促微管聚合蛋白3在肝癌中的表达及其对肝癌细胞生物学功能的影响

Expression of tubulin polymerization promoting protein 3 in hepatocellular carcinoma and its biological function in hepatocellular carcinoma cells

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，Westernblot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表达撤较高的SMMC-7721细胞和QCY-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；裸鼠皮下移悄瘤实验检测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在肝癌组织巾的表达量（0．374±0．046）显著高于配对癌旁组织（0．113±0．044，P〈0．01），在肝癌细胞巾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1-移植瘤的生长[肿瘤体积：（3062．800±694．268）mm3比（850．200±469．387）mm3，P〈O．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通过凋广依赖的方式蒯控细胞增殖。此外，TPPP3还可涮控癌细胞转移和体内肿瘤生长。

Objective To examine the expression level of Tubulin polymerizatior, promoting protein 3 （ TPPP3 ） in hepatocellular carcinoma and the biological role of TPPP3 in hepatocellular carcinoma （ltCC） 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 eytometry was employed for cell apoptosis as- say. Transwell analysis was used for determining the migration ability. XenogrMted subcutaneous tumor ex- periment 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 ceils 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 （lot 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. 011 and lung metastasis （ 1. 400 ± 1. 140 vs. 0. 000 ±0. 000 ,P 〈0. 01 ）. Conclusion TPPP3 was overex- pressed in HCC ceils. 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.