摘要
[目的]研究肌醇多聚磷酸-4-磷酸酶Ⅱ型(inositol polyphosphate 4-phosphatase typeⅡ,INPP4B)的表达对结肠癌预后及氟尿嘧啶(5-fluorouracil,5-FU)药物敏感性的影响及其机制。[方法]通过免疫组织化学方法和R2在线工具分析INPP4B在结肠癌组织的表达及其与预后的关系。构建敲减或过表达INPP4B质粒,分别转染结肠癌细胞株WiDr细胞和SW480细胞,通过实时荧光定量PCR技术检测INPP4B mRNA的表达情况。通过Western blot技术检测细胞中INPP4B、磷酸化的Akt(pAkt)和p27的表达情况。使用CCK-8检测细胞的增殖能力。采取流式细胞术检测细胞的凋亡率和细胞周期的分布。[结果]INPP4B高表达与结肠癌患者预后不良相关(P<0.05)。敲减INPP4B后,5-FU处理细胞24 h,发现与WiDr shControl细胞相比,Wi Dr shINPP4B细胞存活率显著降低(P=0.011),凋亡率显著增高(P<0.001),G_(0)/G_(1)期细胞比例显著升高(P<0.001)。过表达INPP4B后,与SW480空载细胞相比,SW480 INPP4B细胞存活率显著升高(P=0.002),凋亡率显著降低(P=0.002),G_(0)/G_(1)期的细胞比例显著降低(P=0.001)。5-FU处理结肠癌细胞后,与WiDr shControl细胞相比,WiDr shINPP4B细胞中pAkt(Ser473)的水平显著降低(P=0.003),pAkt(Thr308)的水平显著降低(P<0.001),p27蛋白的表达显著升高(P<0.001);与SW480空载细胞相比,SW480INPP4B细胞中pAkt(Ser473)的水平显著升高(P<0.001),pAkt(Thr308)的水平显著升高(P<0.001),p27蛋白的表达显著降低(P<0.001)。[结论]INPP4B高表达与结肠癌患者不良预后相关,并通过激活Akt/p27信号轴降低结肠癌细胞对5-FU的敏感性。
[Purpose]To investigate the relationship of inositol polyphosphate 4-phosphatase typeⅡ(INPP4B)with prognosis of colon cancer and its effect on chemosensitivity of colon cancer cells.[Methods]The correlation between expression of INPP4B and prognosis of colon cancer patients was analyzed with bioinformatics methods.Lentivirus-mediated knock-down or overexpressing INPP4B cell lines were constructed with human colon cancer WiDr or SW480 cells.The expression of NPP4B mRNA and protein was detected by real-time quantitative PCR(qPCR)and Western blotting,respectively;the phosphorylated Akt(pAkt)and p27 were detected by Western blotting;cell proliferation ability was measured with cytotoxicity kit-8(CCK-8);the apoptotic rate and cell cycle distribution were determined with flow cytometry.[Results]Kaplan-Meier survival analysis showed that the high expression of INPP4B was correlated with poor prognosis of colon cancer patients(P<0.05).Compared to control cells,INPP4B knockdown increased the sensitivity of colon cancer cells to 5-FU significantly(P=0.011),enhancing 5-FU-induced cell apoptosis(P<0.001)and increasing 5-FU-induced G_(1) arrest(P<0.001)of colon cancer cells.Compared to vector control,over-expression of INPP4B decreased the sensitivity of colon cancer cells to 5-FU significantly(P=0.002),attenuated 5-FU-induced cell apoptosis(P=0.002)and decreasing 5-FU-induced G_(1) arrest(P=0.001)of colon cancer cells.Knockdown of INPP4B in colon cancer cells significantly decreased p-AKT protein expression(P=0.003)and enhanced p27 protein expression(P<0.001)compared to control cells.Overexpression of INPP4B in colon cancer cells increased p-AKT protein expression(P<0.001)and reduced p27 protein expression(P<0.001).[Conclusion]INPP4B is correlated with poor prognosis of colon cancer patients and it can decrease the sensitivity of colon cancer cells to 5-FU via AKT/p27 signal pathway.
作者
孙奋琪
曹小梅
邹佳杰
王福花
杨瑞红
郭素堂
SUN Fen-qi;CAO Xiao-mei;ZOU Jia-jie;WANG Fu-hua;YANG Rui-hong;GUO Su-tang(Basic Medical College,Shanxi Medical University,Taiyuan 030001,China;Shanxi Cancer Hospital,Taiyuan 030013,China)
出处
《中国肿瘤》
CAS
CSCD
北大核心
2022年第4期311-320,共10页
China Cancer
基金
国家自然科学基金(81972840)。
