摘要
目的探讨阿司匹林(ASA)是否增强顺铂(DDP)对食管鳞癌(ESCC)的治疗作用及其相关机制。方法培养ESCC细胞,实验分为空白组(DMSO组)、实验组(ASA组,DDP组,ASA+DDP组)。CCK8检测ASA和DDP单用或联用对ESCC细胞(Eca-109、Kyse30、TE-10)活性的影响,Annexin-V-FITC/PI检测ASA和DDP单用或联用对ESCC细胞凋亡的影响;转染PHY-LVOE1.6-SH2B1上调Eca-109细胞SH2B1的表达水平,CCK8及细胞划痕实验检测过表达SH2B1对Eca-109细胞活性及迁移的影响,免疫荧光及Western blot检测Eca-109细胞内SH2B1、β-catenin及其相关通路关键分子(GSK-3β、CyclinD1、TCF-4)蛋白变化。结果给予5mmol/L ASA处理后,与空白组相比Eca-109、Kyse30、TE-10细胞活性及凋亡率均无明显差异(Ps>0.05);与DDP5组相比,ASA5+DDP5组3株细胞活性最低(t=3.49,P<0.05;t=2.80,P<0.05;t=3.10,P<0.05)、凋亡率最高(t=11.44,P<0.05;t=11.68,P<0.05;t=12.24,P<0.05),差异具有统计学差异。DDP(5μmol/L)联合梯度ASA(0、5、10mmol/L)处理Eca-109细胞,SH2B1的表达量随ASA浓度上升而下降(F=54.0,P<0.05)。与SH2B1组相比,ASA(10mmol/L)和DDP(5μmol/L)联合处理后,Eca-109细胞SH2B1(t=2.62,P<0.05)、GSK-3β(t=0.96,P<0.05)、CyclinD1(t=1.25,P<0.05)、TCF-4(t=1.98,P<0.05)蛋白表达下降;与Vector组相比,过表达SH2B1,Eca-109细胞活性及迁移能力增强(Ps<0.05),β-catenin表达量及入核增加(t=2.52,P<0.05),GSK-3β(t=2.50,P<0.05)、CyclinD1(t=1.49,P<0.05)、TCF-4(t=2.62,P<0.05)表达增加;可部分逆转ASA和DDP联用处理所带来的上述蛋白抑制作用。结论ASA通过抑制SH2B1/β-catenin信号通路在一定程度上增强ESCC细胞对DDP的化疗敏感性,协同DDP可以抑制细胞活性及促进细胞凋亡。
Objective To investigate the adjuvant chemotherapy role of aspirin(ASA)whether potentiates the therapeutic effect of cisplatin(DDP)on esophageal squamous cell carcinoma(ESCC)and explore its mechanisms.Methods Esophageal squamous cell carcinoma cell lines(Eca-109,Kyse30,TE-10)were treated with 5μmol/L cisplatin,or/and plus aspirin at a concentration of 0,5,10mmol/L.Divide the cultured ESCC cells into blank group(DMSO group),experimental groups(ASA group,DDP group,ASA+DDP group,DDP group).Aspirin and cisplatin and their synergistic effects were assessed by measuring cell viability,death,protein and mRNA expression.CCK8 assay and apoptosis assay were employed to analyze the effect of aspirin to potentiate cisplatin treatment in esophageal squamous cell carcinoma(ESCC).Transfection of PHY-LVOE1.6-SH2B1 upregulated the expression of SH2B1 in Eca-109 cells,CCK8 and cell scratch assay were used to detect the effects of overexpressed SH2B1 on the viability and migration of Eca-109 cells,and the protein of SH2B1,β-catenin and its key components(GSK-3β,CyclinD1 and TCF-4)were detected by Immunofluorescence and Western blot.Results Compared with DDP alone,the Eca-109,Kyse30,TE-10 cell viability of ASA+DDP group was the lowest(t=3.49,P<0.05,t=2.80,P<0.05,t=3.10,P<0.05)and the apoptosis rate was the highest(t=11.44,P<0.05,t=11.68,P<0.05,t=12.24,P<0.05).Eca-109 cells were treated with DDP(5μmol/L)and gradient ASA(0,5,10mmol/L),the expression of SH2B1(F=54.0,P<0.05)was negatively correlated with the concentration of ASA.In Eca-109 cells treated with ASA(10mmol/L)and DDP(5μmol/L),the protein expression of SH2B3(t=2.62,P<0.05),GSK-3β(t=0.96,P<0.05),CyclinD1(t=1.25,P<0.05),TCF-4(t=1.98,P<0.05)were decreased.The activity and migration ability of Eca-109 cells overexpression of SH2B1 were enhanced(Ps<0.05)and the expression ofβ-catenin(t=2.52,P<0.05),GSK-3β(t=2.50,P<0.05),CyclinD1(t=1.49,P<0.05),TCF-4(t=2.62,P<0.05)was increased.Overexpression of SH2B1 can partially reverse the inhibitory effect of the above proteins induced by the combination of ASA and DDP.Mechanistically,aspirin reversed cisplatin resistance through the SH2B1/β-catenin pathway.Conclusion Aspirin potentiated the sensitivity of cisplatin and inhibits the growth of esophageal squamous cell carcinoma by inhibiting SH2B1/β-catenin pathway at the cellular level.
作者
张黎明
王少强
王丽娜
马鸣
魏海翔
ZHANG Liming;WANG Shaoqiang;WANG Lina;MA Ming;WEI Haixiang(School of Clinical Medicine,Jining Medical University,Jining 272013,China;Department of Thoracic Surgery,Weifang People′s Hospital,Weifang 261041,China;Medical Research Center,Affilited Hospital of Jining Medical University,Jining 272029,China;Department of Thoracic Surgery,Affiliated Hospital of Jining Medical University,Jining 272029,China)
出处
《济宁医学院学报》
2023年第3期153-158,共6页
Journal of Jining Medical University
基金
国家自然科学基金(81802290)
济宁医学附属医院“苗圃”科研计划(MP-ZD-2020-002)。
