曲古抑菌素A诱导胰腺癌细胞株凋亡和Notch信号通路改变的研究  

Trichostatin A induces apoptosis and alteration of Notch pathway in human pancreatic cancer cellline PANC-1

作　　者：陈宗静[1] 杨云秀[1] 吕万治[1] 白永恒 张行 刘彪[1] 王本泉[1] 梁勇 郑建建 陈必成 

机构地区：[1]温州医学院附属第一医院肝胆胰外科,浙江省325000 [2]温州市重点实验室,浙江省325000

出　　处：《中华普通外科杂志》2012年第12期1006-1009,共4页Chinese Journal of General Surgery

摘　　要：目的研究曲古抑菌素A（TrichostatinA，TSA）干预对胰腺癌细胞株PANC．1凋亡及相关基因表达的影响，探讨其作用机制。方法采用0．1～0．4μmol／L不同浓度TSA处理胰腺癌细胞。四甲基偶氮唑蓝（MTr）法检测各组细胞的存活率，通过Hoechst33258染色直接观察细胞凋亡。实时荧光定量PCR（1iT-qPCR）方法检测包括Notch信号通路相关的基因、肿瘤增殖相关基因eat2和肿瘤转移相关基因cytohesin1、2、3、4的表达。蛋白质印迹法检测细胞caspase-3、bel-2、bax以及Notch-1的胞内活性形式NICD蛋白表达。结果TSA对PANC-1的增殖抑制作用呈明显浓度和时间依赖性，0．1、0．2、0．4μmol／LTSA作用PANC-124h后存活率分别为72％、58％和39％。显微镜观察发现随着TSA浓度增加和作用时间延长，PANC一1细胞死亡逐渐增加，Hoechst33258染色可见凋亡典型特征的PANC-1细胞比例增加。TSA作用PANC-1后，蛋白质印迹法结果显示caspase-3、bax以及NICD蛋白表达增加，而bcl-2蛋白表达下降。qRT-PCR结果显示Notch通路中相关基因numb、hes6mRNA表达升高，gcn512、d113mRNA表达下降（P〈0．05），而NotchlmRNA表达没有变化。肿瘤增殖相关基因eaf2以及肿瘤转移相关基因cytohesinmRNA表达未发现有变化（P〉0．05）。结论TSA可诱导胰腺癌PANC-1细胞凋亡，且呈剂量依赖性。Notch通路相关基因可能参与了TSA诱导细胞凋亡的过程。Objective To investigate the efficiency of Trichostatin A （TSA） in inducing cell apoptosis and altering the Notch pathway genes expression in PANC-1 cells line. Methods The survival rate and apoptosis of PANC-1 ceils were measured by MTY assay and Hoechst 33258 staining, respectively. mRNA expression levels of the genes, numb, gcnSl2, dl13, hes6, eai2, cytohesins, in PANC-1 cells were assessed by real-time quantitive PCR. Western blot was used to measure the expression of bcl-2, bax, actived caspase-3 and NICD protein which was the biologically active form of Notch-1. Results After culturing with 0. 1, 0. 2, and 0. 4 p.mol/L TSA for 24 hours, the cellular survival rate of PANC-1 cells significantly decreased to 72% ,58% and 39%, respectively. The survival rate of PANC-1 was negatively correlated to time length of culture with TSA. Increased apoptosis of PANC-1 cells after 12, 24 and 36 h culture with TSA was detected by Hoechst 33258 staining. Western blotting showed that the expression of bax, actived caspase-3 and NICD protein increased while the bel-2 protein decreased after culture with TSA. In real time quantitive PCR assessment, the mRNA expression of numb and hes6 in PANC-1 cells were up- regulated by TSA （P 〈0. 05）, while the mRNA expression of gcn512 and dll3 were down-regulated by TSA （P 〈 0. 05 ）. While mRNA expressions of ear2 and cytohesinl, 2, 3,4 were not affected by TSA. Condusions ＇USA induces apoptosis of pancreatic cancer cell line PANC-1. The Notch signal pathway may be involved in inducing cellular anootosis of PANC-1 when cultured with TSA.

关 键 词：胰腺肿瘤 细胞凋亡 受体 NOTCH 曲古抑菌素A 

分 类 号：R735.9[医药卫生—肿瘤]