富含虫草素的冬虫夏草乙醇提取物对骨肉瘤的抑制作用研究  

Antiproliferative Effects of Cordycepin-rich Ethanol Extract from Cordyceps Militaries on Osteosarcoma

作　　者：漆伟 孙强 张大伟 陈永锋 Qi Wei;Sun Qiang;Zhang Dawei;Chen Yongfeng(Department of Orthopedics,Xijing Hospital,Air Force Military Medical University,Xi'an 710032,China)

机构地区：[1]空军军医大学附属西京医院骨科,西安710032

出　　处：《骨科》2024年第6期549-554,共6页ORTHOPAEDICS

基　　金：陕西省重点研发计划项目(2024SF-YBXM-364)。

摘　　要：目的评价富含虫草素的冬虫夏草乙醇提取物(CRECM)对骨肉瘤的抑制作用。方法制备CRECM,并采用高效液相色谱(HPLC)评估CRECM中虫草素的含量。体外实验,以不同浓度(1、2、4μg/mL)的CRECM诱导LM8细胞,采用细胞计数试剂盒(cell counting kit,CCK)-8检测细胞活力;采用流式细胞术分析细胞凋亡情况;通过划痕实验测定细胞迁移能力;采用实时荧光定量聚合酶链反应(RTqPCR)、Western blot法测定基质金属蛋白酶(MMP)-2的表达。体内实验采用昆明小鼠建立骨肉瘤LM8的肿瘤模型,观察不同浓度(1、2、4 mg/kg)CRECM刺激的小鼠在不同时间点的肿瘤体积、体重变化和总体生存情况。结果CRECM的虫草素含量为33350.676μg/mL。不同浓度的CRECM均可显著降低LM8细胞活力,且呈剂量依赖性;CRECM引起LM8细胞的凋亡率显著增加,并且通过划痕实验确定CRECM显著降低了细胞迁移能力;经CRECM诱导后,MMP-2 mRNA水平显著降低。在小鼠模型中,CRECM在体内抑制骨肉瘤细胞生长,且未出现明显的药物毒性。结论CRECM剂量依赖性地抑制骨肉瘤的增殖,其抗癌作用通过诱导细胞凋亡和抑制细胞迁移来实现;CRECM在体内抑制肿瘤生长,且无明显的药物毒性。Objective To evaluate the antiproliferative effect of cordycepin-rich ethanol extract from cordyceps militaris(CRECM)on osteosarcoma.Methods CRECM was prepared and the cordycepin content in CRECM was assessed using high-performance liquid chromatography(HPLC).The in vitro experiments were performed to treat LM8 cells with varying concentrations(1,2,and 4μg/mL)of CRECM,and cell viability was evaluated utilizing a cell counting kit-8(CCK-8)assay.Apoptosis was examined by flow cytometry.The migratory capacity of the cells was assessed via a wound healing assay.The expression levels of matrix metalloproteinase-2(MMP-2)were quantified using real-time fluorescence quantitative polymerase chain reaction(RT-qPCR)and Western blotthg.The in vivo studies were conducted to establish an LM8 osteosarcoma tumor model in Kunming mice,and the changes in tumor volume,body weight,and overall survival of mice stimulated with CRECM at different concentrations(1,2,and 4 mg/kg)at different time points were observed.Results The cordycepin concentration in CRECM was 33350.676μg/mL.Treatment with CRECM at varying concentrations significantly diminished the viability of LM8 cells in a dose-dependent manner and markedly increased the rate of apoptosis among these cells.Additionally,CRECM substantially impaired cell migration capabilities as demonstrated by scratch assays.Following induction with CRECM,there was a significant reduction in MMP-2 mRNA levels.In the in vivo mouse model,CRECM effectively inhibited the growth of osteosarcoma cells without exhibiting notable drug toxicity.Conclusion CRECM dose-dependently inhibits the proliferation of osteosarcoma,and its anti-cancer effect is achieved by inducing cell apoptosis and inhibiting cell migration.CRECM inhibits tumor growth in vivo without significant drug toxicity.

关 键 词：冬虫夏草 富含虫草素的冬虫夏草乙醇提取物 骨肉瘤 虫草素 基质金属蛋白酶2 

分 类 号：R285.5[医药卫生—中药学]