摘要目的:构建人源胰腺癌细胞株PANC1与鼠源胰腺癌细胞株PANC02的吉西他滨耐药细胞株，并探究其生物学行为改变情况。方法:采用浓度梯度递增法建立人源胰腺癌细胞PANC1及鼠源胰腺癌细胞PANC02的吉西他滨耐药细胞株，即PANC1-GR、PANC02-GR。采用细胞计数试剂(CCK-8)、流式细胞术、细胞划痕及Transwell实验检测四组细胞的耐药、增殖、细胞周期、迁移与侵袭等，并与亲本细胞比较。结果:PANC1-GR及PANC02-GR的耐药指数分别为153.3和185.4。CCK-8检测结果显示耐药细胞的增殖速度明显增快，PANC1-GR较PANC1的细胞群体倍增时间缩短(1.5±0.1)d比(2.4±0.2)d，差异有统计学意义( t＝8.00， P<0.001)。流式细胞仪结果显示两种耐药细胞处于S期与G2/M期的细胞比例增加，G0/G1期的比例降低。细胞划痕及Transwell实验显示，与亲本细胞相比，PANC1-GR和PANC02-GR细胞的24 h迁移率增高(47.6±2.4)%比(28.7±6.3)%、(53.6±3.2)%比(30.1±1.4)%，穿膜数量(放大200倍的单个视野)也增加(269.7±30.9)个比(62.7±10.1)个、(172.0±30.8)个比(36.3±4.9)个，差异均有统计学意义(均 P<0.05)。 结论:浓度梯度递增法可建立胰腺癌吉西他滨耐药细胞株，耐药细胞株的增殖能力、迁移、侵袭性比亲本更强，为耐药机制研究提供基础。

abstractsObjective:To construct the gemcitabine resistant cell lines of human pancreatic cancer cell line (PANC1) and mouse pancreatic cancer cell line (PANC02), and to investigate their biological behavior changes.Methods:Gemcitabine-resistant cell lines PANC1-GR of human pancreatic cancer and PANC02-GR of mouse pancreatic cancer were induced by concentration gradient increment method. Cell count assay (CCK-8), flow cytometry, cell scratch assay and Transwell assay were used to detect the drug resistance, proliferation, cell cycle, migration and invasion of the four groups of cell lines. The drug-resistant cells were also compared with the parent cells.Results:The resistance indices of PANC1-GR and PANC02-GR were 153.3 and 185.4, respectively. The results of CCK-8 showed that with the increase of gemcitabine concentration, the proliferation of resistant cells changed significantly compared with parental cells, the population doubling time of PANC1-GR was significantly shorter than that of PANC1 (1.5±0.1) d vs (2.4±0.2) d ( t=8.00, P<0.001). The proportion of cells in S and G2/M phase increased, and the proportion of cells in G0/G1 phase decreased. The cell scratch and Transwell experiments indicated that the 24h mobility of PANC1-GR and PANC02-GR was higher than that of parent cells (47.6±2.4)% vs (28.7±6.3)% and (53.6±3.2)% vs (30.1±1.4)%, the number of individual field (200 times magnification) penetrating membrane cells was also higher than that of parent cells (269.7±30.9) vs (62.7±10.1) and (172.0±30.8) vs (36.3±4.9), with statistical significance (all P<0.05). Conclusion:Concentration gradient increment method can successfully establish gemcitabine-resistant pancreatic cancer cell lines, which have stronger proliferation, migration and invasiveness, and can be used to study the mechanism of drug resistance in pancreatic cancer.