摘要
目的:三溴丙酮酸(3-bromopyruvate,3-BrPA)作为对糖酵解途径具有抑制作用的一种新型的潜在抗癌药物,在癌症治疗领域得到了广泛关注,但3-BrPA在喉癌中的作用尚不清楚。本研究基于磷脂酰肌醇3-激酶(phosphoinositide 3-kinase,PI3K)/蛋白激酶(protein kinases B,Akt)/哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,mTOR)信号通路探讨3-BrPA对人Hep2喉癌细胞增殖、迁移、侵袭的影响以及可能的作用机制。方法:体外培养人Hep2喉癌细胞,并将其分为对照组、3-BrPA(20、40、60、80、100、120μmol/L)组,分别作用24、48、72 h,观察3-BrPA对人Hep2喉癌细胞增殖的影响,确定高、中、低浓度和作用时间进行后续实验。划痕、Transwell实验分别观察3-BrPA对人Hep2喉癌细胞迁移和侵袭的影响。利用STRING数据库分析己糖激酶-Ⅱ(hexokinase-Ⅱ,HK-2)与PI3K、Akt、m TOR蛋白之间的相关性。采用real-time RT-PCR和蛋白质印迹法检测HK-2、PI3K、Akt和mTOR的mRNA和蛋白的表达情况。结果:细胞计数试剂盒8(cell counting-kit 8,CCK-8)检测结果表明:与对照组相比,40~120μmol/L 3-BrPA作用24、48、72 h后人Hep2喉癌细胞增殖活力明显下降,差异均具有统计学意义(均P<0.05)。选择95、115、135μmol/L作为3-BrPA低、中、高浓度组作用24 h进行后续实验,CCK-8检测结果发现:随着药物浓度的增加,3-BrPA低、中、高浓度组的人Hep2喉癌细胞增殖活力、迁移率和侵袭数量逐渐下降,与对照组相比,差异均具有统计学意义(均P<0.01)。STRING数据库分析表明:人Hep2喉癌细胞中HK-2与Akt、mTOR蛋白相关性较高。Real-time RT-PCR和蛋白质印迹法结果显示:与对照组相比,3-BrPA低、中、高浓度组HK-2、PI3K、Akt、mTOR mRNA和蛋白表达均降低,差异均有统计学意义(均P<0.05)。结论:3-BrPA能够抑制人Hep2喉癌细胞增殖、迁移和侵袭,这与下调HK-2的表达,抑制PI3K/Akt/mTOR信号通路有关。
Objective:3-Bromopyruvate(3-BrPA)is a novel potential anticancer agent that inhibits glycolytic and has gained considerable attention in cancer treatment.However,its effects in laryngeal cancer remain unclear.This study investigates the impact of 3-BrPA on the proliferation,migration,and invasion of human Hep2 laryngeal cancer cells and explores its potential mechanisms via the phosphoinositide 3-kinase(PI3K)/protein kinase(Akt)/mammalian rapacity target protein(mTOR)signaling pathway.Methods:Human Hep2 laryngeal cancer cells were cultured in vitro and divided into a control group and 3-BrPA(20,40,60,80,100,120μmol/L)treatment groups for 24,48,and 72 hours to observe the effects on cell proliferation.Optimal concentrations and exposure times were determined for further experiments.Scratch and Transwell assays were used to assess cell migration and invasion.The STRING database was used to analyze correlations between hexokinase-II(HK-2)and PI3K,Akt,and mTOR proteins.Real-time RT-PCR and Western blotting analyses were performed to measure mRNA and protein expression of HK-2,PI3K,Akt,and mTOR.Results:Cell counting-kit 8(CCK-8)assays indicated a significant decrease in proliferation of Hep2 cells exposed to 40−120μmol/L 3-BrPA at 24,48,and 72 hours compared to the control(all P<0.05).Concentrations of 95,115,and 135μmol/L were selected as low,medium,and high for subsequent 24-hour treatment experiments.CCK-8 results revealed that as 3-BrPA concentration increased,Hep2 cell proliferation,migration rate,and invasion number significantly decreased compared to the control group(all P<0.01).STRING analysis indicated a strong correlation between HK-2 and Akt/mTOR proteins in Hep2 cells.Real-time RT-PCR and Western blotting results demonstrated that HK-2,PI3K,Akt,and mTOR mRNA and protein expression significantly decreased in the low,medium,and high 3-BrPA treatment groups compared to the control(all P<0.05).Conclusion:3-BrPA inhibits the proliferation,migration,and invasion of Hep2 laryngeal cancer cells,potentially through down-regulation of the PI3K/Akt/mTOR signaling pathway.
作者
赵静
王磊
刘喆
张立
季洁
李亚玲
舍雅莉
ZHAO Jing;WANG Lei;LIU Zhe;ZHANG Li;JI Jie;LI Yaling;SHE Yali(First School of Clinical Medical,Gansu University of Chinese Medicine,Lanzhou 730030;Basic Medicine College,Gansu University of Chinese Medicine,Lanzhou 730030;Department of Otolaryngology,First People’s Hospital of Lanzhou City,Lanzhou 730050,China)
出处
《临床与病理杂志》
CAS
2024年第8期1059-1068,共10页
Journal of Clinical and Pathological Research
基金
甘肃省自然科学基金(21JR11RA141)。
