Anethole promotes osteogenic differentiation of human bone marrow mesenchymal stem cells

doi:10.12307/2026.847

Abstract

Abstract: BACKGROUND: While anethole has been shown to inhibit receptor activator of nuclear factor kappa-B ligand-induced osteoclastogenesis and ameliorate osteoporosis, its potential to promote bone formation remains unclear.
OBJECTIVE: To explore the effect of anethole on promoting osteogenic differentiation of human bone marrow mesenchymal stem cell and its mechanism.
METHODS: CCK-8 assay was employed to assess cytotoxicity of anethole and identify optimal concentration for further study. Human bone marrow mesenchymal stem cells were then stimulated with either 100 μM anethole or its vehicle, dimethyl sulfoxide added in osteogenic differentiation medium, and their effects on osteogenic differentiation of human bone marrow mesenchymal stem cells were observed. Human bone marrow mesenchymal stem cells were stimulated with 1 μmol/L bone morphogenetic protein 2-transforming growth factor β-Smad2/3 signaling pathway inhibitor LDN193189 or phosphate-buffered saline (control) in osteogenic differentiation medium containing 100 μmol/L anethole, and the effect of the signaling pathway inhibitor on osteogenic differentiation of human bone marrow mesenchymal stem cells was observed. Alizarin red staining was used to observe calcium nodule formation in human bone marrow mesenchymal stem cells. RT-qPCR and western blot assay were used to detect the expression of related osteogenic differentiation markers, including type I collagen α1 chain, Runt-related transcription factor 2, and genes and proteins related to the bone morphogenetic protein 2, transforming growth factor β, and Smad2/3.
RESULTS AND CONCLUSION: (1) CCK-8 assay showed that 200 μmol/L anethole had a cytotoxic effect on human bone marrow mesenchymal stem cells, while 50 and 100 μmol/L anethole had no cytotoxic effect. Therefore, 100 μmol/L anethole was selected for subsequent exploratory experiments. (2) 100 μmol/L anethole promoted calcium nodule formation and increased the expression of osteogenic differentiation markers, including type I collagen α1 chain and Runt-related transcription factor 2, in human bone marrow mesenchymal stem cells, as well as promoted the increased expression of genes related to the bone morphogenetic protein 2-transforming growth factor β-Smad2/3 signaling pathway. (3) After blocking the signaling pathway with the bone morphogenetic protein 2-transforming growth factor β-Smad2/3 signaling pathway inhibitor LDN193189, the effect of anethole on promoting calcium nodule formation and increasing the expression of osteogenic differentiation markers (type I collagen α1 chain and Runt-related transcription factor 2) in human bone marrow mesenchymal stem cells was inhibited. The results indicate that anethole can activate the bone morphogenetic protein 2-transforming growth factor β-Smad2/3 signaling pathway to promote the osteogenic differentiation of human bone marrow mesenchymal stem cells.

Key words: osteoporosis, bone marrow mesenchymal stem cell, anethole, osteogenic differentiation, bone morphogenetic protein 2-transforming growth factor β-Smad2/3 signaling pathway, network pharmacology analysis, osteogenic differentiation markers

CLC Number:

Lu Chuan, Yang Jiale, Li Youliang, Li Chao. Anethole promotes osteogenic differentiation of human bone marrow mesenchymal stem cells[J]. Chinese Journal of Tissue Engineering Research, 2026, 30(31): 8041-8046.

Figures/Tables 5

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