Requirement of TGFβ Signaling for Effect of Fluoride on Osteoblastic Differentiation
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Research focused on transforming growth factor β (TGFβ) signaling in osteoblast is gradually increasing, whereas literature is rare in terms of fluorosis. This work aimed to investigate how TGFβ signaling participated in regulation of the osteoblast by different doses of fluoride treatment. Bone marrow stem cells (BMSCs) were developed into osteoblastic cells and exposed to 1, 4, and 16 mg/L F− with and without 10 ng/mL of TGFβ. Cell viability and differentiation state of osteoblast under different settings were measured by means of cell counting kit and analysis of alkaline phosphatase (ALP) activity as well as formation of mineral nodules. Real-time PCR was utilized to test expression of ALP and Runt-related transcription factor 2 (Runx2) at gene level. The gene expression of TGFβ signaling effectors was also investigated, such as TGFβ receptors (TβRs), smad3, and mitogen-activated protein kinases (MAPK). Results demonstrated that fluoride treatment exhibited action on osteoblast viability and osteogenic differentiation and upregulated expression of TβR2, smad3, and MAPK in this process. Administration of TGFβ strengthened ALP activity but attenuated formation of mineral nodules. Co-treatment of TGFβ and low-dose fluoride increased ALP activity compared to same dose of single fluoride treatment, whereas it inhibited mineral nodule formation. Administration of TGFβ reversed the suppression of high-dose fluoride on osteogenic differentiation of BMSCs. Taken together, studies revealed that TβR2 acted as a target for fluoride and TGFβ treatment on BMSCs, and smad3 and MAPK were involved in the mechanism of fluoride regulating osteogenic differentiation. Together, our data indicated that TGFβ receptor-mediated signaling through smad3 and MAPK was required for modulation of fluoride on osteoblast viability and differentiation, and activating TβR2-smad3 signaling pathway reversed suppression of osteoblasts differentiation by high dose of fluoride treatment.
KeywordsTransforming growth factor beta Osteoblast Alkaline phosphatase Transforming growth factor beta receptor Mitogen-activated protein kinase Smad3
This study was funded by project (Study on role of osteocyte and PTH/TGF-beta signaling pathway in the mechanism of bone turnover occurred in skeletal fluorosis) supported by the National Natural Science Foundation of China (grant number 81673111). This study was supported by a grant from Technical Innovation Project of Health and Family Planning Commission in Jilin Province of China (2016J054). This study was supported by the Natural Science Foundation of Jilin Province of China (20180101151JC).
Compliance with Ethical Standards
All procedures performed in studies involving mice were in accordance with the ethical standards of the Animal Welfare Ethical Committee of Jilin University guidelines and regulations.
Conflict of Interest
The authors declare that they have no conflict of interest.
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