抑制Src酪氨酸激酶活性对人肺癌A549/DDP细胞耐药性及MDR1和LRP表达的影响

摘要

背景与目的 本研究旨在探讨抑制Src酪氨酸激酶活性对人肺癌A549/DDP细胞耐药性及多药耐药蛋白( muti-drug resistance 1,MDR1)和肺耐药相关蛋白(lung resistance-related protein,LRP)表达的影响.方法 以Src酪氨酸激酶抑制剂作用于A549/DDP细胞,应用Western blot法检测肿瘤细胞Src酪氨酸激活性的变化,CellTiter-Glo发光法检测肿瘤细胞药物敏感性的变化,流式细胞仪检测肿瘤细胞Rh-123含量变化.Western blot法和RT-PCR检测肿瘤细胞MDR1和LRP表达变化.结果 Src酪氨酸激酶抑制剂可下调A549/DDP细胞中Src酪氨酸激活性,2.SμM和10 μM Src酪氨酸激酶抑制剂作用肿瘤细胞后,肿瘤细胞药物敏感性提高,逆转倍数(reversal fold,RF)分别为1.59倍和2.10倍,肿瘤细胞中 Rh-123含量分别提高了1.21倍和1.59倍,MDR1 mRNA表达分别是对照组的53.8％和27.5％,LRP mRNA表达分别是对照组的59.3％和21.4％,MDR1和LRP蛋白表达水平明显下降.结论 抑制A549/DDP细胞中Src酪氨酸激酶活性可逆转肿瘤细胞多药耐药性,提高肿瘤细胞药物敏感性,其机制可能与降低细胞MDR1和LPP表达有关%Background and objective The aim of this study is to investigate the effect of Src tyrosine kinase inhibition on the drug-resistance as well as the expression of multidrug resistance 1 (MDR1) and lung resistance-related protein (LRP) of the human cis-platinum-resistant lung cancer cell line A549/DDP. Methods 4-AnilinoquirazoIine was used to inhibit Src tyrosine kinase activity in A549/DDP. Western blot analysis was used to detect the Src tyrosine kinase activity. CellTiter-Glo assay was used to detect the drug sensitivity of tumor cells. Flow cytometry was used to detect the intracellular Rh-123 content. Western blot and real-time PCR assay were used to detect the expression of tumor MDRl and LRP. Results 4-Anilinoquirazoline can down-regulate the cellular Src tyrosine kinase activity in A549/DDP. After treatment with 2.5 μM and 10 uM of 4-anilinoquirazoline, the cells became more sensitive to the drug and the reversal folds (RFs) oftumor cell sensitivity to the drug were 1.59. and 2.10-fold, respectively. The intracellular content ofRh-123 improved by 1.21- and l.S9-fold, respectively. The mRNA levels of MDRl were 53.8% and 27.5% of the control, respectively. The mRNA level of LRP was 59.3% and 21.4% of the control, respectively. The expression of MDRl and LRP protein significantly decreased. Conclusion The inhibition of Src tyrosine kinase activity in A549/DDP cells can reverse multi-drug resistance and increase the sensitivity of the cells to the drug. The mechanism may be related to the down-regulation of cellular MDRl and LRP.