Oleanolic acid inhibits RANKL-induced osteoclastogenesis via ER alpha/miR-503/RANK signaling pathway in RAW264.7 cells
Oleanolic acid inhibits RANKL-induced osteoclastogenesis via ER alpha/miR-503/RANK signaling pathway in RAW264.7 cells
复制标题
齐墩果酸通过 ER α/miR-503/RANK 信号通路抑制 RAW264.7 细胞中 RANKL 诱导的破骨细胞生成
DOI：
10.1016/j.biopha.2019.109045
复制
发表时间：
2019-09-01
影响因子：
7.5
通讯作者：
Gan, Guo-xing
中科院分区：
文献类型：
--
作者：
Xie, Bao-ping;Shi, Li-ying;Gan, Guo-xing
齐墩果酸(OA)近年来成为治疗多种人类疾病的研究热点，尤其是骨质疏松症和关节炎。然而，其作用机制尚未完全阐明。我们的目的是阐明OA抑制破骨细胞分化的靶点和机制。采用TRAP染色和甲苯胺蓝染色分别检测OA对破骨细胞形成和骨吸收的影响。检测破骨细胞分化相关基因雌激素受体α(ERα)和miR-503的表达水平。我们用ERα的特异性阻断剂甲基哌啶基吡唑(MPP)来阻断ERα。我们用Anagomir-503-5P拮抗miR-503的作用。利用RT-PCR和ELISA试剂盒从基因和蛋白水平研究了OA对miR-503的形成以及成熟相关酶DICER和DROSHA的影响。结果提示，骨钙素可抑制破骨细胞生成和骨吸收。OA上调ERα和miR-503的表达水平，抑制RANK的表达。MPP可显著减弱骨性关节炎的作用，包括抑制破骨细胞生成、抑制骨吸收和上调miR-503的表达。结果表明，ERα是骨关节炎的靶点，骨关节炎通过ERα上调miR-503的表达。Antag omir-503-5p可抑制miR-503的功能，并减弱OA对破骨细胞生成的抑制作用，提示OA通过上调miR-503的表达抑制破骨细胞的形成。此外，OA通过上调DICER的表达上调miR-503的表达。综上所述，OA通过ERα/miR-503/RANK信号通路抑制RANKL诱导的RAW264.7细胞破骨细胞生成。
Oleanolic acid (OA) has recently become a research hotspot in the treatment of many human diseases, especially osteoporosis and arthritis. However, the mechanisms are not elucidated completely. We aimed to elucidate the target and the mechanism via which OA inhibited osteoclast differentiation. We used TRAP staining and toluidine blue dye to test OA effect on osteoclastogenesis and bone resorption respectively. We detected the expression level of osteoclast differentiation related genes, estrogen receptor alpha (ER alpha) and miR-503. We blocked ER alpha with its specific blocker, methylpiperidino pyrazole (MPP). We antagonized the function of miR-503 with antagomir-503-5p. RT-PCR and ELISA kits were used to investigate the effects of OA on miR-503 formation and maturation-relevant enzymes Dicer and Drosha at gene and protein levels. The data suggested that OA inhibited osteoclastogenesis and bone resorption. OA upregulated ER alpha and miR-503 expression levels, inhibited RANK expression. MPP significantly attenuated the OA effect including inhibiting osteoclastogenesis, inhibiting bone resorption and up-regulating miR-503 expression. It showed that ER alpha was the target of OA and OA up-regulated miR-503 expression through ER alpha. Antagomir-503-5p inhibited the function of miR-503 and attenuated the inhibition of OA on osteoclastogenesis, suggesting that OA inhibited osteoclast by up-regulating miR-503 expression. In addition, OA up-regulated miR-503 by up-regulating Dicer expression. In conclusion, OA inhibits RANKL-induced osteoclastogenesis via ER alpha/miR-503/ RANK signaling pathway in RAW264.7 cells.