中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2013年
1期
61-63
,共3页
王涌%刘雅辉%姜建帅%崔翰斌
王湧%劉雅輝%薑建帥%崔翰斌
왕용%류아휘%강건수%최한빈
癌,肝细胞%肿瘤干细胞%苦参碱%增殖%转移
癌,肝細胞%腫瘤榦細胞%苦參堿%增殖%轉移
암,간세포%종류간세포%고삼감%증식%전이
Carcinoma,hepatocellular%Tumor stem cell%Matrine%Proliferation%Metastasis
目的 观察苦参碱对肝癌干细胞增殖及侵袭转移能力的影响,探讨苦参碱对肝癌干细胞的作用机制.方法 应用无血清肿瘤干细胞培养基,悬浮培养人肝癌细胞株SMMC-7721形成克隆,收集后裸鼠皮下接种并予以顺铂筛选,取耐药细胞继续悬浮培养获得干细胞克隆.采用不同浓度苦参碱体外作用72 h后,噻唑蓝(MTT)比色法观察肝癌干细胞的增殖能力变化,荧光定量聚合酶链反应(PCR)、Westem blot法检测侵袭转移相关基因细胞黏附调节基因(CAR)、E-钙黏蛋白(E-cadherin)、层粘连蛋白(Laminin)、纤连蛋白(Fibronectin)表达水平的变化.结果 在肿瘤干细胞培养基和顺铂的双重筛选下,SMMC-7721细胞能形成悬浮生长的干细胞克隆.5、50、100、200 mg/L苦参碱体外作用72 h后,可明显抑制肝癌干细胞的增殖,抑制率分别为(3.0±0.3)％、(26.9±0.5)％、(55.1±0.6)％、(77.4±0.8)％.荧光定量PCR结果显示,50 mg/L苦参碱可诱导肿瘤干细胞CAR、E-cadherin,Laminin、Fibronectin基因表达上调至未处理组的15.2、8.5、4.9和6.3倍,差异有统计学意义,与Western blot的结果一致.结论 一定浓度的苦参碱可明显抑制肝癌干细胞的体外增殖,并抑制其侵袭转移能力.
目的 觀察苦參堿對肝癌榦細胞增殖及侵襲轉移能力的影響,探討苦參堿對肝癌榦細胞的作用機製.方法 應用無血清腫瘤榦細胞培養基,懸浮培養人肝癌細胞株SMMC-7721形成剋隆,收集後裸鼠皮下接種併予以順鉑篩選,取耐藥細胞繼續懸浮培養穫得榦細胞剋隆.採用不同濃度苦參堿體外作用72 h後,噻唑藍(MTT)比色法觀察肝癌榦細胞的增殖能力變化,熒光定量聚閤酶鏈反應(PCR)、Westem blot法檢測侵襲轉移相關基因細胞黏附調節基因(CAR)、E-鈣黏蛋白(E-cadherin)、層粘連蛋白(Laminin)、纖連蛋白(Fibronectin)錶達水平的變化.結果 在腫瘤榦細胞培養基和順鉑的雙重篩選下,SMMC-7721細胞能形成懸浮生長的榦細胞剋隆.5、50、100、200 mg/L苦參堿體外作用72 h後,可明顯抑製肝癌榦細胞的增殖,抑製率分彆為(3.0±0.3)％、(26.9±0.5)％、(55.1±0.6)％、(77.4±0.8)％.熒光定量PCR結果顯示,50 mg/L苦參堿可誘導腫瘤榦細胞CAR、E-cadherin,Laminin、Fibronectin基因錶達上調至未處理組的15.2、8.5、4.9和6.3倍,差異有統計學意義,與Western blot的結果一緻.結論 一定濃度的苦參堿可明顯抑製肝癌榦細胞的體外增殖,併抑製其侵襲轉移能力.
목적 관찰고삼감대간암간세포증식급침습전이능력적영향,탐토고삼감대간암간세포적작용궤제.방법 응용무혈청종류간세포배양기,현부배양인간암세포주SMMC-7721형성극륭,수집후라서피하접충병여이순박사선,취내약세포계속현부배양획득간세포극륭.채용불동농도고삼감체외작용72 h후,새서람(MTT)비색법관찰간암간세포적증식능력변화,형광정량취합매련반응(PCR)、Westem blot법검측침습전이상관기인세포점부조절기인(CAR)、E-개점단백(E-cadherin)、층점련단백(Laminin)、섬련단백(Fibronectin)표체수평적변화.결과 재종류간세포배양기화순박적쌍중사선하,SMMC-7721세포능형성현부생장적간세포극륭.5、50、100、200 mg/L고삼감체외작용72 h후,가명현억제간암간세포적증식,억제솔분별위(3.0±0.3)％、(26.9±0.5)％、(55.1±0.6)％、(77.4±0.8)％.형광정량PCR결과현시,50 mg/L고삼감가유도종류간세포CAR、E-cadherin,Laminin、Fibronectin기인표체상조지미처리조적15.2、8.5、4.9화6.3배,차이유통계학의의,여Western blot적결과일치.결론 일정농도적고삼감가명현억제간암간세포적체외증식,병억제기침습전이능력.
Objective To investigate the effect of matrine on proliieration,invasion and metastasis of hepatocellular carcinoma (HCC) stem ceils in vitroand the action mechanims.Methods SMMC-7721cells were cultured in tumor stem cell specific medium to form sphere,harvested,inoculated subcutaneously into the nude mice,and treated with different concentrations of cisplatin.The drug resistant cells were resuspended and purified.Methyl thiazol tetrazolium (MTT) test was performed to detect the inhibitory effects of matrine against tumor proliferation.Real-time polymerase chain reaction (real-time PCR) and Western blotting were used to detect the changes of CAR,E-cadherin,Laminin and Fibronectin.Results Through the double filter of TSC medium and cisplatin-resistance,SMMC-7721 stem cells could form spheres in TSC medium.The inhibition ratio of HCC stem ceils was (3.0 ± 0.3) ％,(26.9 ± 0.5) ％,(55.1 ± 0.6) ％,and (77.4 ± 0.8) ％,respectively in the presence of matrine (5,50,100,200 mg/L),for 72 h in vitro.The real-time PCR revealed that 50 mg/L matrine increased the expression of cell adhesion regulates gene (CAR),E-cadherin,Laminin and Fibronectin up to 15.2,8.5,4.9 and 6.3 times respectively,which was consistent to the result of Western blotting.Conclusion Appropriate concentration of matrine could significantlv inhibite the proliferation and invasion of HCC stem cells.
