中国中医骨伤科杂志2026，Vol.34Issue(2)：12-21,10.DOI:10.20085/j.cnki.issn1005-0205.260202
益气散瘀解毒方有效成分抑制骨肉瘤的机制研究
Study on the Mechanism of the Effective Components of the Yi-Qi-San-Yu-Jie-Du Formula in Inhibiting Osteosarcoma
摘要
Abstract
Objective:To explore the mechanism by which the main active components of the Yi-Qi-San-Yu-Jie-Du formula inhibit the occurrence and development of osteosarcoma through regulating the REGγ/RNF149/BRAF/MEK/ERK signa-ling pathway.Methods:Mass spectrometry and immunohistochemical experiments were conducted on samples of adjacent normal tissues and osteosarcoma tissues from 9 osteosarcoma patients.Osteosarcoma cell lines 143B and MG63 were used as research subjects.The expression of REGγ protein was regulated by shRNA interference and transient transfection methods.Western Blot and RT-PCR were used to verify the transfection efficiency and detect the expression levels of RNF149,BRAF,MEK,and ERK.Co-immunoprecipitation experiments and in vitro degradation experiments were per-formed to verify the interaction between REGγ and RNF149 proteins.The main components of Astragalus,Coptis,and Polyphyllin in the compound were identified by high-performance liquid chromatography.After treating the cells with these components,CCK-8,cell plate cloning,Transwell,Western Blot,and RT-PCR were used to detect cell proliferation ability and the expression levels of the REGγ/RNF149/BRAF/MEK/ERK signaling pathway.REGγ overexpression was achieved by transient transfection,and CCK-8 and cell plate cloning experiments were used to detect the cytotoxic effects of berberine,polyphyllin Ⅰ,and the compound on the two osteosarcoma cell lines.Western Blot and RT-PCR were used to detect changes in gene expression levels.C16-PAF,an activator of the MEK/ERK pathway,was used to treat the cells alone or in combination with the compound,and CCK-8 was used to detect cell survival rates.Results:REGγ was highly expressed in osteosarcoma tissues.Knockdown of REGγ significantly inhibited the proliferation and invasion abilities of the two osteosarcoma cell lines.The mRNA level of RNF149 did not change significantly,but the protein level increased significantly.The protein levels of BRAF,p-MEK,and p-ERK decreased significantly.REGγ was involved in regulating the ubiquitin and ATP-independent protein degradation process of RNF149.Berberine and polyphyllin Ⅰ were the main components of the Yi-Qi-San-Yu-Jie-Du formula that exerted anti-cancer effects,and the compound more significantly inhibited the proliferation ability of the two osteosarcoma cell lines than the two effective monomer components,which was consistent with the changes in protein expression levels after REGγ knockdown.Overexpression of REGγ significantly inhibited the cytotoxic effects of berberine,polyphyllin Ⅰ,and the compound on the two osteosarcoma cell lines.Overex-pression of REGγ significantly inhibited the ability of the compound to upregulate RNF149 protein expression and down-regulate BRAF,p-MEK,and p-ERK protein expression levels.C16-PAF significantly inhibited the cytotoxic effects of berberine,polyphyllin Ⅰ,and the compound on the two osteosarcoma cell lines.Conclusion:Yi-Qi-San-Yu-Jie-Du formula participates in the regulation of osteosarcoma occurrence and development through the REGγ/RNF149/BRAF/MEK/ERK signaling pathway,and berberine and polyphyllin Ⅰ are its main active components.关键词
益气散瘀解毒方/黄连素/重楼皂苷Ⅰ/骨肉瘤/信号通路Key words
Yi-Qi-San-Yu-Jie-Du formula/berberine/polyphyllin Ⅰ/osteosarcoma/signaling pathway分类
医药卫生引用本文复制引用
孙蕴初,周琳,张磊,王树强,尹萌辰,于文龙,吕凯,陈定榜,黄凯鹏,李建明,吴佳俊,葛志超,孙浩..益气散瘀解毒方有效成分抑制骨肉瘤的机制研究[J].中国中医骨伤科杂志,2026,34(2):12-21,10.基金项目
上海市卫生健康委员会临床行业专项(20204Y0165,20224Y0165)国家自然科学基金项目(82205145)上海市"医苑新星"青年医学临床人才计划(2023-062)中华中医药学会青年人才托举工程项目(2023-QNRC2-A03)上海中医药大学"访名校拜名师"项目(079) （20204Y0165,20224Y0165）
