CAJ | 학술논문

目的检测结肠癌转移相关基因1(MACC1)在卵巢癌细胞株中的表达,并探讨用siRNA技术抑制其表达对卵巢癌细胞生物学行为的影响.方法应用实时荧光定量PCR (RT-qPCR)及Western blot法检测OVCAR3、ES-2、SKOV3、HO-8910卵巢癌细胞株中MACC1的表达,合成MACC1特异性siRNA并转染OVCAR3细胞,利用RT-qPCR筛选并鉴定MACC 1基因有效沉默后,应用体外黏附实验、Transwell迁移及侵袭实验、体外血管拟态实验检测MACC1基因沉默后OVCAR3细胞的体外黏附、迁移、侵袭及血管生成能力的变化.结果 OVCAR3细胞较其他卵巢癌细胞株高表达MACC1.MACC1基因沉默后,OVCAR3细胞的体外黏附能力受到不同程度的抑制;Transwell迁移实验中,MACC1 siRNA干扰的OVCAR3细胞(干扰组)转入底层膜的细胞数为(245.5±12.8)个,低于阴性对照组和空白对照组[分别为(500.3±16.5)、(496.3±13.1)个,均P＜0.05]; Transwell侵袭实验中,干扰组转入底层膜的细胞数为(185.3±14.1)个,低于阴性对照组和空白对照组[分别为(405.7±9.1)、(416.3±11.5)个,均P＜0.05];体外血管拟态显示干扰组细胞多呈散在分布,连接减少,形成的完整结构少.结论利用siRNA技术抑制MACC1基因表达可有效抑制卵巢癌OVCAR3细胞的体外转移和侵袭能力,MACC1有望成为卵巢癌治疗的靶基因.
목적 검측결장암전이상관기인1(MACC1)재란소암세포주중적표체,병탐토용siRNA기술억제기표체대란소암세포생물학행위적영향.방법 응용실시형광정량PCR (RT-qPCR)급Western blot법검측OVCAR3、ES-2、SKOV3、HO-8910란소암세포주중MACC1적표체,합성MACC1특이성siRNA병전염OVCAR3세포,이용RT-qPCR사선병감정MACC 1기인유효침묵후,응용체외점부실험、Transwell천이급침습실험、체외혈관의태실험검측MACC1기인침묵후OVCAR3세포적체외점부、천이、침습급혈관생성능력적변화.결과 OVCAR3세포교기타란소암세포주고표체MACC1.MACC1기인침묵후,OVCAR3세포적체외점부능력수도불동정도적억제;Transwell천이실험중,MACC1 siRNA간우적OVCAR3세포(간우조)전입저층막적세포수위(245.5±12.8)개,저우음성대조조화공백대조조[분별위(500.3±16.5)、(496.3±13.1)개,균P＜0.05]; Transwell침습실험중,간우조전입저층막적세포수위(185.3±14.1)개,저우음성대조조화공백대조조[분별위(405.7±9.1)、(416.3±11.5)개,균P＜0.05];체외혈관의태현시간우조세포다정산재분포,련접감소,형성적완정결구소.결론 이용siRNA기술억제MACC1기인표체가유효억제란소암OVCAR3세포적체외전이화침습능력,MACC1유망성위란소암치료적파기인.
Objective To examine the expression of metastasis-associated in colon cancer 1 (MACC1) gene in ovarian cancer cell lines and investigate its effect on biological behaviors of ovarian cancer cells.Methods The expression of MACC1 was examined by qRT-PCR and Western blot analysis in four ovarian cancer cell lines inculding OVCAR3,ES-2,SKOV3 and HO-8910.When the MACC1 was transfected to OVCAR3 cells,fluorogenic quantitative PCR was used to filter and identify MACC1 gene after the efficient silencing.Changes of adhesion in the cells were analyzed by an adhesion assay.Transwell migration and invasion assay and in vitro vascular mimicry assay were used to detect migration,invasion and angiogenesis of OVCAR3 cells in vitro.Results The expression of MACC1 gene was higher in OVCAR3 compared to other cell lines.qRT-PCR confirmed that the expression of MACC1 was silenced successfully after transient transfected MACC1-siRNA into OVCAR3 cells.After successful silencing the MACC1 expression,the adhesion ability was inhibited to some degree.In transwell migration assay,the numbers of cells in upper chamber passing through the membrane in transfected group were less than control groups (245.5 ±12.8,500.3±16.5 and 496.3±13.1 respectively),while in transwell invasion assay,the numbers of cells in upper chamber passing through the membrane in transfected group were less than the negative group and control group (185.3±14.1,405.7±9.1 and 416.3±11.5 respectively),both with markedly differences among the three groups.In tube formation assay,the distrubition of HUVECs was diffused with less junctions,and the average number of complete tubular structure was decreased in transfected group compared to the corresponding controls.Conclusion RNA interference inhibits the expression of MACC1 and effectively inhibits the metastasis and invasion abilities of ovarian cancer cells in vitro,and MACC1 is expected to become the target gene of ovarian cancer treatment.