CAJ | 학술논문

目的探讨失巢凋亡抑制因子酪氨酸激酶受体B(TrkB)介导的卵巢上皮性癌(卵巢癌)OVCAR3细胞的失巢凋亡抑制与其侵袭的关系.方法采用RT-PCR和实时定量PCR技术检测4种细胞包括3种不同病理类型的卵巢癌细胞系OVCAR3、SKOV3(两者均为卵巢浆液性乳头状囊腺癌细胞系)和ES2(卵巢透明细胞癌细胞系)细胞及神经母细胞瘤细胞系sK-N-DZ细胞中,以及经不同方式[贴壁培养和立体培养(培养后形成团簇细胞)]培养的OVCAR3细胞中TrkB mRNA的表达.转染TrkB小分子干扰RNA(siRNA)后,流式细胞仪检测OVCAR3细胞的凋亡率,以转染无意义的(scrambled)siRNA者为阴性对照;体内、体外侵袭实验检测不同方式培养的OVCAR3细胞的侵袭能力.结果 (1)OVCAR3细胞中TrkB mRNA表达水平为0.0240±0.0017,明显高于SKOV3细胞的0.0030±0.0006、ES2细胞的0.0027±0.0009及SK-N-DZ细胞的0.0087±0.0003(P＜0.01);OVCAR3团簇细胞中TrkB mRNA表达水平为0.0437±0.0021,明显高于贴壁细胞的0.0240±0.0017(P＜0.01).(2)转染TrkB siRNA后,OVCAR3细胞的凋亡率为(46.3±5.9)%,明显高于阴性对照的(9.0±0.8)%(P＜0.01).(3)体外侵袭实验显示,OVCAR3团簇细胞的穿膜细胞数为(71.8±0.8)个,明显多于贴壁细胞的(47.7±0.8)个(P＜0.01);体内侵袭实验显示,转染TrkB siRNA后,OVCAR3细胞在裸鼠体内形成的肿瘤体积为(6.0±1.4)mm3,明显小于阴性对照的(16.3±4.7)mm3(P＜0.01).结论 TrkB可能通过诱导卵巢癌OVCAR3细胞的失巢凋亡抑制而赋予其高侵袭能力.
목적 탐토실소조망억제인자락안산격매수체B(TrkB)개도적란소상피성암(란소암)OVCAR3세포적실소조망억제여기침습적관계.방법 채용RT-PCR화실시정량PCR기술검측4충세포포괄3충불동병리류형적란소암세포계OVCAR3、SKOV3(량자균위란소장액성유두상낭선암세포계)화ES2(란소투명세포암세포계)세포급신경모세포류세포계sK-N-DZ세포중,이급경불동방식[첩벽배양화입체배양(배양후형성단족세포)]배양적OVCAR3세포중TrkB mRNA적표체.전염TrkB소분자간우RNA(siRNA)후,류식세포의검측OVCAR3세포적조망솔,이전염무의의적(scrambled)siRNA자위음성대조;체내、체외침습실험검측불동방식배양적OVCAR3세포적침습능력.결과 (1)OVCAR3세포중TrkB mRNA표체수평위0.0240±0.0017,명현고우SKOV3세포적0.0030±0.0006、ES2세포적0.0027±0.0009급SK-N-DZ세포적0.0087±0.0003(P＜0.01);OVCAR3단족세포중TrkB mRNA표체수평위0.0437±0.0021,명현고우첩벽세포적0.0240±0.0017(P＜0.01).(2)전염TrkB siRNA후,OVCAR3세포적조망솔위(46.3±5.9)%,명현고우음성대조적(9.0±0.8)%(P＜0.01).(3)체외침습실험현시,OVCAR3단족세포적천막세포수위(71.8±0.8)개,명현다우첩벽세포적(47.7±0.8)개(P＜0.01);체내침습실험현시,전염TrkB siRNA후,OVCAR3세포재라서체내형성적종류체적위(6.0±1.4)mm3,명현소우음성대조적(16.3±4.7)mm3(P＜0.01).결론 TrkB가능통과유도란소암OVCAR3세포적실소조망억제이부여기고침습능력.
Objective To study the relationship between tyrosine kinase receptor B (TrkB)expression and anoikis-suppression and invasion in OVCAR3 ovarian cancer ceils. Methods The expression of TrkB mRNA in OVCAR3 ovarian cancer cells under two culture conditions :adhesive cells and ceil-spheroids were evaluated by RT-PCR and real-time PCR.The relationship between TrkB expression and anoikis-suppression of OVCAR3 ovarian cancer ceils was examined by RNA interference (RNAi) technic,anchorage independent culture and fluorescence-activated ceil sorting analysis.The difference in invasion and metastatic ability of OVCAR3 cells under two culture conditions and with or without TrkB silenced by small interfering RNA (siRNA) was investigated by matrigel invasion assay and in vivo studies.Results The expression of TrkB mRNA was highest in OVCAR3 ovarian cancer ceils,0.0240 ~ 0.0017,compared with the other three cell lines,0.0030±0.0006,0.0027±0.0009 and 0.0087±0.0003 respectively,andthe expression in OVCAR3 multicellular spheroids was significantly higher than that in ceils under monolayer adhesive culture,0.0437±0.0021 versus 0.0240±0.0017 (P＜0.01) . TrkB mediated anoikissuppression in OVCAR3 ovarian cancer ceils.OVCAR3 multiceilular spheroids had a higher invasion ability than OVCAR3 cells under monolayer adhesive culture,and the penetrating cells of the two groups were 71.8± 0.8 and 47.7±0.8 respectively (P＜0.01 ).The metastatic ability of OVCAR3 cells was attenuated when TrkB was silenced,and the volume of the tumors developed by OVCAR3 adhesive cells and OVCAR3 adhesive cells with TrkB silenced were (16.3±4.7) mm3 and(6.0±1.4) mm3 respectively (P＜0.01).Conclusion As an anoikis-suppressur,TrkB may increase the invasion and metastasis of OVCAR3 ovarian cancer cells.