Department of Spine Surgery and Orthopaedics, Xiangya Hospital, Central South University, Changsha, Hunan 410008, China; National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University,Changsha, Hunan 410008, China.
Hunan Provincial Key Laboratory of Regional Hereditary Birth Defects Prevention and Control, Changsha Hospital for Maternal & Child Health Care Affiliated to Hunan Normal University, Changsha, China.
Phytomedicine. 2024 Feb;124:155284. doi: 10.1016/j.phymed.2023.155284. Epub 2023 Dec 17.
Osteoporosis is a systemic skeletal disorder characterized by decreased bone density and the degradation of bone tissue microarchitecture. Ginsenoside Rg1, derived from Panax ginseng, has been a part of traditional Chinese medicine in China for centuries, particularly for treating osteoporosis. However, there remains limited research on the osteogenic potential of Rg1 within the glucocorticoid-induced osteoporosis (GIOP) model and its specific mechanisms.
The primary objective of this study is to investigate the osteogenic potential of Rg1 within the GIOP model and explore the signaling pathways associated with its in vivo and in vitro effects.
Cell proliferation, differentiation and mineralization were evaluated by the Cell counting kit 8(CCK8) assay, alkaline phosphatase (ALP) test and Alizarin Red S staining, respectively. The qPCR technique was used to determine the relative expression of mRNA and the western blot was used to determine the relative expression of protein. In vivo experiments, spinal vertebrae staining in zebrafish larvae was accomplished by alizarin red S staining.
Zebrafish larvae's hatching, survival, malformation, and heart rate were unaffected by 50 μM of Rg1 in vivo, while the MEC3T3-E1 cell line's proliferation was unaffected by 50 μM of Rg1 in vitro. Meanwhile, Rg1 was shown to improve osteogenic differentiation or bone formation as well as the level of mRNA expression of osteogenic markers in vivo and in vitro. Treatment with Rg1 significantly increased the expression of G protein-coupled estrogen receptor (GPER) and pAKT. In addition, the GPER inhibitor G15 could significantly reduce the mRNA and protein expression levels of GPER and phosphorylated AKT, LY294002, a PI3K/AKT pathway inhibitor, markedly suppresses the expression of phosphorylated AKT, yet shows no significant impact on GPER expression. Both G15 and LY294002 can significantly blocked the Rg1-mediated enhancement of osteogenesis capacity in the GIOP model. In contrast, when both the agonists G1 of GPER and LY294002 were added, G1 increased the relative expression of mRNA and protein of GPER, but not the expression of osteogenic capacity and osteogenic markers.
This study investigates the mineralization effects and mechanisms of Ginsenoside Rg1 both in vitro and in vivo. For the first time, we propose that Rg1 might regulate osteogenesis by modulating AKT phosphorylation through mediating GPER expression within the PI3K/AKT pathway in the GIOP model. This discovery introduces novel targets and avenues for osteoporosis treatment.
骨质疏松症是一种以骨密度降低和骨组织微结构降解为特征的系统性骨骼疾病。人参皂苷 Rg1 来源于人参，在中国传统医学中已使用了数个世纪，特别是用于治疗骨质疏松症。然而，在糖皮质激素诱导的骨质疏松症（GIOP）模型中，Rg1 的成骨潜力及其具体机制的研究仍然有限。
本研究的主要目的是研究 Rg1 在 GIOP 模型中的成骨潜力，并探讨与其体内和体外作用相关的信号通路。
通过细胞计数试剂盒 8（CCK8）测定法、碱性磷酸酶（ALP）试验和茜素红 S 染色分别评估细胞增殖、分化和矿化。采用 qPCR 技术测定 mRNA 的相对表达，采用 Western blot 技术测定蛋白的相对表达。体内实验通过在斑马鱼幼虫中进行茜素红 S 染色来完成脊椎骨染色。
体内实验中，50 μM 的 Rg1 对斑马鱼幼虫的孵化、存活、畸形和心率没有影响，而体外实验中，50 μM 的 Rg1 对 MEC3T3-E1 细胞系的增殖没有影响。同时，Rg1 被证明可改善体内和体外的成骨分化或骨形成以及成骨标志物的 mRNA 表达水平。Rg1 处理可显著增加 G 蛋白偶联雌激素受体（GPER）和 pAKT 的表达。此外，GPER 抑制剂 G15 可显著降低 GPER 和磷酸化 AKT 的 mRNA 和蛋白表达水平，PI3K/AKT 通路抑制剂 LY294002 显著抑制磷酸化 AKT 的表达，但对 GPER 表达没有显著影响。G15 和 LY294002 均可显著阻断 Rg1 在 GIOP 模型中增强成骨能力。相比之下，当同时添加 GPER 的激动剂 G1 和 LY294002 时，G1 增加了 GPER 的 mRNA 和蛋白的相对表达，但没有增加成骨能力和成骨标志物的表达。
本研究在体内和体外研究了人参皂苷 Rg1 的矿化作用及其机制。首次提出，Rg1 可能通过调节 PI3K/AKT 通路中的 GPER 表达来调节 AKT 磷酸化，从而调节 GIOP 模型中的成骨作用。这一发现为骨质疏松症的治疗提供了新的靶点和途径。
