活性氧抑制剂增强胰腺癌细胞对吉西他滨敏感性的作用机制研究

Mechanism of reactive oxygen species inhibitor in enhancing the sensitivity of pancreatic cancer cell lines to gemcitabine

目的探讨活性氧抑制剂在胰腺癌细胞中促进吉西他滨敏感性的作用机制。方法胰腺癌细胞株Miapaca-2分为对照组、吉西他滨组、N-乙酰半胱氨酸(N-acetyl-L-cysteine,NAC)组和吉西他滨+NAC组,分别采用PBS、吉西他滨、NAC和吉西他滨+NAC进行处理。处理48h后,采用流式细胞仪检测4组细胞中活性氧表达情况及细胞凋亡率,采用显微镜观察4组药物处理后存活细胞数,采用实时定量PCR法检测4组细胞中p21、白细胞介素(interleukin,IL)-8mRNA表达情况。结果处理48h后,吉西他滨组活性氧阳性表达率[(67.32±3.75)%]、p21mRNA(52.26±1.53)和IL-8mRNA表达量(27.05±0.74)明显高于对照组[(25.32±2.23)%、0.95±0.08、0.90±0.10]、NAC组[(36.73±5.42)%、5.26±0.89、3.23±0.97]和吉西他滨+NAC组[(45.32±7.23)%、31.24±3.24、7.33±0.56](P<0.05),吉西他滨+NAC组高于对照组(P<0.05);吉西他滨+NAC组存活细胞数[(1.06±0.07)×104个/mL]明显少于对照组[(4.23±0.40)×104个/mL]、吉西他滨组[(1.97±0.03)×104个/mL]和NAC组[(4.17±0.56)×104个/mL](P<0.05),吉西他滨组少于对照组和NAC组(P<0.05),对照组与NAC组比较差异无统计学意义(P>0.05);吉西他滨+NAC组细胞凋亡率[(49.23±3.07)%]明显高于对照组[(9.63±3.37)%]、吉西他滨组[(25.32±5.15)%]和NAC组[(15.57±5.76)%](P<0.05),吉西他滨组高于对照组(P<0.05),对照组、吉西他滨组与NAC组比较差异无统计学意义(P>0.05)。结论活性氧抑制剂NAC可提高吉西他滨对胰腺癌细胞株的敏感性,可能与活性氧抑制剂能明显促胰腺癌细胞凋亡及抑制细胞中IL-8、p21基因表达有关。

Objective To explore the mechanism of reactive oxygen species(ROS)inhibitor,in enhancing the sensitivity of pancreatic cancer cell lines to gemcitabine.Methods Pancreatic cancer cell lines Miapaca-2 were divided into control group,gemcitabine group,N-acetyl-L-cysteine(NAC)group and gemcitabine+NAC group.After 48-h treatment,the ROS expression and cell apoptosis were detected by flow cytometry,the survived cell numbers were observed by microscopy,and the expressions of p21 and interleukin-8(IL-8)mRNA were detected by real-time quantitive PCR in four groups.Results After 48-h treatment,the expressions of ROS,p21 mRNA and IL-8 mRNA were significantly higher in gemcitabine group((67.32±3.75)%,52.26±1.53,27.05±0.74)than those in control group((25.32±2.23)%,0.95±0.08,0.90±0.10),NAC group((36.73±5.42)%,5.26±0.89,3.23±0.97)and gemcitabine+NAC group((45.32±7.23)%,31.24±3.24,7.33±0.56)(P<0.05),and in gemcitabine+NAC group than those in control group(P<0.05).The survived cell was significantly fewer in gemcitabine+NAC group((1.06±0.07)×104 cell/mL)than that in control group((4.23±0.40)×104 cell/mL),gemcitabine group((1.97±0.03)×104 cell/mL)and NAC group((4.17±0.56)×104 cell/mL)(P<0.05),was fewer in gemcitabine group than that in control group and NAC group(P<0.05),and showed no significant difference between control group and NAC group(P>0.05).The cell apoptosis rate was significantly higher in gemcitabine+ NAC group((49.23±3.07)%)than that in control group((9.63±3.37)%),gemcitabine group((25.32±5.15)%)and NAC group((15.57±5.76)%)(P<0.05),was higher in gemcitabine group than that in control group(P<0.05),and showed no significant difference in control group and gemcitabine group in comparison with NAC group(P>0.05).Conclusion ROS inhibitor can enhance the sensitivity of pancreatic cancer cell lines to gemcitabine,probably because NAC can significantly induce the cell apoptosis and inhibit p21 and IL-8 expressions in human pancreatic cancer cell lines.