Lithospermic Acid Promotes Osteoblast Proliferation and Differentiation through the Wnt/β-Catenin Signaling Pathway

Jianfeng Wang^#, Zhongqing Hu^#, Jiandong Guo, Xin Jin, Lei Cai, Jian Li, Jinxi Zhang^*, DONGAN HE^*
Department of Orthopaedics, Hangzhou Ninth Hospital, Hangzhou, 311225, China
* Corresponding Author: Jinxi Zhang. Email: email ; DONGAN HE. Email: email
# These authors contributed equally to this work

BIOCELL https://doi.org/10.32604/biocell.2025.072227

Received 22 August 2025; Accepted 30 October 2025; Published online 05 January 2026

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Abstract

Objectives: Therapeutic strategies for enhancing bone regeneration and combating osteoporosis remain a significant unmet medical need. This study aims to elucidate Lithospermic acid (LA)’s regulatory effects on osteoblast proliferation and differentiation, investigating its viability as a bone-healing agent. Methods: This study employed various cellular and molecular biology experiments to assess the effects of LA on the viability, proliferation, cell cycle, apoptosis, differentiation, mineralization, and migration of MC3T3-E1 osteoblasts. Immunofluorescence and Western blot analyses were conducted to detect the expression of proteins related to the Wnt/β-catenin signaling pathway, investigating the regulatory mechanisms by which LA promotes osteoblast proliferation and differentiation. Additionally, Wnt inhibitor dickkopf-1 (DKK-1) and β-catenin-silenced cell models were used to further validate the role of LA in modulating this signaling pathway. Results: LA significantly promoted osteoblast proliferation without apparent cytotoxicity. Flow cytometry showed that LA regulated the cell cycle by reducing G0/G1 phase arrest and promoting G2/M phase progression. Western blot results indicated that LA upregulated the expression of proteins associated with cell proliferation and enhanced osteoblast differentiation and mineralization. Immunofluorescence and Western blot analyses further confirmed that LA markedly increased the expression of Wnt and β-catenin, facilitating β-catenin nuclear translocation. Treatment with the DKK-1 inhibitor significantly diminished the proliferative and differentiation-promoting effects of LA, confirming the critical role of this pathway. β-catenin knockdown experiments further substantiated its central role in LA-mediated regulation. Conclusion: This study confirms that LA promotes osteoblast proliferation, differentiation, mineralization, and migration by activating the Wnt/β-catenin signaling pathway.