摘要
目的建立紫杉醇(PTX)耐药人胃癌细胞HGC-27/PTX,探讨耐药前后细胞特征的变化并初步分析紫杉醇耐药机制。方法逐步递增紫杉醇浓度并间歇作用于人胃癌细胞系HGC-27,建立紫杉醇耐药细胞HGC-27/PTX。用CCK-8及流式细胞仪检测细胞的半数抑制浓度(IC50)及细胞周期分布,RNAseq法筛选紫杉醇耐药前后差异表达基因及通路。结果 9个月后,相同浓度紫杉醇对HGC-27/PTX细胞的增殖抑制作用较亲本HGC-27细胞明显减弱(P<0.05),耐药细胞的形态较亲本细胞略有不同。与亲本HGC-27细胞相比,HGC-27/PTX细胞的S期及G2/M期细胞比例明显增多(P<0.01)。HGC-27/PTX细胞较亲本HGC-27细胞具有274个显著差异表达基因(DEGs),表达上调与下调基因分别有130个和144个,差异基因主要富集在ECM-receptor interaction通路(P<0.001)和PI3KAkt信号通路(P<0.05),这为逆转紫杉醇耐药提供有力线索。结论成功建立紫杉醇耐药胃癌细胞HGC-27/PTX并可体外稳定传代,为深入研究耐药机制提供了理想的体外实验模型。
Objective To establish the paclitaxel-resistant gastric cancer cell（HGC-27/PTX） and to investigate the changes of characteristics before and after resistance, as well as the possible resistant mechanisms. Methods The paclitaxel-resistant gastric cancer cell HGC-27/PTX was established by increasing paclitaxel dose gradually and in- termittently. The IC50（50% inhibitory concentration） and cell cycle were determined by CCK-8 assay and flow cy- tometry, respectively. The differentially expressed genes （DEGs） and signaling pathways were analyzed using RNAseq. Results The establishment of HGC-27/PTX cells lasted 9 months, and the sensitivity of paclitaxel of HGC-27/PTX cells was significantly lower than parental cells （P〈0. 05 ）. Compared to parental cells, the morphol- ogy of HGC-27/PTX cells was slightly different, and the proportion of S and G2/M phase was obviously increased（ P〈 0. 01 ）. A total of 274 DEGs were identified between the resistant and parental cells with 130 genes up-regulated and 144 genes down-regnlated.DEGs were significantly enriched in extracellular matrix （ECM）-receptor interaction （P〈 0. 001） and PI3K-Akt signaling pathways （P〈0. 05）, which could provide evidences for reversing paclitaxel resist- ance. Conclusions The paclitaxel-resistant gastric cancer cells HGC-27/PTX was established with stable culturein vitro, which provides an ideal model for future study on the mechanism of drug resistance.
出处
《基础医学与临床》
CSCD
2017年第8期1146-1151,共6页
Basic and Clinical Medicine
