start-ver=1.4 cd-journal=joma no-vol=155 cd-vols= no-issue= article-no= start-page=105797 end-page= dt-received= dt-revised= dt-accepted= dt-pub-year=2023 dt-pub=202311 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=Ruxolitinib altered IFN-β induced necroptosis of human dental pulp stem cells during osteoblast differentiation en-subtitle= kn-subtitle= en-abstract= kn-abstract=Objective: This study aimed to evaluate the role of ruxolitinib in the interferon beta (IFN-β) mediated osteoblast differentiation using human dental pulp stem cells (hDPSCs).
Design: hDPSCs from five deciduous teeth of healthy patients were stimulated by adding human recombinant IFN-β protein (1 or 2 ng/ml) to the osteogenic differentiation induction medium. Substrate formation was determined using Alizarin Red staining, calcium concentration, and osteoblast marker expression levels. Ruxolitinib was used to inhibit the Janus kinase/signal transducers and activators of transcription (JAK-STAT) pathway. Apoptosis was detected using terminal deoxynucleotidyl nick-end labeling (TUNEL) staining, and necroptosis was detected using propidium iodide staining and phosphorylated mixed lineage kinase domain-like protein (pMLKL) expression.
Results: In the IFN-β-treated group, substrate formation was inhibited by a reduction in alkaline phosphatase (ALP) expression in a concentration-dependent manner. Although the proliferation potency was unchanged between the IFN-β-treated and control groups, the cell number was significantly reduced in the experimental group. TUNEL-positive cell number was not significantly different; however, the protein level of necroptosis markers, interleukin-6 (IL-6) and pMLKL were significantly increased in the substrate formation. Cell number and ALP expression level were improved in the group administered ruxolitinib, a JAK-STAT inhibitor. Additionally, ruxolitinib significantly suppressed IL-6 and pMLKL levels.
Conclusion: Ruxolitinib interfered with the IFN-β-mediated necroptosis and osteogenic differentiation via the JAK-STAT pathway. en-copyright= kn-copyright= en-aut-name=TanakaAtsuko en-aut-sei=Tanaka en-aut-mei=Atsuko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=HayanoSatoru en-aut-sei=Hayano en-aut-mei=Satoru kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=NagataMasayo en-aut-sei=Nagata en-aut-mei=Masayo kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=KosamiTakahiro en-aut-sei=Kosami en-aut-mei=Takahiro kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= en-aut-name=WangZiyi en-aut-sei=Wang en-aut-mei=Ziyi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=5 ORCID= en-aut-name=KamiokaHiroshi en-aut-sei=Kamioka en-aut-mei=Hiroshi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=6 ORCID= affil-num=1 en-affil=Department of Orthodontics, Okayama University Hospital kn-affil= affil-num=2 en-affil=Department of Orthodontics, Okayama University Hospital kn-affil= affil-num=3 en-affil=Department of Orthodontics, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University kn-affil= affil-num=4 en-affil=Department of Orthodontics, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University kn-affil= affil-num=5 en-affil=Department of Molecular Biology and Biochemistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences kn-affil= affil-num=6 en-affil=Department of Orthodontics, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama University kn-affil= en-keyword=Type-I interferon kn-keyword=Type-I interferon en-keyword=Janus kinase/signal transducers and activators of transcription pathway kn-keyword=Janus kinase/signal transducers and activators of transcription pathway en-keyword=Osteoblast kn-keyword=Osteoblast en-keyword=Necroptosis kn-keyword=Necroptosis en-keyword=Singleton-Merten Syndrome kn-keyword=Singleton-Merten Syndrome END start-ver=1.4 cd-journal=joma no-vol=2021 cd-vols= no-issue= article-no= start-page=1 end-page=11 dt-received= dt-revised= dt-accepted= dt-pub-year=2021 dt-pub=2021929 dt-online= en-article= kn-article= en-subject= kn-subject= en-title= kn-title=Investigation of the molecular causes underlying physical abnormalities in Diamond‐Blackfan anemia patients with RPL5 haploinsufficiency en-subtitle= kn-subtitle= en-abstract= kn-abstract=Diamond-Blackfan anemia (DBA) is a genetic disorder caused by mutations in genes encoding ribosomal proteins and characterized by erythroid aplasia and various physical abnormalities. Although accumulating evidence suggests that defective ribosome biogenesis leads to p53-mediated apoptosis in erythroid progenitor cells, little is known regarding the underlying causes of the physical abnormalities. In this study, we established induced pluripotent stem cells from a DBA patient with RPL5 haploinsufficiency. These cells retained the ability to differentiate into osteoblasts and chondrocytes. However, RPL5 haploinsufficiency impaired the production of mucins and increased apoptosis in differentiated chondrocytes. Increased expression of the pro-apoptotic genes BAX and CASP9 further indicated that RPL5 haploinsufficiency triggered p53-mediated apoptosis in chondrocytes. MDM2, the primary negative regulator of p53, plays a crucial role in erythroid aplasia in DBA patient. We found the phosphorylation level of MDM2 was significantly decreased in RPL5 haploinsufficient chondrocytes. In stark contrast, we found no evidence that RPL5 haploinsufficiency impaired osteogenesis. Collectively, our data support a model in which RPL5 haploinsufficiency specifically induces p53-mediated apoptosis in chondrocytes through MDM2 inhibition, which leads to physical abnormalities in DBA patients. en-copyright= kn-copyright= en-aut-name=FukuiYuko en-aut-sei=Fukui en-aut-mei=Yuko kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=1 ORCID= en-aut-name=HayanoSatoru en-aut-sei=Hayano en-aut-mei=Satoru kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=2 ORCID= en-aut-name=KawanabeNoriaki en-aut-sei=Kawanabe en-aut-mei=Noriaki kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=3 ORCID= en-aut-name=WangZiyi en-aut-sei=Wang en-aut-mei=Ziyi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=4 ORCID= en-aut-name=ShimadaAkira en-aut-sei=Shimada en-aut-mei=Akira kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=5 ORCID= en-aut-name=SaitoMegumu K. en-aut-sei=Saito en-aut-mei=Megumu K. kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=6 ORCID= en-aut-name=AsakaIsao en-aut-sei=Asaka en-aut-mei=Isao kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=7 ORCID= en-aut-name=KamiokaHiroshi en-aut-sei=Kamioka en-aut-mei=Hiroshi kn-aut-name= kn-aut-sei= kn-aut-mei= aut-affil-num=8 ORCID= affil-num=1 en-affil=Department of Orthodontics, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Okayama University kn-affil= affil-num=2 en-affil=Department of Orthodontics, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Okayama University kn-affil= affil-num=3 en-affil=Department of Orthodontics, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Okayama University kn-affil= affil-num=4 en-affil=Department of Orthodontics, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Okayama University kn-affil= affil-num=5 en-affil=Department of Pediatric Hematology/Oncology Okayama University Hospital kn-affil= affil-num=6 en-affil=Department of Clinical Application, Center for iPS Cell Research and Application Kyoto University kn-affil= affil-num=7 en-affil=Department of Fundamental Cell Technology, Center for iPS Cell Research and Application Kyoto University kn-affil= affil-num=8 en-affil=Department of Orthodontics, Graduate School of Medicine, Dentistry and Pharmaceutical Sciences Okayama University kn-affil= en-keyword=iPS cell kn-keyword=iPS cell en-keyword=RPL5 kn-keyword=RPL5 en-keyword=cleft lip and palate kn-keyword=cleft lip and palate en-keyword=chondrocyte kn-keyword=chondrocyte en-keyword=Diamond-Blackfan Anemia kn-keyword=Diamond-Blackfan Anemia END