CAJ | 학술논문

目的探讨携带可溶性肿瘤坏死因子相关凋亡诱导配体(sTRAIL)基因的重组腺相关病毒载体rAAV-sTRAIL对肝癌细胞的增殖抑制及诱导凋亡作用,以及采用超声辐照联合微泡造影剂介导rAAV-sTRAIL转染肝癌细胞的有效性.方法单纯重组腺相关病毒rAAV-sTRAIL感染肝癌细胞株HepG2,以及利用超声辐照联合微泡造影剂介导重组腺相关病毒rAAV-sTRAIL感染HepG2细胞,分别以RT-PCR方法和Westen印迹法检测细胞内sTRAIL基因mRNA及蛋白表达量,噻唑蓝法检测细胞的增殖抑制率,流式细胞术检测细胞凋亡率.结果重组腺相关病毒rAAV-sTRAIL可有效转染HepG2细胞,sTRAIL mRNA表达水平在微泡造影剂组高于单纯病毒组,二者差异有统计学意义(P<0.05);rAAV-srrRAIL可抑制HepG2细胞增殖,其抑制率在微泡造影剂组高于单纯病毒组,二者差异有统计学意义(P<0.05);rAAV-sTRAIL可诱导HepG2细胞凋亡,其凋亡率在微泡造影剂组高于单纯病毒组,二者差异有统计学意义(P<0.05).结论构建出的重组腺相关病毒rAAV-sTRAIL在肝癌细胞中能有效表达,sTRAIL基因对肝癌细胞有明显的抑制增殖和促凋亡作用;联合超声微泡造影剂及低强度超声,可有效提高腺相关病毒载体在肝癌细胞中的感染率,从而增强sTRAIL对肝癌细胞的增殖抑制及诱导凋亡作用.
목적 탐토휴대가용성종류배사인자상관조망유도배체(sTRAIL)기인적중조선상관병독재체rAAV-sTRAIL대간암세포적증식억제급유도조망작용,이급채용초성복조연합미포조영제개도rAAV-sTRAIL전염간암세포적유효성.방법 단순중조선상관병독rAAV-sTRAIL감염간암세포주HepG2,이급이용초성복조연합미포조영제개도중조선상관병독rAAV-sTRAIL감염HepG2세포,분별이RT-PCR방법화Westen인적법검측세포내sTRAIL기인mRNA급단백표체량,새서람법검측세포적증식억제솔,류식세포술검측세포조망솔.결과 중조선상관병독rAAV-sTRAIL가유효전염HepG2세포,sTRAIL mRNA표체수평재미포조영제조고우단순병독조,이자차이유통계학의의(P<0.05);rAAV-srrRAIL가억제HepG2세포증식,기억제솔재미포조영제조고우단순병독조,이자차이유통계학의의(P<0.05);rAAV-sTRAIL가유도HepG2세포조망,기조망솔재미포조영제조고우단순병독조,이자차이유통계학의의(P<0.05).결론 구건출적중조선상관병독rAAV-sTRAIL재간암세포중능유효표체,sTRAIL기인대간암세포유명현적억제증식화촉조망작용;연합초성미포조영제급저강도초성,가유효제고선상관병독재체재간암세포중적감염솔,종이증강sTRAIL대간암세포적증식억제급유도조망작용.
Objective To evaluate the impact of the recombined adeno-associated virus encoding soluble tumor necrosis factor related apoptosis inducing ligand gene(rAAV-sTRAIL)on proliferation and apoptosis of human hepatocellular carcinoma(HCC)cells,and to investigate the feasibility and efficiency of transfection of rAAV-sTRAIL into human HCC cells by ultrasound microbubble intensitler.Methods Human HCC cells of the line HepG2 were transfected with rAAV-sTRAIL or rAAV-sTRAIL combined with microbubble echocontrast agent and appropriate dose of ultrasound irradiation.RT-PCR and Western blotting were used to detect the mRNA and protein expression of sTRAIL gene.MTT method Was used to detect the proliferation inhibition rate.and the apoptosis rate of the HepG2 cells Was evaluated by flow cytometry.Results The expression levels of sTRAIL mRNA and protein were higher in the rAAV-TRAIL combined with altrasound microbubble group than in the rAAV-sTRAIL group(both P<0.05).The proliferation inhibition rate of the rAAV-TRAIL combined with ultrasound microbubble group Was significandy higher than that of the rAAV-sTRAIL group(P<0.05).The apoptotic effect of the rAAV-TRAIL combined with ultrasound microbubble group was greater than that of the rAAV-sTRAIL group(P<0.05).Condusion TRAIL has a potential role to inhibit e proliferation and induce apoptosis of human hepatocellular carcinoma cells.Ultrasound and microbubble echocontrast agent increase the transfection rate of rAAV vector into HCC cells.