CAJ | 학술논문

目的检测促微管聚合蛋白3(TPPP3)在肝癌组织中的表达,观察靶向沉默TPPP3对肝癌细胞生物学功能的影响.方法应用实时荧光定量聚合酶链反应(FQ-PCR)技术检测23例肝癌组织和配对癌旁组织,Western blot和FQ-PCR技术检测肝癌细胞和正常永生化的肝细胞中TPPP3的表达.设计并筛选靶向沉默TPPP3的有效短发夹RNA(shRNA),并选取TPPP3表达量较高的SMMC-7721细胞和QGY-7703细胞进行功能实验.细胞计数试剂盒(CCK-8)法检测细胞增殖能力的改变;流式细胞术检测细胞凋亡率的改变;Transwell实验检测细胞迁移能力的改变;裸鼠皮下移植瘤实验检测TPPP3的体内生物学效应;裸鼠肺转移试验检测TPPP3对在体肝癌转移能力的影响.结果 TPPP3在肝癌组织中的表达量(0.374±0.046)显著高于配对癌旁组织(0.113 ±0.044,P＜0.01),在肝癌细胞中的表达量显著高于正常肝细胞;靶向沉默TPPP3可显著抑制肝癌细胞的增殖能力(QGY-7703:0.873 ±0.044比0.631±0.037;SMMC-7721:0.829±0.049比0.502±0.048)并增加细胞的凋亡率[SMMC-7721:(8.702±1.131)％比(4.213±0.098)％;QGY-7703:(14.731±2.247)％比(7.013±1.098)％];靶向沉默TPPP3可抑制肝癌细胞的转移能力(SMMC-7721:105±23比75±14,P＜0.05;QGY-7703:94±20比42±10,P＜0.01);靶向沉默TPPP3可显著抑制裸鼠皮下移植瘤的生长[肿瘤体积:(3 062.800±694.268) mm3比(850.200±469.387) mm3,P＜0.01;肿瘤重量:(1.401±0.202)g比(0.803±0.097)g,P＜0.01]以及肺转移(1.400±1.140比0.000±0.000,P＜0.01).结论肝癌中TPPP3过表达,TPPP3通过凋亡依赖的方式调控细胞增殖.此外,TPPP3还可调控癌细胞转移和体内肿瘤生长.
목적 검측촉미관취합단백3(TPPP3)재간암조직중적표체,관찰파향침묵TPPP3대간암세포생물학공능적영향.방법 응용실시형광정량취합매련반응(FQ-PCR)기술검측23례간암조직화배대암방조직,Western blot화FQ-PCR기술검측간암세포화정상영생화적간세포중TPPP3적표체.설계병사선파향침묵TPPP3적유효단발협RNA(shRNA),병선취TPPP3표체량교고적SMMC-7721세포화QGY-7703세포진행공능실험.세포계수시제합(CCK-8)법검측세포증식능력적개변;류식세포술검측세포조망솔적개변;Transwell실험검측세포천이능력적개변;라서피하이식류실험검측TPPP3적체내생물학효응;라서폐전이시험검측TPPP3대재체간암전이능력적영향.결과 TPPP3재간암조직중적표체량(0.374±0.046)현저고우배대암방조직(0.113 ±0.044,P＜0.01),재간암세포중적표체량현저고우정상간세포;파향침묵TPPP3가현저억제간암세포적증식능력(QGY-7703:0.873 ±0.044비0.631±0.037;SMMC-7721:0.829±0.049비0.502±0.048)병증가세포적조망솔[SMMC-7721:(8.702±1.131)％비(4.213±0.098)％;QGY-7703:(14.731±2.247)％비(7.013±1.098)％];파향침묵TPPP3가억제간암세포적전이능력(SMMC-7721:105±23비75±14,P＜0.05;QGY-7703:94±20비42±10,P＜0.01);파향침묵TPPP3가현저억제라서피하이식류적생장[종류체적:(3 062.800±694.268) mm3비(850.200±469.387) mm3,P＜0.01;종류중량:(1.401±0.202)g비(0.803±0.097)g,P＜0.01]이급폐전이(1.400±1.140비0.000±0.000,P＜0.01).결론 간암중TPPP3과표체,TPPP3통과조망의뢰적방식조공세포증식.차외,TPPP3환가조공암세포전이화체내종류생장.
Objective To examine the expression level of Tubulin polymerization promoting protein 3 (TPPP3) in hepatocellular carcinoma and the biological role of TPPP3 in hepatocellular carcinoma (HCC) cells.Methods Real-time fluorescent quantitative polymerase chain reaction (FQ-PCR) was employed to detect the expression level of TPPP3 in 28 cancer tissues and paired adjacent tissues.Western blotting and FQ-PCR were used to detect the level of TPPP3 in HCC cells and immortalized hepatocytes.Effective short hairpin RNA (shRNA) for silencing TPPP3 was designed and screened.SMMC-7721 and QGY-7703 cells,which had a high level of TPPP3,were used for functional assay.Cell counting kit-8 (CCK-8) was employed for cell proliferation assay.Flow cytometry was employed for cell apoptosis assay.Transwell analysis was used for determining the migration ability.Xenografted subcutaneous tumor experiment was used for examining the in vivo function of TPPP3.Lung metastasis assay of nude mice was employed for evaluating the role of TPPP3 in in vivo metastasis.Results TPPP3 was highly expressed in tumor tissues as compared with adjacent normal tissues (0.374 ± 0.046 vs.0.113 ± 0.044,P ＜ 0.01),and in HCC cell lines as compared with immortalized hepatocytes.Silencing TPPP3 in SMMC-7721 and QGY-7703 cells could inhibit cell proliferation (for QGY-7703:0.873 ± 0.044 vs.0.631 ± 0.037;for SMMC-7721:0.829 ±0.049 vs.0.502 ±0.048),which was in accordance with an increase of apoptosis rate [SMMC-7721:(8.702 ± 1.131) ％ vs.(4.213 ± 0.098) ％;for QGY-7703:(14.731 ± 2.247) ％ vs.(7.013 ± 1.098)％].Silencing TPPP3 could inhibit the migration ability of HCC cells (for SMMC-7721:105 ±23 vs.75 ± 14,P＜0.05;for QGY-7703:94 ± 20 vs.42± 10,P＜0.01).Silencing TPPP3 significantly inhibited xenograft tumor growth [for tumor volume:(3 062.800 ± 694.268) mm3 vs.(850.200±469.387) mm3,P＜0.01;for tumor weight:(1.401 ±0.202) g vs.(0.803 ±0.097) g,P＜ 0.01] and lung metastasis (1.400 ± 1.140 vs.0.000 ±0.000,P ＜0.01).Conclusion TPPP3 was overexpressed in HCC cells.TPPP3 could regulate apoptosis-dependent cell proliferation.Furthermore,TPPP3 could regulate cell migration and in vivo tumor growth.TPPP3 might be a novel oncogene in HCC.