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目的 :探索微RNA-21(micro RNA-21,mi R-21)以及与Bcl-2相互作用的细胞死亡调节子(Bcl-2 interacting mediator of cell death,BIM)蛋白在人肺腺癌细胞发生表皮生长因子受体(epidermal growth factor receptor,EGFR)-酪氨酸激酶抑制剂(tyrosine kinase inhibitors,TKIs)获得性耐药过程中的作用及调控关系。方法 :将重组质粒pc DNA3.1-BIM和空质粒pc DNA3.1分别瞬时转染至吉非替尼耐药的肺腺癌细胞株PC9R中,采用实时荧光定量PCR法检测各组细胞中mi R-21的表达水平,细胞计数试剂盒-8(cell counting kit-8,CCK-8)法检测PC9R细胞对吉非替尼的敏感性变化。同时,采用慢病毒感染的方法干扰PC9R细胞株中mi R-21的表达,然后采用实时荧光定量PCR法和蛋白质印迹法检测细胞中BIM基因的表达水平,CCK-8法检测PC9R细胞对吉非替尼的敏感性变化。另外,在干扰mi R-21表达的PC9R细胞中转染pc DNA3.1-BIM重组质粒,然后采用CCK-8法检测PC9R细胞对吉非替尼的敏感性变化。结果 :重组质粒pc DNA3.1-BIM转染后,PC9R细胞中BIM的表达水平明显提高(P<0.01),mi R-21的表达水平也相应升高(P<0.01)。慢病毒干扰mi R-21表达后,PC9R细胞中mi R-21的表达水平明显降低(P<0.05),BIM的表达水平也相应降低(P<0.05)。下调mi R-21水平和上调BIM表达均能提高PC9R细胞对吉非替尼的敏感性(P值均<0.05),而在下调mi R-21表达的同时上调BIM表达,更加提高了细胞对吉非替尼的敏感性(P<0.05)。结论 :mi R-21和BIM基因在逆转吉非替尼耐药过程中可能起关键作用,并且二者存在相互拮抗的作用。
OBJECTIVE: To explore the role of microRNA-21 (mi R-21) and Bcl-2 interacting mediator of cell death (BIM) proteins in human lung adenocarcinoma The role and regulation of epidermal growth factor receptor (EGFR) -type tyrosine kinase inhibitors (TKIs) in acquired drug resistance. Methods: The recombinant plasmid pcDNA3.1-BIM and empty plasmid pcDNA3.1 were transiently transfected into the gefitinib-resistant lung adenocarcinoma cell line PC9R. Real-time fluorescence quantitative PCR was used to detect the mi R-21 and cell counting kit-8 (CCK-8) were used to detect the sensitivity of PC9R cells to gefitinib. At the same time, lentivirus infection interfered with the expression of mi R-21 in PC9R cell line, and then the expression of BIM gene was detected by real-time fluorescence quantitative PCR and Western blot. The CCK-8 assay was used to detect the expression of mi R- Changes in the sensitivity of tini. In addition, pcDNA3.1-BIM recombinant plasmid was transfected into PC9R cells that interfered with mi R-21 expression, and then the sensitivity of PC9R cells to gefitinib was tested by CCK-8 assay. Results: After transfection with recombinant plasmid pcDNA3.1-BIM, the expression of BIM in PC9R cells was significantly increased (P <0.01) and the expression of mi R-21 was also increased (P <0.01). The expression of mi R-21 in PC9R cells was significantly decreased (P <0.05) after lentiviral interference with mi R-21 expression, and the expression level of BIM was also decreased accordingly (P <0.05). Down-regulation of mi R-21 and up-regulation of BIM increased the sensitivity of PC9R cells to gefitinib (all P <0.05), while up-regulated BIM expression while down-regulating the expression of mi R-21, Gefitinib sensitivity (P <0.05). CONCLUSION: The mi R-21 and BIM genes may play key roles in reversing the resistance to gefitinib, and their antagonism may exist.