CAJ | 학술논문

目的：通过表达水通道蛋白-8（AQP-8）真核表达载体，观察水通道蛋白AQP-8对HT-29细胞凋亡及凋亡抑制蛋白（ IAPs）表达的影响。方法取对数生长期的人结肠癌HT-29细胞株用于实验。构建AQP-8真核表达载体并转染HT-29细胞，通过Western blot检测GFP-AQP-8转染效率；采用MTT法检测各组细胞增殖抑制率；流式细胞术检测细胞凋亡率；通过Real Time-PCR和Western blot检测转染GFP-AQP-8的HT-29细胞凋亡抑制蛋白（ IAPs）家族成员c-IAP1、c-IAP2、XIAP、NIAP、Survivin和Livin表达水平。结果 Western blot结果显示，GFP-AQP-8转染后结肠癌HT-29细胞AQP-8基因表达显著上调（ P ＜0{．05）。 MTT分析结果显示，转染了GFP-AQP-8的结肠癌HT-29细胞增殖抑制率显著增加（ P ＜0．05）；流式细胞分析发现，转染了GFP-AQP-8的HT-29细胞凋亡率显著增加（ P ＜0．05）。Real Time-PCR和Western blot结果显示，与转染GFP-N1的阴性对照组相比较，转染了GFP-AQP-8的HT-29细胞c-IAP1、c-IAP1、XIAP、Livin和Survivin的mRNA和蛋白表达水平下降（ P ＜0．05），NIAP表达变化不明显（ P ＜0．05）。结论过表达AQP-8可抑制HT-29细胞生长，并能通过下调c-IAP1、c-IAP1、XIAP、Livin和Survivin表达诱导HT-29细胞凋亡。
목적：통과표체수통도단백-8（AQP-8）진핵표체재체，관찰수통도단백AQP-8대HT-29세포조망급조망억제단백（ IAPs）표체적영향。방법취대수생장기적인결장암HT-29세포주용우실험。구건AQP-8진핵표체재체병전염HT-29세포，통과Western blot검측GFP-AQP-8전염효솔；채용MTT법검측각조세포증식억제솔；류식세포술검측세포조망솔；통과Real Time-PCR화Western blot검측전염GFP-AQP-8적HT-29세포조망억제단백（ IAPs）가족성원c-IAP1、c-IAP2、XIAP、NIAP、Survivin화Livin표체수평。결과 Western blot결과현시，GFP-AQP-8전염후결장암HT-29세포AQP-8기인표체현저상조（ P ＜0{．05）。 MTT분석결과현시，전염료GFP-AQP-8적결장암HT-29세포증식억제솔현저증가（ P ＜0．05）；류식세포분석발현，전염료GFP-AQP-8적HT-29세포조망솔현저증가（ P ＜0．05）。Real Time-PCR화Western blot결과현시，여전염GFP-N1적음성대조조상비교，전염료GFP-AQP-8적HT-29세포c-IAP1、c-IAP1、XIAP、Livin화Survivin적mRNA화단백표체수평하강（ P ＜0．05），NIAP표체변화불명현（ P ＜0．05）。결론과표체AQP-8가억제HT-29세포생장，병능통과하조c-IAP1、c-IAP1、XIAP、Livin화Survivin표체유도HT-29세포조망。
Objective To investigate the effects of aquaporin-8 ( AQP-8 ) on cell apoptosis and expression of inhibitors of apoptosis proteins( IAPs) by means of eukaryotic expression vector that expresses AQP-8 in human colon cancer HT-29 cells in vitro.Methods Human colon cancer HT-29 cells at logarithm growth period were used in the experiment. AQP-8 eukaryotic expression vector was constructed and transfected into HT-29 cells.Western Blot was used to detect the transfection efficiency;MTT assay was used to detect the cell growth inhibition rate;flow cytometry( FCM) was used to measure the cell apoptosis rate.The expression levels of mRNA and protein of IAPs including c-IAP1,c-IAP2,XIAP,NIAP,Survivin and Livin were detected by fluorescence quantitative Real-time RT-PCR and Western Blot.Results The results by Western Blot showed that the expression levels of AQP-8 were significantly up-regulated in colon cancer HT-29 cells after GFP-AQP-8 transfection ( P <0.05).MTT analysis showed that the proliferation inhibition rate was increased significantly in HT-29 cells after GFP-AQP-8 transfection,as compared with that in GFP-N1 control group ( P <0.05).FCM analysis showed that GFP-AQP-8 transfection obviously induced HT-29 cell apoptosis ( P <0.05 ) .The results of fluorescence Real-time quantitative RT-PCR and Western Blot showed that the expression levels of mRNA and protein of c-IAP1, c-IAP1, XIAP, Livin,and survivin in HT-29 cells were significantly decreased by AQP-8 overexpression ( P <0.05).However,the expression levels of NIAP mRNA and protein had no obvious change ( P >0.05).Conclusion The overexpression of AQP-8 can inhibit growth of HT-29 cell and can induce HT-29 cell apoptosis by down-regulating the expression levels of c-IAP1,c-IAP1,XIAP, Livin and survivin.