中华普通外科杂志
中華普通外科雜誌
중화보통외과잡지
CHINESE JOURNAL OF GENERAL SURGERY
2012年
12期
1006-1009
,共4页
陈宗静%杨云秀%吕万治%白永恒%张行%刘彪%王本泉%梁勇%郑建建%陈必成
陳宗靜%楊雲秀%呂萬治%白永恆%張行%劉彪%王本泉%樑勇%鄭建建%陳必成
진종정%양운수%려만치%백영항%장행%류표%왕본천%량용%정건건%진필성
胰腺肿瘤%细胞凋亡%受体,notch%曲古抑菌素A
胰腺腫瘤%細胞凋亡%受體,notch%麯古抑菌素A
이선종류%세포조망%수체,notch%곡고억균소A
Pancreatic neoplasms%Apoptosis%Receptors,notch%Trichostatin A
目的 研究曲古抑菌素A(Trichostatin A,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 ～0.4 μmol/L不同浓度TSA处理胰腺癌细胞.四甲基偶氮唑蓝(MTT)法检测各组细胞的存活率,通过Hoechst 33258染色直接观察细胞凋亡.实时荧光定量PCR(RT-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eaf2和肿瘤转移相关基因cytohesin 1、2、3、4的表达.蛋白质印迹法检测细胞caspase-3、bcl-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、0.4 μmol/L TSA作用PANC-1 24 h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-1细胞死亡逐渐增加,Hoechst 33258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、dll3 mRNA表达下降(P＜0.05),而Notch1 mRNA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cytohesin mRNA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.
目的 研究麯古抑菌素A(Trichostatin A,TSA)榦預對胰腺癌細胞株PANC-1凋亡及相關基因錶達的影響,探討其作用機製.方法 採用0.1 ～0.4 μmol/L不同濃度TSA處理胰腺癌細胞.四甲基偶氮唑藍(MTT)法檢測各組細胞的存活率,通過Hoechst 33258染色直接觀察細胞凋亡.實時熒光定量PCR(RT-qPCR)方法檢測包括Notch信號通路相關的基因、腫瘤增殖相關基因eaf2和腫瘤轉移相關基因cytohesin 1、2、3、4的錶達.蛋白質印跡法檢測細胞caspase-3、bcl-2、bax以及Notch-1的胞內活性形式NICD蛋白錶達.結果 TSA對PANC-1的增殖抑製作用呈明顯濃度和時間依賴性,0.1、0.2、0.4 μmol/L TSA作用PANC-1 24 h後存活率分彆為72％、58％和39％.顯微鏡觀察髮現隨著TSA濃度增加和作用時間延長,PANC-1細胞死亡逐漸增加,Hoechst 33258染色可見凋亡典型特徵的PANC-1細胞比例增加.TSA作用PANC-1後,蛋白質印跡法結果顯示caspase-3、bax以及NICD蛋白錶達增加,而bcl-2蛋白錶達下降.qRT-PCR結果顯示Notch通路中相關基因numb、hes6mRNA錶達升高,gcn512、dll3 mRNA錶達下降(P＜0.05),而Notch1 mRNA錶達沒有變化.腫瘤增殖相關基因eaf2以及腫瘤轉移相關基因cytohesin mRNA錶達未髮現有變化(P＞0.05). 結論 TSA可誘導胰腺癌PANC-1細胞凋亡,且呈劑量依賴性.Notch通路相關基因可能參與瞭TSA誘導細胞凋亡的過程.
목적 연구곡고억균소A(Trichostatin A,TSA)간예대이선암세포주PANC-1조망급상관기인표체적영향,탐토기작용궤제.방법 채용0.1 ～0.4 μmol/L불동농도TSA처리이선암세포.사갑기우담서람(MTT)법검측각조세포적존활솔,통과Hoechst 33258염색직접관찰세포조망.실시형광정량PCR(RT-qPCR)방법검측포괄Notch신호통로상관적기인、종류증식상관기인eaf2화종류전이상관기인cytohesin 1、2、3、4적표체.단백질인적법검측세포caspase-3、bcl-2、bax이급Notch-1적포내활성형식NICD단백표체.결과 TSA대PANC-1적증식억제작용정명현농도화시간의뢰성,0.1、0.2、0.4 μmol/L TSA작용PANC-1 24 h후존활솔분별위72％、58％화39％.현미경관찰발현수착TSA농도증가화작용시간연장,PANC-1세포사망축점증가,Hoechst 33258염색가견조망전형특정적PANC-1세포비례증가.TSA작용PANC-1후,단백질인적법결과현시caspase-3、bax이급NICD단백표체증가,이bcl-2단백표체하강.qRT-PCR결과현시Notch통로중상관기인numb、hes6mRNA표체승고,gcn512、dll3 mRNA표체하강(P＜0.05),이Notch1 mRNA표체몰유변화.종류증식상관기인eaf2이급종류전이상관기인cytohesin mRNA표체미발현유변화(P＞0.05). 결론 TSA가유도이선암PANC-1세포조망,차정제량의뢰성.Notch통로상관기인가능삼여료TSA유도세포조망적과정.
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 cells were measured by MTT assay and Hoechst 33258 staining,respectively.mRNA expression levels of the genes,numb,gcn512,dll3,hes6,eaf2,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 μmol/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 bcl-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 upregulated 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 eaf2 and cytohesin1,2,3,4 were not affected by TSA.Conclusions TSA induces apoptosis of pancreatic cancer cell line PANC-1.The Notch signal pathway may be involved in inducing cellular apoptosis of PANC-1 when cultured with TSA.
