中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2015年
6期
1276-1278
,共3页
朱建强%张昌文%张蒙%盛镔%张志宏%徐勇
硃建彊%張昌文%張矇%盛鑌%張誌宏%徐勇
주건강%장창문%장몽%성빈%장지굉%서용
前列腺癌%增殖%CtBP2%RNA干扰
前列腺癌%增殖%CtBP2%RNA榦擾
전렬선암%증식%CtBP2%RNA간우
Prostate cancer%Proliferation%C-terminal-binding protein%RNA interference
目的 观察下调CtBP2对人前列腺癌C4-2细胞体外增殖及体内成瘤的影响并讨论其机制.方法 通过实时荧光定量聚合酶链反应(FQ-PCR)检测前列腺癌组织和癌旁组织中CtBP2mRNA表达量;将靶向CtBP2基因短发卡RNA(shRNA)质粒及阴性对照(NC)质粒分别转入C4-2细胞,FQ-PCR和Western blot法分别检测CtBP2 mRNA及蛋白的表达水平;细胞计数试剂盒(CCK-8)法和流式细胞仪分别检测下调CtBP2表达对C4-2细胞体外增殖能力和凋亡的影响;裸鼠皮下移植瘤实验观察下调CtBP2对成瘤的影响;通过Western blot法检测下调CtBP2对c-myc和HSPC 111蛋白表达的影响.结果 CtBP2 mRNA在前列腺癌组织中的表达水平显著高于癌旁组织(4.08±1.25比1.0 ±0.46)(P＜0.05);与NC组细胞比较,CtBP2-shRNA组细胞CtBP2的mRNA(0.21 ±0.03比0.97 ±0.09)和蛋白(0.45±0.02比1.02±0.07)表达量均显著降低(P＜0.05);CCK-8检测结果显示,在24、48、72 h CtBP2-shRNA组细胞450nm处吸光度值分别为0.45±0.05,1.37±0.09和1.64±0.07,而NC组细胞为0.78±0.08,2.35±0.18和2.99±0.28,下调CtBP2后细胞增殖水平分别降至NC组细胞的(60.1±13.3)％、(59.4±6.3)％和(56.0±6.6)％;通过流式细胞检测发现下调CtBP2可显著促进C4-2细胞的凋亡,凋亡率为(32.57±1.98)％ (P ＜0.05),而空白对照组和NC组细胞凋亡率分别为(15.63±0.38)％和(15.87±0.99)％;体内实验显示下调CtBP2表达显著抑制裸鼠皮下移植瘤的生长;通过Western blot发现下调CtBP2可显著降低C4-2细胞中c-myc(0.55 ±0.03比0.96±0.02)及其下游靶蛋白HSPC111(0.56 ±0.02比1.04 ±0.10)的表达量(P＜0.05).结论 CtBP2可通过c-myc-HSPC111通路参与调控前列腺癌C4-2细胞的增殖能力及体内成瘤能力.
目的 觀察下調CtBP2對人前列腺癌C4-2細胞體外增殖及體內成瘤的影響併討論其機製.方法 通過實時熒光定量聚閤酶鏈反應(FQ-PCR)檢測前列腺癌組織和癌徬組織中CtBP2mRNA錶達量;將靶嚮CtBP2基因短髮卡RNA(shRNA)質粒及陰性對照(NC)質粒分彆轉入C4-2細胞,FQ-PCR和Western blot法分彆檢測CtBP2 mRNA及蛋白的錶達水平;細胞計數試劑盒(CCK-8)法和流式細胞儀分彆檢測下調CtBP2錶達對C4-2細胞體外增殖能力和凋亡的影響;裸鼠皮下移植瘤實驗觀察下調CtBP2對成瘤的影響;通過Western blot法檢測下調CtBP2對c-myc和HSPC 111蛋白錶達的影響.結果 CtBP2 mRNA在前列腺癌組織中的錶達水平顯著高于癌徬組織(4.08±1.25比1.0 ±0.46)(P＜0.05);與NC組細胞比較,CtBP2-shRNA組細胞CtBP2的mRNA(0.21 ±0.03比0.97 ±0.09)和蛋白(0.45±0.02比1.02±0.07)錶達量均顯著降低(P＜0.05);CCK-8檢測結果顯示,在24、48、72 h CtBP2-shRNA組細胞450nm處吸光度值分彆為0.45±0.05,1.37±0.09和1.64±0.07,而NC組細胞為0.78±0.08,2.35±0.18和2.99±0.28,下調CtBP2後細胞增殖水平分彆降至NC組細胞的(60.1±13.3)％、(59.4±6.3)％和(56.0±6.6)％;通過流式細胞檢測髮現下調CtBP2可顯著促進C4-2細胞的凋亡,凋亡率為(32.57±1.98)％ (P ＜0.05),而空白對照組和NC組細胞凋亡率分彆為(15.63±0.38)％和(15.87±0.99)％;體內實驗顯示下調CtBP2錶達顯著抑製裸鼠皮下移植瘤的生長;通過Western blot髮現下調CtBP2可顯著降低C4-2細胞中c-myc(0.55 ±0.03比0.96±0.02)及其下遊靶蛋白HSPC111(0.56 ±0.02比1.04 ±0.10)的錶達量(P＜0.05).結論 CtBP2可通過c-myc-HSPC111通路參與調控前列腺癌C4-2細胞的增殖能力及體內成瘤能力.
목적 관찰하조CtBP2대인전렬선암C4-2세포체외증식급체내성류적영향병토론기궤제.방법 통과실시형광정량취합매련반응(FQ-PCR)검측전렬선암조직화암방조직중CtBP2mRNA표체량;장파향CtBP2기인단발잡RNA(shRNA)질립급음성대조(NC)질립분별전입C4-2세포,FQ-PCR화Western blot법분별검측CtBP2 mRNA급단백적표체수평;세포계수시제합(CCK-8)법화류식세포의분별검측하조CtBP2표체대C4-2세포체외증식능력화조망적영향;라서피하이식류실험관찰하조CtBP2대성류적영향;통과Western blot법검측하조CtBP2대c-myc화HSPC 111단백표체적영향.결과 CtBP2 mRNA재전렬선암조직중적표체수평현저고우암방조직(4.08±1.25비1.0 ±0.46)(P＜0.05);여NC조세포비교,CtBP2-shRNA조세포CtBP2적mRNA(0.21 ±0.03비0.97 ±0.09)화단백(0.45±0.02비1.02±0.07)표체량균현저강저(P＜0.05);CCK-8검측결과현시,재24、48、72 h CtBP2-shRNA조세포450nm처흡광도치분별위0.45±0.05,1.37±0.09화1.64±0.07,이NC조세포위0.78±0.08,2.35±0.18화2.99±0.28,하조CtBP2후세포증식수평분별강지NC조세포적(60.1±13.3)％、(59.4±6.3)％화(56.0±6.6)％;통과류식세포검측발현하조CtBP2가현저촉진C4-2세포적조망,조망솔위(32.57±1.98)％ (P ＜0.05),이공백대조조화NC조세포조망솔분별위(15.63±0.38)％화(15.87±0.99)％;체내실험현시하조CtBP2표체현저억제라서피하이식류적생장;통과Western blot발현하조CtBP2가현저강저C4-2세포중c-myc(0.55 ±0.03비0.96±0.02)급기하유파단백HSPC111(0.56 ±0.02비1.04 ±0.10)적표체량(P＜0.05).결론 CtBP2가통과c-myc-HSPC111통로삼여조공전렬선암C4-2세포적증식능력급체내성류능력.
Objective To investigate the effect of C-terminal-binding protein 2 (CtBP2) downregulation on the proliferation and tumor growth of human prostate cancer cell line C4-2.Methods The expression of CtBP2 mRNA in 60 prostate cancer tissues and 60 adjacent normal prostate tissues were assessed by real-time florescent quantitative polymerase chain reaction (FQ-PCR).Human prostate cancer C4-2 cells were infected with lentivirus carrying short hairpin RNA (shRNA) targeting CtBP2 expression or the negative control plasmids.The expression of CtBP2 mRNA and protein were assessed by FQ-PCR and western blotting,respectively.The proliferation and apoptosis rate of C4-2 cells was detected by cell counting kit-8 (CCK-8) assay and flow cytometry.The effects of CtBP2 downregulation on tumor growth in vivo was determined in nude mice.The effects of CtBP2 downregulation on the expression of c-myc and HSPC111 were assessed by western blotting.Results The expression level of CtBP2 mRNA in prostate cancer tissues was significantly higher than in the adjacent normal prostate tissues (4.08 ± 1.25 vs.1.00 ± 0.46,P ＜ 0.05).Stable shRNA CtBP2 transfection significantly down-regulated the expression of mRNA (0.21 ± 0.03 vs.0.97 ± 0.09) and protein (0.45 ± 0.02 vs.1.02 ± 0.07) levels of CtBP2 when compared with NC cells (P ＜ 0.05).The absorbance at 450 nm of CtBP2-shRNA cells at 24,48,and 72 h was 0.45 ±0.05,1.37 ± 0.09,and 1.64 ± 0.07,and that in NC cells was 0.78 ± 0.08,2.35 ± 0.18,and 2.99 ± 0.28 respectively.The cell proliferation was decreased by (60.1 ± 13.3)％,(59.4±6.3)％,and (56.0±6.6)％ at 24,48,and 72h respectively in CtBP2-shRNA cells as compared with NC cells (P ＜ 0.05).Tumor growth in nude mice was also significantly inhibited by CtBP2 knockdown (P ＜ 0.05).The apoptosis rate of CtBP2-shRNA cells was significantly increased when compared that in NC cells [(32.57 ± 1.98) ％ vs.(15.63 ± 0.38) ％] (P ＜ 0.05).The expression levels of c-myc (0.55 ± 0.03 vs.0.96 ± 0.02) and it's downstream target protein HSPC111 (0.56 ± 0.02 vs.1.04 ± 0.10) decreased when CtBP2 was knockdown (P ＜ 0.05).Conclusion CtBP2 participates in the modulation of prostate cancer cell proliferation and tumor growth through c-myc-HSPC111 pathway.
