原花青素通过自噬途径增强喉癌细胞对顺铂化疗敏感性的研究  

Procyanidins enhance the chemotherapeutic sensitivity of laryngeal carcinoma cells to cisplatin through autophagy pathway

作　　者：于锋[1,2,3] 刘炜 龚小蓉 周毅波 林颖[1,2,3] YU Feng;LIU Wei;GONG Xiaorong;ZHOU Yibo;LIN Ying(Institute of Otolaryngology Head and Neck Surgery Affiliated to Guangzhou Medical Univer- sity, Guangzhou, 510620;Guangzhou Hospital of Otolaryngology Head and Neck Surgery;Guangzhou 12th People＇s Hospital)

机构地区：[1]广州医科大学耳鼻咽喉头颈外科研究所,广州510620 [2]广州市耳鼻咽喉头颈外科医院 [3]广州市第十二人民医院

出　　处：《临床耳鼻咽喉头颈外科杂志》2018年第6期447-451,456,共6页Journal of Clinical Otorhinolaryngology Head And Neck Surgery

基　　金：广州市喉癌临床医学研究与转化中心试点建设项目(No:2060404)

摘　　要：目的:探究原花青素(OPC)对喉癌TU686细胞自噬的影响,并从自噬和凋亡的角度讨论OPC增强喉癌细胞对顺铂(DDP)化疗敏感性。方法:CCK-8法检测不同浓度OPC和DDP对TU686细胞活力的影响;实验分组TU686细胞分为CON组、DDP组,OPC组、MIX组;Annexin-V-FITC/PI双染行流式细胞术检测各实验组对细胞凋亡的作用;细胞免疫荧光染色检测细胞自噬的形成;Western blot检测自噬及凋亡相关蛋白的表达;使用自噬抑制剂(3-MA)研究自噬对细胞凋亡的影响。结果:CCK-8结果显示OPC和DDP作用TU686细胞24h后,都以浓度依赖性方式抑制TU686细胞增殖活力。LC3-Ⅱ蛋白染色观察到,与CON和DDP组、OPC组相比,MIX组作用TU686细胞后显著诱导自噬形成(P<0.05)。流式检测结果提示相比CON组,DDP组、OPC组和MIX组诱导TU686细胞凋亡,其中MIX组促凋亡作用明显高于OPC组和DDP组(P<0.05),使用3-MA预处理后,OPC组及MIX组对喉癌细胞的凋亡作用显著下降(P<0.05)。Western blot的结果提示DDP、OPC及MIX组的LC3-Ⅱ与Caspase-3蛋白的表达明显高于CON组(P<0.05),而其中MIX组较之于单药组,其LC3-Ⅱ与Caspase-3表达的增加亦差异有统计学意义(P<0.05)。使用3-MA抑制自噬后,LC3-Ⅱ的表达显著下降(P<0.05),Caspase-3的表达伴随LC3-Ⅱ发生降低,但Caspase-3表达的下降只在OPC及MIX组具有显著意义(P<0.05)。结论:OPC可以通过诱导喉癌TU686细胞自噬,促进其凋亡,进而增强喉癌细胞对顺铂化疗敏感性。Objective：To investigate the effect of Procyanidins（OPCs）on the autophagy of laryngeal cancer cell line TU686 and to explore the effect of OPCs on the chemosensitivity of laryngeal cancer cells to DDP in terms of autophagy and apoptosis.Method：CCK-8 was used to detected the effect of different concentrations of OPC and DDP on TU686 cell viability.Experimental grouping：Both kinds of cells were divided into CON group,DDP group,OPC group and MIX group.Annexin-V-FITC/PI double staining of flow cytometry was used to detected the effect of each experimental group on the apoptosis.Cell immunofluorescence staining was used to detect the formation of autophagy.Western blot was used to detect the expression of autophagy-related and apoptosis-related proteins.Autophagy inhibitors（3-MA）were used to study the effect of autophagy on apoptosis.Result：The results of CCK-8 showed that TU686 cells were inhibited by OPC and DDP in a concentration-dependent manner for24 hours.LC3-Ⅱ protein staining showed that compared with CON group,DDP group and OPC group,MIX group significantly induced autophagy formation in TU686 cells（P〈0.05）.Flow cytometry showed that compared with CON group,apoptosis of TU686 cells was induced in DDP group,OPC group and MIX group.And the effect of MIX on apoptosis was significantly higher than that of OPC and DDP groups（P〈0.05）.After pretreatment with 3-MA,the apoptotic effect of OPC group and MIX group on TU686 cells was significantly decreased（P〈0.05）.Western blot results showed that the expression of LC3-Ⅱ and Caspase-3 in DDP,OPC and MIX groups was significantly higher than that in CON group（P〈0.05）.In MIX group,the expression of LC3-Ⅱ and Caspase-3 also had significant difference（P〈0.05）compared with single drug group.After using 3-MA to inhibit autophagy,the expression of LC3-Ⅱ was significantly decreased（P〈0.05）,and the expression of Caspase-3 was decreased along with LC3-Ⅱ,but the decrease of Caspase-3 expression was only significant in OPC

关 键 词：原花青素 喉癌 自噬 凋亡 

分 类 号：R739.65[医药卫生—肿瘤]