CAJ | 학술논문

目的探讨葡萄糖神经酰胺合成酶基因(glucosylceramide synthase gene,GCS)和多药耐药基因(multidrug resistance 1 gene,MDR1)在人白血病多药耐药细胞株K562/A02耐药形成中的相关性,以便为逆转白血病细胞耐药提供新的思路.方法应用逆转录聚合酶链反应(reverse transcription-PCR,RT-PCR)方法检测野生株K562及其耐药株K562/A02中GCS、MDR1基因的表达情况;采用RNA干扰技术(RNA interference,RNAi),将靶基因为GCS的小干扰RNA(GCS siRNA)和靶基因为MDR1的小干扰RNA(MDR1 siRNA)分别传染K562/A02细胞后,通过定性和荧光实时定量PCR的方法观察GCS及MDR1 mRNA的基因沉默现象及两者之间的交互影响.结果 KS62/A02细胞GCS和MDR1 mRNA的表达明显强于药物敏感株K562细胞;GCS siRNA组K562/A02细胞GCS和MDR1基因表达均被抑制,抑制率分别为73%(59%～82%)和67%(38%～82%);MDR1 siRNA组K562/A02细胞MDR1基因被抑制,抑制率为81%(63%～91%),GCS基因表达无改变.结论 GCS与MDR1 mRNA的表达在K562/A02细胞呈正相关,特异性的沉默GCS基因可以下调MDR1 mRNA的表达水平.
목적 탐토포도당신경선알합성매기인(glucosylceramide synthase gene,GCS)화다약내약기인(multidrug resistance 1 gene,MDR1)재인백혈병다약내약세포주K562/A02내약형성중적상관성,이편위역전백혈병세포내약제공신적사로.방법 응용역전록취합매련반응(reverse transcription-PCR,RT-PCR)방법검측야생주K562급기내약주K562/A02중GCS、MDR1기인적표체정황;채용RNA간우기술(RNA interference,RNAi),장파기인위GCS적소간우RNA(GCS siRNA)화파기인위MDR1적소간우RNA(MDR1 siRNA)분별전염K562/A02세포후,통과정성화형광실시정량PCR적방법관찰GCS급MDR1 mRNA적기인침묵현상급량자지간적교호영향.결과 KS62/A02세포GCS화MDR1 mRNA적표체명현강우약물민감주K562세포;GCS siRNA조K562/A02세포GCS화MDR1기인표체균피억제,억제솔분별위73%(59%～82%)화67%(38%～82%);MDR1 siRNA조K562/A02세포MDR1기인피억제,억제솔위81%(63%～91%),GCS기인표체무개변.결론 GCS여MDR1 mRNA적표체재K562/A02세포정정상관,특이성적침묵GCS기인가이하조MDR1 mRNA적표체수평.
Objective To investigate the correlation of glucosylceramide synthase (GCS) gene and multidrug resistance 1 ( MDR1 ) gene in inducing multidrug resistance in human multidrug-resistant K562/ A02 cell line, and search for a novel strategy for reversing multidrug resistance of leukemia cells. Methods The expression levels of GCS and MDR1 mRNA in K562 and K562/A02 cells were assayed by RT-PCR. siRNAs targeting the GCS and MDR1 gene were transfected into K562/A02 cells with liposome, respectively. The differential expression of GCS and MDR1 mRNAs, as well as their correlation, were detected by RT-PCR and real time quantitative-PCR(QPCR). Results The expression level of GCS and MDR1 mRNA was dramatically lower in drug-sensitive K562 cells compared with the K562/A02 cells. The GCS mRNA was inhibited by 73%(59%-82%) and MDR1 mRNA expression was down regulated by 67% (38%-82%) in K562/A02 cells after being transfected with GCS siRNA. The expression level of MDR1 mRNA was inhibited by 81 %(63%-91%) and GCS mRNA expression had no apparent change in K562/A02 cells treated with MDR1 small interference RNA(siRNA). Conclusion Positive correlation was detected between the expression of GCS and MDR1 mRNA in K562/A02 cells and MDR1 mRNA expression was down regulated after silencing the GCS gene expression.