β-榄香烯诱导人骨肉瘤143B细胞增殖、凋亡及自噬的实验研究  

The investigation ofβ-elemene on proliferation,apoptosis and autophagy of human osteosarcoma 143B cells

作　　者：赵茗[1] 周庄[1] 齐典文[1] 孙涛[1] 郭昶志 赵桂松 马天晓 刘思源[1] 胡彤宇[1] 张国川[1] ZHAO Ming;ZHOU Zhuang;QI Dianwen;SUN Tao;GUO Changzhi;ZHAO Guisong;MA Tianxiao;LIU Siyuan;HU Tongyu;ZHANG Guochuan(Department of Bone and Soft Tissue Oncology, the Third Hospital of Hebei Medical University, Shijiazhuang 050051, China;Department of Orthopedics, Chest Hospital of Hebei Province, Shijiazhuang 050051, China)

机构地区：[1]河北医科大学第三医院骨与软组织肿瘤科,河北石家庄050051 [2]河北省胸科医院骨科,河北石家庄050051

出　　处：《中国肿瘤外科杂志》2021年第3期267-271,共5页Chinese Journal of Surgical Oncology

基　　金：河北省引进留学人员资助项目(CY201715)。

摘　　要：目的探讨β-榄香烯在骨肉瘤中诱导自噬发生的信号通路及其对肿瘤细胞增殖及凋亡的影响。方法以不同浓度(0、10、20、50、100、150μg/ml)的β-榄香烯处理人骨肉瘤143B细胞24、48 h后,应用CCK-8法检测细胞存活率;用0、20、50μg/mlβ-榄香烯处理143B细胞48 h后,应用流式细胞仪使用AnnexinⅤ-FITC/PI双染法检测细胞凋亡率,并通过荧光显微镜及蛋白印迹法(Western blotting)检测细胞自噬相关蛋白微管相关蛋白1轻链3(LC3B)的分布及表达,Western blotting检测143B细胞中丝氨酸/苏氨酸激酶(AKT)、磷酸化丝氨酸/苏氨酸激酶(p-AKT)、哺乳动物雷帕霉素靶蛋白(mTOR)及磷酸化哺乳动物雷帕霉素靶蛋白(p-mTOR)蛋白表达水平。结果β-榄香烯呈浓度依赖性地显著抑制骨肉瘤143B细胞的增殖,其对143B细胞24 h和48 h时的半数抑制浓度(IC50)分别为58.89μg/ml和18.48μg/ml;143B细胞经20、50μg/mlβ-榄香烯处理48 h后,细胞凋亡率分别为(15.43±0.99)%和(19.21±0.82)%,明显高于0μg/mlβ-榄香烯组(6.87±1.02)%;且其细胞Cleaved caspase-3蛋白表达水平亦明显高于0μg/mlβ-榄香烯组,差异均有统计学意义(P<0.05);经20、50μg/mlβ-榄香烯处理骨肉瘤143B细胞48 h后,细胞质内的自噬体数量明显增多,其LC3-Ⅱ/LC3-Ⅰ表达水平较0μg/mlβ-榄香烯组均明显增高,差异均有统计学意义(P<0.05),β-榄香烯可以抑制骨肉瘤细胞p-AKT和p-mTOR的表达,且呈时间及剂量依赖性。结论β-榄香烯可以诱导人骨肉瘤143B细胞增殖抑制及凋亡,还可以通过抑制AKT/mTOR通路激活细胞自噬。Objective To investigate the effects ofβ-elemene on proliferation,apoptosis and autophagy of osteosarcoma cells and its potential mechanisms.Methods Human osteosarcoma cell line 143B was treated with different concentrations ofβ-elemene(0,10,20,50,100 and 150μg/ml).Cell proliferation was evaluated by CCK-8 method.After the cells were treated with 0,20 and 50μg/mlβ-elemene for 48 h,cell apoptotic rate was estimated by AnnexinⅤ-FITC/PI double staining with flow cytometry.Distribution of autophagy associated protein LC3B was observed by fluorescence microscope.The protein levels of LC3B,AKT,p-AKT,mTOR and p-mTOR were examined by Western blotting.Resultsβ-elemene significantly inhibited cell proliferation in a dose-dependent manner.IC50 ofβ-elemene on 143B cells for 24 h and 48 h were 58.89μg/ml and 18.48μg/ml.After treatment with 20 or 50μg/mlβ-elemene for 48 h,the apoptosis rate of 143B cells was(15.43±0.99)%and(19.21±0.82)%,respectively,which was significantly higher than that of the 0μg/mlβ-elemene group(6.87±1.02)%(P<0.05).The expression level of Cleaved caspase-3 was significantly higher than that of the 0μg/mlβ-elemene group(P<0.05).The numbers of autophagosome in the cytoplasm increased,and the expression level of LC3-Ⅱ/LC3-Ⅰwas significantly higher than that of the 0μg/mlβ-elemene group(P<0.05).β-elemene could inhibit the activation of AKT and mTOR in osteosarcoma cells in a time and dose dependent manner.β-elemene induced autophagy in 143B cells by inhibiting the activation of AKT/mTOR pathway in a time-and dose-dependent manner.Conclusionsβ-elemene could inhibit cell proliferation,induce apoptosis and autophagy of 143B cells by inhibiting AKT/mTOR pathway.

关 键 词：Β-榄香烯 增殖 凋亡 自噬 骨肉瘤细胞 

分 类 号：R73[医药卫生—肿瘤]