丁香提取物对放射抗拒食管癌细胞的体外抗肿瘤作用及机制研究  

In vitro anti-tumor effect and mechanism of clove extracts against radioresistant esophagus cancer cell

作　　者：龚盈盈 谢玲 朱圣明 吴峰 段奇文 邓鑫州 柯青 骆志国 GONG Yingying;XIE Ling;ZHU Shengming;WU Feng;DUAN Qiwen;DENG Xinzhou;KE Qing;LUO Zhiguo(Postgraduate Training Basement of Jinzhou Medicical University,Taihe Hospital,Hubei University of Medicine,Shiyan,Hubei 442000,P.R.China;Department of Dermatology,Taihe Hospital of Shiyan City,Affiliated Hospital of Hubei University of Medicine,Shiyan,Hubei 442000,P.R.China;Department of Oncology,Taihe Hospital of Shiyan City,Affiliated Hospital of Hubei University of Medicine/Hubei Provincial Center for Precision Medicine of Cancer,Shiyan,Hubei 442000,P.R.China)

机构地区：[1]锦州医科大学十堰市太和医院研究生培养基地(湖北医药学院附属医院),湖北十堰442000 [2]十堰市太和医院(湖北医药学院附属医院)皮肤科,湖北十堰442000 [3]十堰市太和医院(湖北医药学院附属医院)肿瘤科/湖北省肿瘤精准医学研究中心,湖北十堰442000

出　　处：《华西医学》2021年第1期66-75,共10页West China Medical Journal

基　　金：国家自然科学基金(81602391);湖北医药学院自由探索项目(FDFR201802)。

摘　　要：目的探讨丁香提取物(clove extract,CE)对放射抗拒食管癌细胞KYSER的体外抗肿瘤疗效及其机制。方法制备丁香花蕾乙醇提取物,采用气相色谱法鉴定CE中的主要活性组分;采用体外细胞培养法观察不同浓度CE对食管癌放射抗拒细胞KYSER生长的影响;采用噻唑蓝法检测不同浓度CE对KYSER存活的影响及方式;采用透射电子显微镜观察CE处理KYSER后细胞器微观结构的变化;采用Transwell小室法检测CE对KYSER迁徙能力的影响;采用流式细胞术定量测定CE诱导KYSER的凋亡率及细胞周期分布;采用克隆形成实验检测CE处理后,KYSER的克隆形成能力变化。结果气相色谱法显示:CE的主要成分为丁香酚、丁香油烃和乙酸丁香酚酯。体外细胞培养法观察到:0.4%CE即可对KYSER生长产生抑制作用。噻唑蓝法显示:CE浓度≥0.5%时即可对KYSER的存活起到抑制作用,抑制作用呈明显的浓度依赖性。透射电子显微镜观察到:0.5%CE处理KYSER后,肿瘤细胞微绒毛变短、变宽,细胞质中核糖体减少,线粒体萎缩,形成大量自噬小体。Transwell跨膜迁徙实验显示:0.5%和0.6%的CE处理细胞后,KYSER迁徙率分别为对照的(65±4)%和(41±3)%,分别与对照组比,差异均有统计学意义(P均<0.001)。流式细胞术凋亡检测显示:对照组、0.5%CE和0.6%CE处理组细胞的凋亡率分别为(5.63±0.50)%、(11.77±0.42)%和(19.44±0.19)%,处理组与对照组相比差异均有统计学意义(P均<0.001)。流式细胞术细胞周期检测显示:对照组、0.5%CE处理组和0.6%CE处理组细胞G1期比例分别为(61.99±1.20)%、(75.38±1.50)%和(78.81±1.00)%,处理组分别与对照组相比,差异均有统计学意义(P均<0.001)。克隆形成实验显示:CE处理细胞24 h后,对照组、0.5%CE处理组和0.6%CE处理组细胞克隆形成率分别为(80.5±1.0)%、(18.1±0.8)%和(5.0±0.5)%,处理组与对照组相比,差异有统计学意义(P均<0.001)。结论CE可通过诱导细胞自噬和促进凋�Objective To explore the in vitro anti-tumor effect of clove extracts(CEs)on radioresistant esophageal cancer cell KYSER and its mechanism.Methods Ethanol extracts of clove bud were prepared.Gas chromatography was used to identify the main active components of CEs.In vitro cell culture method was used to observe the effect of CEs at different concentrations on KYSER cell growth.Methyl thiazolyl tetrazolium(MTT)method was used to detect the effect of different concentration CEs on KYSER’s survival and its’manner.Transmission electron microscope(TEM)was used to observe the changes of KYSER’s organelle microstructure after CEs treated.Transwell chamber method was used to detect the impact of CEs on KYSER’s migration ability.The apoptosis rate and cell cycle distribution of KYSER treated by CEs were quantitatively determined by flow cytometry.Clone formation experiment was used to detect the clone formation ability of KYSER treaded by CEs.Results The main components of CEs were eugenol,eugenol hydrocarbon,and eugenol acetate.In vitro cell culture showed that 0.4%CEs could inhibit KYSER growth.MTT assay showed that the concentration of CEs≥0.5%could inhibit the survival of KYSER in a dosedependent manner.TEM assay showed that after treated by 0.5%CEs,KYSER’s microvilli became shorter and wider,ribosomes in the cytoplasm decreased,mitochondria atrophied,and a large number of autophagosomes were formed.Transwell migration assay showed that relative migration rates of KYSER after treated by 0.5%CEs and 0.6%CEs were(65±4)%and(41±3)%,respectively.Compared with the control group,the differences were statistically significant(P<0.001).Flow cytometry showed that the apoptosis rates of the control group,the 0.5%CEs treated group,and the 0.6%CEs treated group were(5.63±0.50)%,(11.77±0.42)%,and(19.44±0.19)%,respectively,and the differences between the control group and the two CEs treated groups were statistically significant(P<0.001).Flow cytometry showed that the G1 phase ratios of cells in the control group,t

关 键 词：放射抗拒 丁香 丁香酚 食管癌 

分 类 号：R285.5[医药卫生—中药学]