Inhibition of Sphingosine-1-Phosphate Receptor 2 by JTE013 Promoted Osteogenesis by Increasing Vesicle Trafficking, Wnt/Ca<sup>2+</sup>, and BMP/Smad Signaling
Sphingosine-1-phosphate receptor 2 (S1PR2) is a G protein-coupled receptor that regulates various immune responses. Herein, we determine the effects of a S1PR2 antagonist (JTE013) or a S1PR2 shRNA on osteogenesis by culturing murine bone marrow stromal cells (BMSCs) in osteogenic media with JTE013,...
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| Format: | Article |
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MDPI AG
2021-11-01
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| Series: | International Journal of Molecular Sciences |
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| Online Access: | https://www.mdpi.com/1422-0067/22/21/12060 |
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| author | Simon Lin Subramanya Pandruvada Hong Yu |
| author_facet | Simon Lin Subramanya Pandruvada Hong Yu |
| author_sort | Simon Lin |
| collection | DOAJ |
| description | Sphingosine-1-phosphate receptor 2 (S1PR2) is a G protein-coupled receptor that regulates various immune responses. Herein, we determine the effects of a S1PR2 antagonist (JTE013) or a S1PR2 shRNA on osteogenesis by culturing murine bone marrow stromal cells (BMSCs) in osteogenic media with JTE013, dimethylsulfoxide (DMSO), a S1PR2 shRNA, or a control shRNA. Treatment with JTE013 or the S1PR2 shRNA increased alkaline phosphatase and alizarin red s staining, and enhanced alkaline phosphatase, RUNX2, osteocalcin, and osterix mRNA levels in BMSCs compared with the controls. Protein analysis revealed that a high dose of JTE013 (4 or 8 μM) increased vesicle trafficking-associated proteins (F-actin, clathrin, Early Endosome Antigen 1 (EEA1), and syntaxin 6) and Wnt/Ca2+ signaling. On the other hand, a low dose of JTE013 (1 to 2 μM) increased BMP/Smad signaling. In contrast, the S1PR2 shRNA reduced vesicle trafficking-associated proteins and attenuated Wnts and BMP/Smad signaling, but enhanced p-CaMKII compared with the control, suggesting that the S1PR2 shRNA influenced osteogenesis via different signaling pathways. Moreover, inhibiting protein trafficking by brefeldin A in BMSCs suppressed Wnts and BMPRs expressions. These data supported that enhanced osteogenesis in JTE013-treated BMSCs is associated with increased vesicle trafficking, which promotes the synthesis and transport of osteogenic protein and matrix vesicles and enhances matrix mineralization. |
| first_indexed | 2024-03-10T05:59:30Z |
| format | Article |
| id | doaj.art-28f43dc6f0ea46f0a3b480db16276085 |
| institution | Directory Open Access Journal |
| issn | 1661-6596 1422-0067 |
| language | English |
| last_indexed | 2024-03-10T05:59:30Z |
| publishDate | 2021-11-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | International Journal of Molecular Sciences |
| spelling | doaj.art-28f43dc6f0ea46f0a3b480db162760852023-11-22T21:02:13ZengMDPI AGInternational Journal of Molecular Sciences1661-65961422-00672021-11-0122211206010.3390/ijms222112060Inhibition of Sphingosine-1-Phosphate Receptor 2 by JTE013 Promoted Osteogenesis by Increasing Vesicle Trafficking, Wnt/Ca<sup>2+</sup>, and BMP/Smad SignalingSimon Lin0Subramanya Pandruvada1Hong Yu2Department of Oral Health Sciences, College of Dental Medicine, Medical University of South Carolina, Charleston, SC 29425, USADepartment of Oral Health Sciences, College of Dental Medicine, Medical University of South Carolina, Charleston, SC 29425, USADepartment of Oral Health Sciences, College of Dental Medicine, Medical University of South Carolina, Charleston, SC 29425, USASphingosine-1-phosphate receptor 2 (S1PR2) is a G protein-coupled receptor that regulates various immune responses. Herein, we determine the effects of a S1PR2 antagonist (JTE013) or a S1PR2 shRNA on osteogenesis by culturing murine bone marrow stromal cells (BMSCs) in osteogenic media with JTE013, dimethylsulfoxide (DMSO), a S1PR2 shRNA, or a control shRNA. Treatment with JTE013 or the S1PR2 shRNA increased alkaline phosphatase and alizarin red s staining, and enhanced alkaline phosphatase, RUNX2, osteocalcin, and osterix mRNA levels in BMSCs compared with the controls. Protein analysis revealed that a high dose of JTE013 (4 or 8 μM) increased vesicle trafficking-associated proteins (F-actin, clathrin, Early Endosome Antigen 1 (EEA1), and syntaxin 6) and Wnt/Ca2+ signaling. On the other hand, a low dose of JTE013 (1 to 2 μM) increased BMP/Smad signaling. In contrast, the S1PR2 shRNA reduced vesicle trafficking-associated proteins and attenuated Wnts and BMP/Smad signaling, but enhanced p-CaMKII compared with the control, suggesting that the S1PR2 shRNA influenced osteogenesis via different signaling pathways. Moreover, inhibiting protein trafficking by brefeldin A in BMSCs suppressed Wnts and BMPRs expressions. These data supported that enhanced osteogenesis in JTE013-treated BMSCs is associated with increased vesicle trafficking, which promotes the synthesis and transport of osteogenic protein and matrix vesicles and enhances matrix mineralization.https://www.mdpi.com/1422-0067/22/21/12060S1PR2JTE013osteogenesismatrix vesiclevesicle traffickingWnt |
| spellingShingle | Simon Lin Subramanya Pandruvada Hong Yu Inhibition of Sphingosine-1-Phosphate Receptor 2 by JTE013 Promoted Osteogenesis by Increasing Vesicle Trafficking, Wnt/Ca<sup>2+</sup>, and BMP/Smad Signaling International Journal of Molecular Sciences S1PR2 JTE013 osteogenesis matrix vesicle vesicle trafficking Wnt |
| title | Inhibition of Sphingosine-1-Phosphate Receptor 2 by JTE013 Promoted Osteogenesis by Increasing Vesicle Trafficking, Wnt/Ca<sup>2+</sup>, and BMP/Smad Signaling |
| title_full | Inhibition of Sphingosine-1-Phosphate Receptor 2 by JTE013 Promoted Osteogenesis by Increasing Vesicle Trafficking, Wnt/Ca<sup>2+</sup>, and BMP/Smad Signaling |
| title_fullStr | Inhibition of Sphingosine-1-Phosphate Receptor 2 by JTE013 Promoted Osteogenesis by Increasing Vesicle Trafficking, Wnt/Ca<sup>2+</sup>, and BMP/Smad Signaling |
| title_full_unstemmed | Inhibition of Sphingosine-1-Phosphate Receptor 2 by JTE013 Promoted Osteogenesis by Increasing Vesicle Trafficking, Wnt/Ca<sup>2+</sup>, and BMP/Smad Signaling |
| title_short | Inhibition of Sphingosine-1-Phosphate Receptor 2 by JTE013 Promoted Osteogenesis by Increasing Vesicle Trafficking, Wnt/Ca<sup>2+</sup>, and BMP/Smad Signaling |
| title_sort | inhibition of sphingosine 1 phosphate receptor 2 by jte013 promoted osteogenesis by increasing vesicle trafficking wnt ca sup 2 sup and bmp smad signaling |
| topic | S1PR2 JTE013 osteogenesis matrix vesicle vesicle trafficking Wnt |
| url | https://www.mdpi.com/1422-0067/22/21/12060 |
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