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SOX2 suppresses osteoblast differentiation of MC3T3-E1 cells through activating the transcription of LGR4

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Abstract

Osteogenic differentiation is a crucial process of new bone formation. This study aimed to explore the roles and mechanism of SRY-Box Transcription Factor 2 (SOX2) on proliferation and osteogenic differentiation of MC3T3-E1 cells. Bone morphogenetic protein 2 (BMP2) was used to induce the osteogenic differentiation of MC3T3-E1 cells. The expression of SOX2 was determined by quantitative real-time PCR (RT-PCR) at different time points after induction. The SOX2 overexpression plasmids were constructed and transfected into MC3T3-E1 cells. Osteogenic differentiation was evaluated by Alizarin Red S staining and alkaline phosphatase (ALP) assay. The expressions of osteogenic differentiation markers including runt-related transcription factor 2 (Runx2), osteopontin (OPN), and osteocalcin (OCN) were detected by western blot assay. Luciferase reporter and CHIP assays were used to confirm that SOX2 regulated the transcriptional activation of leucine-rich repeat-containing G protein–coupled receptor 4 (LGR4). We found that SOX2 was down-regulated upon BMP2-induced osteogenic differentiation in MC3T3-E1 cells. Overexpression of SOX2 effectively inhibited osteogenic differentiation with decreased ALP activity, calcification, and osteogenic differentiation markers’ expression including Runx2, OPN, and OCN. LGR4 was identified as a target of SOX2, and the inhibitory effect of SOX2 on osteogenic differentiation was reversed by knockdown of LGR4. The present study confirmed that SOX2 suppressed osteogenic differentiation of MC3T3-E1 cells through targeting LGR4, which possesses a therapeutic strategy for bone formation and generation.

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Data availability

The data used and/or analyzed during the current study are available from the corresponding author on reasonable request.

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Acknowledgements

We are grateful to all the colleagues in our laboratory for their constructive help.

Funding

This work was financially supported by General Projects of Fujian Provincial Natural Fund (2021J011312).

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Authors and Affiliations

  1. The Third Clinical Medical College, Fujian Medical University, No. 88, Jiaotong Road, Fuzhou, Fujian Province, 350004, People’s Republic of China

    Sunyu Chen

  2. Department of Orthopedics, Fuzhou Second Hospital, Cangshan District, No. 47, Shangteng Road, Fuzhou, Fujian Province, 350007, People’s Republic of China

    Sunyu Chen, Zhanhao Xiao & Wenjin Jiang

  3. Fujian Provincial Clinical Medical Research Center for First Aid and Rehabilitation in Orthopaedic Trauma (2020Y2014), Fuzhou, People’s Republic of China

    Sunyu Chen

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  1. Sunyu Chen
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  2. Zhanhao Xiao
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All authors participated in making the design, performing the experiment, analyzing the data, and drafting the manuscript, and all authors consented to its submission for publication.

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Correspondence to Sunyu Chen.

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Chen, S., Xiao, Z. & Jiang, W. SOX2 suppresses osteoblast differentiation of MC3T3-E1 cells through activating the transcription of LGR4. In Vitro Cell.Dev.Biol.-Animal 59, 1–9 (2023). https://doi.org/10.1007/s11626-022-00740-4

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  • DOI: https://doi.org/10.1007/s11626-022-00740-4

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