Factors secreted from the stem cells of human exfoliated deciduous teeth inhibit osteoclastogenesis through the activation of the endogenous antioxidant system

IF 2.6 Q1 DENTISTRY, ORAL SURGERY & MEDICINE

Journal of Oral Biosciences Pub Date : 2025-01-22 DOI:10.1016/j.job.2025.100618

Cheng Ding , Noboru Hashimoto , Fumiya Kano , Hirofumi Tenshin , Takahiro Arai , Linze Xia , Yang Xu , Houjun Lao , Yifei Wang , Tomonori Iwasaki , Hideharu Hibi , Akihito Yamamoto

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Abstract

Objectives

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.

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