[1]陈利娟,穆晓倩,刘　杰,等.PTEN通过磷脂酰肌醇-3-激酶/蛋白激酶B信号通路对非小细胞肺癌顺铂敏感性的影响[J].新乡医学院学报,2020,37(6):501-508.[doi:10.7683/xxyxyxb.2020.06.001]
　CHEN Lijuan,MU Xiaoqian,LIU Jie,et al.Effect of PTEN on the sensitivity of non-small cell lung cancer to cisplatin through the phosphatidylinositol-3-kinase/protein kinase B signaling pathway[J].Journal of Xinxiang Medical University,2020,37(6):501-508.[doi:10.7683/xxyxyxb.2020.06.001]
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PTEN通过磷脂酰肌醇-3-激酶/蛋白激酶B信号通路对非小细胞肺癌顺铂敏感性的影响
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《新乡医学院学报》[ISSN:1004-7239/CN:41-1186/R]

卷:
37
期数:
2020年6
页码:
501-508
栏目:
基础研究
出版日期:
2020-06-05

文章信息/Info

Title:
Effect of PTEN on the sensitivity of non-small cell lung cancer to cisplatin through the phosphatidylinositol-3-kinase/protein kinase B signaling pathway
作者:
陈利娟1; 穆晓倩1; 刘　杰1; 张体鹏2; 赵艳秋1
（1.郑州大学附属肿瘤医院肿瘤内科，河南 　郑州　450008；2.郑州大学附属郑州中心医院心血管内科，河南　郑州
450052）
Author(s):
CHEN Lijuan1; MU Xiaoqian1; LIU Jie1; ZHANG Tipeng2; ZHAO Yanqiu1
(1.Department of Oncology,Cancer Hospital Affiliated to Zhengzhou University,Zhengzhou 450008,Henan Province,China;2.Department of Cardiology,Zhengzhou Central Hospital Affiliated to Zhengzhou University,Zhengzhou 450052,Henan Province,China)
关键词:
非小细胞肺癌; PTEN; 磷脂酰肌醇-3-激酶/蛋白激酶B信号通路; 顺铂; 化学治疗敏感性
Keywords:
non-small-cell lung carcinoma; PTEN; phosphatidylinositol-3-kinase/protein kinase B signaling pathway; cisplatin; chemosensitivity
分类号:
R734.2
DOI:
10.7683/xxyxyxb.2020.06.001
文献标志码:
A
摘要:
目的　探讨PTEN通过磷脂酰肌醇-3-激酶（PI3K）/蛋白激酶B（AKT）信号通路对非小细胞肺癌（NSCLC）顺铂敏感性的影响。方法　选择2016年1月至2018年1月于郑州大学附属肿瘤医院手术切除的60例NSCLC患者的癌组织标本及对应的癌旁组织（距肿瘤边缘＞5 cm）标本，采用免疫组织化学染色检测癌组织和癌旁组织中PTEN蛋白表达。培养NSCLC细胞株A549，取对数生长期细胞并随机分为blank组（不作任何处理）、pcDNA-PTEN组（转染PTEN过表达质粒）、pcDNA-PTEN对照组（转染PTEN过表达对照质粒）、siRNA-PTEN组（转染siRNA-PTEN质粒）、siRNA-PTEN对照组（转染siRNA-PTEN对照质粒）、wortmannin组（转染PI3K/AKT信号通路抑制剂）、胰岛素样生长因子-1（IGF-1）组（转染PI3K/AKT信号通路激动剂）、pcDNA-PTEN+IGF-1组（共转染pcDNA-PTEN过表达质粒和PI3K/AKT信号通路激动剂），采用Lipofectamine 2000介导细胞转染，转染6 h后更换为完全培养基，37 ℃培养48 h后收集细胞；采用定量反转录聚合酶链反应检测A549细胞中PTEN、PI3K及AKT mRNA表达，Western blot法检测A549细胞中PTEN、PI3K及AKT蛋白表达，Transwell实验检测A549细胞侵袭能力，划痕实验检测A549细胞迁移能力。取各组转染后培养48 h细胞，分别加入0.00、0.75、1.50、3.00、6.00、12.00 μmol·L-1顺铂，培养48 h后应用细胞计数试剂盒-8检测A549细胞对顺铂的敏感性。结果　NSCLC患者癌组织中PTEN蛋白阳性表达率显著低于癌旁组织，癌组织中PI3K、AKT蛋白阳性表达率显著高于癌旁组织（P＜0.05）。blank组、pcDNA-PTEN对照组和siRNA-PTEN对照组A549细胞中PTEN、PI3K、AKT 蛋白及mRNA表达比较差异无统计学意义（P＞0.05）；与pcDNA-PTEN对照组比较，pcDNA-PTEN组A549细胞中PTEN蛋白及mRNA表达增加，PI3K、AKT蛋白及mRNA表达减少（P＜0.05）；与siRNA-PTEN对照组比较，siRNA-PTEN组A549细胞中PTEN蛋白及mRNA表达减少，PI3K、AKT蛋白及mRNA表达增加（P＜0.05）；与pcDNA-PTEN组比较，pcDNA-PTEN+IGF-1组A549细胞中PTEN蛋白及mRNA表达减少，PI3K、AKT蛋白及mRNA表达增加（P＜0.05）；与blank组比较，wortmannin组A549细胞中PTEN蛋白和mRNA表达增加，PI3K、AKT蛋白及mRNA表达减少（P＜0.05）；与blank组和wortmannin组比较，IGF-1组A549细胞中PTEN蛋白和mRNA表达减少，PI3K、AKT蛋白及mRNA表达增加（P＜0.05）。blank组、pcDNA-PTEN对照组和siRNA-PTEN对照组A549细胞侵袭和迁移能力比较差异无统计学意义（P＞0.05）；pcDNA-PTEN组A549细胞侵袭和迁移能力显著低于pcDNA-PTEN对照组（P＜0.05），siRNA-PTEN组A549细胞侵袭和迁移能力显著高于siRNA-PTEN对照组（P＜0.05），IGF-1组A549细胞侵袭和迁移能力显著高于blank组和wortmannin组（P＜0.05），wortmannin组A549细胞侵袭和迁移能力显著低于blank组（P＜0.05），pcDNA-PTEN+ IGF-1组A549细胞侵袭和迁移能力显著高于pcDNA-PTEN组（P＜0.05）。随着顺铂浓度增加，各组A549细胞存活率逐渐降低（P＜0.05）。各浓度顺铂干预下，blank组、pcDNA-PTEN对照组和siRNA-PTEN对照组A549细胞存活率比较差异无统计学意义（P＞0.05），pcDNA-PTEN组A549细胞存活率显著低于pcDNA-PTEN对照组（P＜0.05），siRNA-PTEN组A549细胞存活率显著高于siRNA-PTEN对照组（P＜0.05），wortmannin组A549细胞存活率显著低于blank组（P＜0.05），IGF-1组A549细胞存活率显著高于wortmannin组和blank组（P＜0.05），pcDNA-PTEN+IGF-1组A549细胞存活率显著高于pcDNA-PTEN组（P＜0.05）。结论　NSCLC组织中PTEN蛋白表达降低，上调PTEN基因表达可能通过抑制PI3K/AKT信号通路活性而降低细胞侵袭和迁移能力，提高NSCLC细胞对顺铂的敏感性。
Abstract:
Objective　To investigate the effect of PTEN on the sensitivity of non-small cell lung cancer(NSCLC) to cisplatin through the phosphatidylinositol-3-kinase/protein kinase B(PI3K/AKT) signaling pathway.Methods　The cancer tissue and paracancerous tissue specimens (more than 5 cm from tumor margin) of 60 patients with NSCLC who underwent surgical resection from January 2016 to January 2018 in the Cancer Hospital Affiliated to Zhengzhou University were selected,and the expression of PTEN protein in NSCLC and paracancerous tissues was detected by immunohistochemistry.The NSCLC cells A549 were cultured,and the cells in logarithmic growth phase were obtained and they were randomly divided into blank group (without any treatment),pcDNA-PTEN group (transfected with PTEN overexpression plasmid),pcDNA-PTEN control group (transfected with PTEN overexpression control plasmid),siRNA-PTEN group (transfected with siRNA-PTEN plasmid),siRNA-PTEN control group (transfected with siRNA-PTEN control plasmid),wortmannin group (transfected with PI3K/Akt signal pathway inhibitor),insulin-like growth factor-1 (IGF-1) group (transfected with PI3K/Akt signal pathway activator),pcDNA-PTEN+IGF-1 group (co transfected with pcDNA-PTEN overexpression plasmid and PI3K/Akt signal pathway activator).The cells were transfected by Lipofectamine 2000.After 6 hours of transfection,the cells were cultured in complete medium at 37 ℃ for 48 hours.The expression of PTEN,PI3K and AKT mRNA in A549 cells was detected by quantitative reverse transcription polymerase chain reaction.The expression of PTEN,PI3K and AKT protein in A549 cells was detected by Western blot method.The invasion ability of A549 cells was detected by Transwell test,and the migration ability of A549 cells was detected by scratch test.The cells in each group were cultured for 48 hours after transfection,0.00,0.75,1.50,3.00,6.00,12.00 μmol·L-1 cisplatin for 48 hours respectively,then the sensitivity of A549 cells to cisplatin was detected by cell count kit-8.Results　Compared with the paracancerous tissues,the positive expression rate of PTEN protein was lower,and that of PI3K and AKT protein was higher in NSCLC tissues (P＜0.05).There was no significant difference in the expression of PTEN,PI3K,AKT protein and mRNA among the blank group,pcDNA-PTEN control group and siRNA-PTEN control group (P＞0.05).Compared with the pcDNA-PTEN control group,the expression of PTEN protein and mRNA in A549 cells was higher,while the expression of PI3K,AKT protein and mRNA in A549 cells was lower in the pcDNA-PTEN group (P＜0.05).Compared with the siRNA-PTEN control group,the expression of PTEN protein and mRNA in A549 cells was lower,while the expression PI3K,AKT protein and mRNA in A549 cells was higher in the siRNA-PTEN group (P＜0.05).Compared with the pcDNA-PTEN group,the expression of PTEN protein and mRNA in A549 cells was lower,and the expression of PI3K,AKT protein and mRNA in A549 cells was higher in the pcDNA-PTEN+IGF-1 group (P＜0.05).Compared with the blank group,the expression of PTEN protein and mRNA in A549 cells was higher,and the expression of PI3K,AKT protein and mRNA in A549 cells was lower in the wortmannin group (P＜0.05).Compared with the blank group and wortmannin group,the expression of PTEN protein and mRNA in A549 cells was lower,while the expression of PI3K,AKT protein and mRNA was higher in the IGF-1 group (P＜0.05).There was no significant difference in the invasion and migration of A549 cells among the blank group,pcDNA-PTEN control group and siRNA-PTEN control group (P＞0.05).The invasion and migration ability of A549 cells in the pcDNA-PTEN group was significantly lower than that in the pcDNA-PTEN control group (P＜0.05).The invasion and migration ability of A549 cells in the siRNA-PTEN group was significantly higher than that in the siRNA-PTEN control group (P＜0.05).The invasion and migration ability of A549 cells in the IGF-1 group was significantly higher than that in the blank group and wortmannin group (P＜0.05).The invasion and migration ability of A549 cells in the wortmannin group was significantly lower than that in the blank group (P＜0.05).The invasion and migration ability of A549 cells in the pcDNA-PTEN+IGF-1 group was significantly higher than that in the pcDNA-PTEN group (P＜0.05).With the increase of concentration of cisplatin,the survival rate of A549 cells in each group decreased gradually (P＜0.05).Under the intervention of different concentrations of cisplatin,there was no significant difference in the survival rate of A549 cells between the blank group,and the pcDNA-PTEN control group and the siRNA-PTEN control group (P＞0.05),the survival rate of A549 cells in the pcDNA-PTEN group was significantly lower than that in the pcDNA-PTEN control group (P＜0.05),the survival rate of A549 cells in the siRNA-PTEN group was significantly higher than that in the siRNA-PTEN control group (P＜0.05),the survival rate of A549 cells in the wortmannin group was significantly lower than that in the blank group (P＜0.05),the survival rate of A549 cells in the IGF-1 group was significantly higher than that in the wortmannin group and the blank group (P＜0.05),and the survival rate of A549 cells in the pcDNA-PTEN+IGF-1 group was significantly higher than that in the pcDNA-PTEN group (P＜0.05).Conclusion　The expression of PTEN protein is decreased in NSCLC tissues.The up-regulation of PTEN gene expression may reduce the cell invasion and migration by inhibiting the activity of PI3K/AKT signaling pathway,and enhance the sensitivity of NSCLC cells to cisplatin.

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更新日期/Last Update: 2020-06-05