芍药苷促进 MC3T3-E1 成骨细胞分化的作用及机制。

BACKGROUND: The incidence of osteoporosis and osteoporotic fractures is increasing every year. Traditional Chinese Medicine (TCM) can shed new light on the treatment of osteoporosis. This study aimed to explore the role and mechanism of paeoniflorin in promoting osteogenic differentiation of an osteoblast precursor cell line (MC3T3-E1). METHODS: MC3T3-E1 cells were cultured in osteogenic induction medium (OIM) and OIM combined with different concentrations of paeoniflorin. The optimal dose of paeoniflorin was assessed by a cell counting kit-8 (CCK-8) assay. Then, alkaline phosphatase (ALP) and Alizarin Red S (ARS) staining were performed to assess the osteogenic capacity of paeoniflorin. The transcription of osteogenic genes and the expression of osteogenic proteins were assessed by RT-PCR and Western blotting, respectively. The transcription of Wnt/β-catenin signaling pathway genes and proteins was assessed by RT-PCR and Western blotting, respectively. Finally, Dickkopf-1 (DKK-1), a Wnt/β-catenin signaling pathway inhibitor, was used to identify whether the Wnt/β-catenin signaling pathway was involved in the osteogenic differentiation of paeoniflorin. Osteoclastogenesis in RAW264.7 cells was identified by tartrate-resistant acid phosphatase (TRAP) staining. RESULTS: At concentrations ranging from 0.1 to 100 μM, paeoniflorin was not cytotoxic to MC3T3-E1 cells. Paeoniflorin significantly increased the osteogenic differentiation of MC3T3-E1 cells in a dose-dependent manner. Moreover, paeoniflorin significantly increased osteogenic differentiation gene and protein expression. Through bioinformatic analysis, paeoniflorin-affected genes were found to be involved in different signaling pathways, such as the Wnt/β-catenin signaling pathway. Paeoniflorin enhanced β-catenin and CyclinD1 expression compared with that of the control groups. DKK-1 partially reversed the promoting effects of paeoniflorin in promoting osteogenic differentiation of MC3T3-E1 cells. Moreover, paeoniflorin inhibited the osteoclastogenesis of RAW264.7 cells. CONCLUSION: Paeoniflorin promotes osteogenic differentiation in MC3T3-E1 cells by regulating the Wnt/β-catenin pathway. Paeoniflorin is a potential therapeutic agent for the treatment of osteoporosis.

背景：骨质疏松症和骨质疏松性骨折的发病率逐年上升。中医药可以为骨质疏松症的治疗带来新的思路。本研究旨在探讨芍药苷促进成骨前体细胞系（MC3T3-E1）成骨分化的作用和机制。

方法：将 MC3T3-E1 细胞在成骨诱导培养基（OIM）和 OIM 中加入不同浓度芍药苷中培养。用细胞计数试剂盒-8（CCK-8）法评估芍药苷的最佳剂量。然后，通过碱性磷酸酶（ALP）和茜素红 S（ARS）染色评估芍药苷的成骨能力。通过 RT-PCR 和 Western blot 分别评估成骨基因和蛋白的转录。通过 RT-PCR 和 Western blot 分别评估 Wnt/β-catenin 信号通路基因和蛋白的转录。最后，用 Wnt/β-catenin 信号通路抑制剂 Dickkopf-1（DKK-1）鉴定 Wnt/β-catenin 信号通路是否参与芍药苷的成骨分化。通过抗酒石酸酸性磷酸酶（TRAP）染色鉴定 RAW264.7 细胞中的破骨细胞生成。

结果：在 0.1 至 100 μM 浓度范围内，芍药苷对 MC3T3-E1 细胞无细胞毒性。芍药苷呈剂量依赖性显著增加 MC3T3-E1 细胞的成骨分化。此外，芍药苷显著增加成骨分化基因和蛋白的表达。通过生物信息学分析，发现芍药苷影响的基因参与不同的信号通路，如 Wnt/β-catenin 信号通路。与对照组相比，芍药苷增强了 β-catenin 和 CyclinD1 的表达。DKK-1 部分逆转了芍药苷促进 MC3T3-E1 细胞成骨分化的作用。此外，芍药苷抑制 RAW264.7 细胞的破骨细胞生成。

结论：芍药苷通过调节 Wnt/β-catenin 通路促进 MC3T3-E1 细胞的成骨分化。芍药苷是治疗骨质疏松症的潜在治疗药物。