IL-8 production were evaluated to explore the involvement of these signaling pathways.
Results
β1 integrin induced IL-8 secretion in the Ca9-22 cells by regulating FAK, Erk1/2, and p130Cas proteins. p130Cas was independent of FAK, whereas Erk1/2 functioned downstream of FAK. Inhibition of FAK or Erk1/2 substantially reduced IL-8 secretion, highlighting their pivotal roles in this signaling pathway.
Conclusion
β1 integrin promotes IL-8 secretion in Ca9-22 cells via the β1 integrin/FAK/Erk1/2 signaling pathway. These findings elucidate the pathogenesis of periodontitis and provide a foundation for the development of targeted therapeutic strategies.
{"title":"β1 Integrin/FAK signaling regulates interleukin-8 production in human gingival epithelial Ca9-22 cells","authors":"Meili Mu , Hiroshi Inoue , Dan Mao , Nagako Sougawa , Seiji Goda","doi":"10.1016/j.job.2025.100615","DOIUrl":"10.1016/j.job.2025.100615","url":null,"abstract":"<div><h3>Objectives</h3><div>Interleukin-8 (IL-8), a proinflammatory factor in human tissues, plays an important role in inflammation. Type IV collagen, a key component of the basement membrane, interacts with integrins, which are primary receptors in the extracellular matrix (ECM). Integrins are essential for the regulation of various cellular behaviors and signal transduction pathways. However, the relationship between type IV collagen, β1 integrin, and gingival epithelial cells is poorly understood. The aim in this study was to elucidate the effect of the interaction between type IV collagen and β1 integrin on IL-8 secretion in human gingival epithelial cells (Ca9-22).</div></div><div><h3>Methods</h3><div>Ca9-22 cells were treated with or without type IV collagen, and IL-8 production was assessed using an enzyme-linked immunosorbent assay (ELISA). The role of β1 integrin was investigated using a β1 integrin-neutralizing antibody. Western blotting was performed to measure the phosphorylation levels of the relevant proteins. The effects of the focal adhesion kinase (FAK) inhibitor Y15 and the MEK inhibitor U0126 on β1 integrin/FAK and Erk1/2 MAPK pathways in IL-8 production were evaluated to explore the involvement of these signaling pathways.</div></div><div><h3>Results</h3><div>β1 integrin induced IL-8 secretion in the Ca9-22 cells by regulating FAK, Erk1/2, and p130Cas proteins. p130Cas was independent of FAK, whereas Erk1/2 functioned downstream of FAK. Inhibition of FAK or Erk1/2 substantially reduced IL-8 secretion, highlighting their pivotal roles in this signaling pathway.</div></div><div><h3>Conclusion</h3><div>β1 integrin promotes IL-8 secretion in Ca9-22 cells via the β1 integrin/FAK/Erk1/2 signaling pathway. These findings elucidate the pathogenesis of periodontitis and provide a foundation for the development of targeted therapeutic strategies.</div></div>","PeriodicalId":45851,"journal":{"name":"Journal of Oral Biosciences","volume":"67 1","pages":"Article 100615"},"PeriodicalIF":2.6,"publicationDate":"2025-01-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143013845","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Exposure of gingival epithelial cells to butyrate, a short-chain fatty acid produced by dental plaque bacteria, cause cell death and subsequent damage-associated molecular pattern (DAMP) release. We investigated the effects of curcumin, a polyphenol extracted from turmeric, on butyrate-induced human gingival epithelial Ca9-22 cell death and DAMP release.
Methods
Ca9-22 cells were pretreated with curcumin before butyrate exposure. Cell death was quantified using SYTOX green dye, and histone H3 acetylation was analyzed by Western blot. Conditioned media were collected to detect DAMPs by Western blot. We also assessed the effects of the histone acetyltransferase (HAT) inhibitor C646, instead of curcumin, on butyrate-induced cell death, DAMP release, and histone H3 acetylation, and examined the effects of curcumin pretreatment on cell death, DAMP release, and histone H3 acetylation induced by the histone deacetylase (HDAC) inhibitors, valproate and suberoylanilide hydroxamic acid (SAHA).
Results
Curcumin pretreatment attenuated butyrate-induced Ca9-22 cell death, histone H3 acetylation, and release of the DAMPs. The C646 also attenuated butyrate-induced cell death, DAMP release, and histone H3 acetylation. Curcumin also suppressed cell death, DAMP release, and histone H3 acetylation triggered by the HDAC inhibitors (valproate and SAHA).
Conclusions
Curcumin pretreatment ameliorated butyrate-induced histone H3 acetylation, cell death, and DAMP release. As elevated histone acetylation by HDAC inhibitors correlates with increased cell death, while reduced acetylation by a HAT inhibitor is associated with their attenuation, protective effects of curcumin against butyrate-induced Ca9-22 cell death and subsequent DAMP release may occur via suppression of histone acetylation.
{"title":"Curcumin pretreatment prevents butyrate-induced cell death and release of damage-associated molecular patterns on gingival epithelial Ca9-22 cells","authors":"Takayuki Hirasawa , Kiwa Miyake , Keiji Shinozuka , Yoshiyuki Yonehara , Hiromasa Tsuda","doi":"10.1016/j.job.2025.100613","DOIUrl":"10.1016/j.job.2025.100613","url":null,"abstract":"<div><h3>Objectives</h3><div>Exposure of gingival epithelial cells to butyrate, a short-chain fatty acid produced by dental plaque bacteria, cause cell death and subsequent damage-associated molecular pattern (DAMP) release. We investigated the effects of curcumin, a polyphenol extracted from turmeric, on butyrate-induced human gingival epithelial Ca9-22 cell death and DAMP release.</div></div><div><h3>Methods</h3><div>Ca9-22 cells were pretreated with curcumin before butyrate exposure. Cell death was quantified using SYTOX green dye, and histone H3 acetylation was analyzed by Western blot. Conditioned media were collected to detect DAMPs by Western blot. We also assessed the effects of the histone acetyltransferase (HAT) inhibitor C646, instead of curcumin, on butyrate-induced cell death, DAMP release, and histone H3 acetylation, and examined the effects of curcumin pretreatment on cell death, DAMP release, and histone H3 acetylation induced by the histone deacetylase (HDAC) inhibitors, valproate and suberoylanilide hydroxamic acid (SAHA).</div></div><div><h3>Results</h3><div>Curcumin pretreatment attenuated butyrate-induced Ca9-22 cell death, histone H3 acetylation, and release of the DAMPs. The C646 also attenuated butyrate-induced cell death, DAMP release, and histone H3 acetylation. Curcumin also suppressed cell death, DAMP release, and histone H3 acetylation triggered by the HDAC inhibitors (valproate and SAHA).</div></div><div><h3>Conclusions</h3><div>Curcumin pretreatment ameliorated butyrate-induced histone H3 acetylation, cell death, and DAMP release. As elevated histone acetylation by HDAC inhibitors correlates with increased cell death, while reduced acetylation by a HAT inhibitor is associated with their attenuation, protective effects of curcumin against butyrate-induced Ca9-22 cell death and subsequent DAMP release may occur via suppression of histone acetylation.</div></div>","PeriodicalId":45851,"journal":{"name":"Journal of Oral Biosciences","volume":"67 1","pages":"Article 100613"},"PeriodicalIF":2.6,"publicationDate":"2025-01-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143013830","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-01-15DOI: 10.1016/j.job.2025.100614
Su-Kyung Son, Jung-Sun Moon, Dong-Wook Yang, Na-Ri Jung, Jee-Hae Kang, Bin-Na Lee, Sun-Hun Kim, Min-Seok Kim
Objectives
We investigated the involvement of FOXO3a in lipopolysaccharide (LPS)-induced inflammation in primary human dental pulp cells (HDPCs).
Methods
HDPCs that were isolated from donors undergoing tooth extraction for orthodontic purposes were cultured with or without 1 μg/mL LPS at various intervals. The FOXO3a localization in the HDPCs was verified using immunofluorescence. Proinflammatory cytokines, such as interleukin (IL) 1β, IL6, and IL8, as well as their underlying mechanisms were assessed by observing gene and protein expressions through quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot analyses.
Results
LPS treatment enhanced the expressions of IL1β, IL6, and IL8 in HDPCs, concurrently activating nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB). Furthermore, FOXO3a expression was higher in the LPS-stimulated HDPCs, as confirmed by immunofluorescence localization. The results of loss-/gain-of-function approaches confirmed the regulatory role of FOXO3a in inflammatory HDPCs. FOXO3a knockdown attenuated proinflammatory cytokine expression; FOXO3a overexpression augmented their expression levels. FOXO3a inhibited retinoic acid receptor-related orphan receptor alpha (RORα) expression, thereby inactivating NFκB.
Conclusion
Our findings suggest that FOXO3a contributes to homeostasis in HDPCs through modulating the expression of proinflammatory cytokines under inflammatory conditions.
{"title":"Role of FOXO3a in LPS-induced inflammatory conditions in human dental pulp cells","authors":"Su-Kyung Son, Jung-Sun Moon, Dong-Wook Yang, Na-Ri Jung, Jee-Hae Kang, Bin-Na Lee, Sun-Hun Kim, Min-Seok Kim","doi":"10.1016/j.job.2025.100614","DOIUrl":"10.1016/j.job.2025.100614","url":null,"abstract":"<div><h3>Objectives</h3><div>We investigated the involvement of FOXO3a in lipopolysaccharide (LPS)-induced inflammation in primary human dental pulp cells (HDPCs).</div></div><div><h3>Methods</h3><div>HDPCs that were isolated from donors undergoing tooth extraction for orthodontic purposes were cultured with or without 1 μg/mL LPS at various intervals. The FOXO3a localization in the HDPCs was verified using immunofluorescence. Proinflammatory cytokines, such as interleukin (IL) 1β, IL6, and IL8, as well as their underlying mechanisms were assessed by observing gene and protein expressions through quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot analyses.</div></div><div><h3>Results</h3><div>LPS treatment enhanced the expressions of IL1β, IL6, and IL8 in HDPCs, concurrently activating nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB). Furthermore, FOXO3a expression was higher in the LPS-stimulated HDPCs, as confirmed by immunofluorescence localization. The results of loss-/gain-of-function approaches confirmed the regulatory role of FOXO3a in inflammatory HDPCs. FOXO3a knockdown attenuated proinflammatory cytokine expression; FOXO3a overexpression augmented their expression levels. FOXO3a inhibited retinoic acid receptor-related orphan receptor alpha (RORα) expression, thereby inactivating NFκB.</div></div><div><h3>Conclusion</h3><div>Our findings suggest that FOXO3a contributes to homeostasis in HDPCs through modulating the expression of proinflammatory cytokines under inflammatory conditions.</div></div>","PeriodicalId":45851,"journal":{"name":"Journal of Oral Biosciences","volume":"67 1","pages":"Article 100614"},"PeriodicalIF":2.6,"publicationDate":"2025-01-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143013840","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-01-11DOI: 10.1016/j.job.2025.100612
Mako Sakakibara , Tomoka Hasegawa , Mai Haraguchi-Kitakamae , Yan Shi , Weisong Li , Jiaxin Cui , Xuanyu Liu , Tomomaya Yamamoto , Hiromi Hongo , Norio Amizuka , Yoshiaki Sato , Masanori Kikuchi
Objectives
Hydroxyapatite (HAp)/collagen (Col) cylinders with laminated collagen layers were implanted into the tibial diaphysis of rats and examined histochemically to clarify how the orientation of HAp and Col bone-like nanocomposite fibers in HAp/Col blocks affects bone resorption and formation.
Methods
HAp/Col fibers were synthesized and compressed into cylindrical blocks to mimic bone nanostructures. These were implanted into the cortical bone cavities of 10-week-old male Wistar rats with fiber bundles parallel to the tibial surface. The implants were histologically analyzed at 3, 5, 7, 14, and 28 days after implantation.
Results
TRAP-positive osteoclasts appeared after 3–5 days in the lateral region of the graft, where the fiber ends were exposed, but not in the bottom region, where the HAp/Col fibers were parallel to the surface. Osteoclasts were observed in both regions by day 14. PHOSPHO1-positive osteoblasts were first detected on day 5, appearing slightly away from the cylinder laterally but directly on the bottom surface. A few osteoblasts contacted the block laterally, whereas many were observed on the new bone tissue at the bottom, between days 7 and 14. Bone formation was induced earlier in the bottom region, whereas lateral resorption was dominant. This suggested the uncoupling of bone resorption and formation in the early postimplantation stages. However, bone remodeling shifted to coupling between osteoclasts and osteoblasts throughout the cylinder by day 28.
Conclusion
The orientation of HAp/Col fibers in HAp/Col graft materials substantially affected the preferential induction of bone resorption or formation during the early stages of bone regeneration.
{"title":"Histochemical analysis of osteoclast and osteoblast distributions on hydroxyapatite/collagen bone-like nanocomposite embedded in rat tibiae","authors":"Mako Sakakibara , Tomoka Hasegawa , Mai Haraguchi-Kitakamae , Yan Shi , Weisong Li , Jiaxin Cui , Xuanyu Liu , Tomomaya Yamamoto , Hiromi Hongo , Norio Amizuka , Yoshiaki Sato , Masanori Kikuchi","doi":"10.1016/j.job.2025.100612","DOIUrl":"10.1016/j.job.2025.100612","url":null,"abstract":"<div><h3>Objectives</h3><div>Hydroxyapatite (HAp)/collagen (Col) cylinders with laminated collagen layers were implanted into the tibial diaphysis of rats and examined histochemically to clarify how the orientation of HAp and Col bone-like nanocomposite fibers in HAp/Col blocks affects bone resorption and formation.</div></div><div><h3>Methods</h3><div>HAp/Col fibers were synthesized and compressed into cylindrical blocks to mimic bone nanostructures. These were implanted into the cortical bone cavities of 10-week-old male Wistar rats with fiber bundles parallel to the tibial surface. The implants were histologically analyzed at 3, 5, 7, 14, and 28 days after implantation.</div></div><div><h3>Results</h3><div>TRAP-positive osteoclasts appeared after 3–5 days in the lateral region of the graft, where the fiber ends were exposed, but not in the bottom region, where the HAp/Col fibers were parallel to the surface. Osteoclasts were observed in both regions by day 14. PHOSPHO1-positive osteoblasts were first detected on day 5, appearing slightly away from the cylinder laterally but directly on the bottom surface. A few osteoblasts contacted the block laterally, whereas many were observed on the new bone tissue at the bottom, between days 7 and 14. Bone formation was induced earlier in the bottom region, whereas lateral resorption was dominant. This suggested the uncoupling of bone resorption and formation in the early postimplantation stages. However, bone remodeling shifted to coupling between osteoclasts and osteoblasts throughout the cylinder by day 28.</div></div><div><h3>Conclusion</h3><div>The orientation of HAp/Col fibers in HAp/Col graft materials substantially affected the preferential induction of bone resorption or formation during the early stages of bone regeneration.</div></div>","PeriodicalId":45851,"journal":{"name":"Journal of Oral Biosciences","volume":"67 1","pages":"Article 100612"},"PeriodicalIF":2.6,"publicationDate":"2025-01-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142980205","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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