下调AFAP-1L2表达对人胰腺癌细胞吉西他滨化疗敏感性的影响

Influence of down-regulation of AFAP-1L2 on the chemotherapy sensitivity of human pancreatic cancer cells to gemcitabine

目的观察下调AFAP-1L2表达对胰腺癌细胞吉西他滨(Gem)化疗敏感性的影响及对胰腺癌细胞的杀灭作用。方法构建靶向siAFAP-1L2干扰质粒下调AFA-1L2表达,转染MIAPACA-2细胞,以Western blot法及(qRT)-PCR法检测转染效率;MTT检测AFAP-1L2下调后细胞存活率。对siAFAP-1L2转染MiaPaCa-2细胞给予吉西他滨干预,采用MTT法检测细胞增殖,流式细胞术检测细胞凋亡。结果与siNegative control组比较,给予不同浓度梯度siAFAP-1L2转染MIAPACA-2细胞,siAFAP-1L2组细胞存活率较低(P<0.05);MiaPaCa-2细胞在培养24、48、72、96 h后,转染siAFAP-1L2组细胞在48 h后存活率较低(P<0.05);siAFAP-1L2转染后细胞存活率与siAFAP-1L2转染浓度和时间具有相关性(r=-0.921,r=-0.907)。将化疗药Gem 20 nmol/L加入分别以不同浓度梯度siAFAP-1L2转染的胰腺癌细胞48 h后,与siNegative control组比较,siAFAP-1L2组细胞存活率较低(P<0.05);将化疗药Gem 20 nmol/L加入siAFAP-1L2 100 nmol/L转染的MiaPaCa-2细胞,培养24、48、72、96 h后,与siNegative control组比较,siAFAP-1L2组细胞在72、96 h后存活率较低(P<0.05);siAFAP-1L2转染后,Gem化疗细胞存活率与siAFAP-1L2转染浓度和时间具有相关性(r=-0.967,r=-0.984)。siAFAP-1L2+Gem组与siAFAP-1L2组凋亡率分别为37.28%、11.65%,差异有统计学意义(P<0.05)。结论下调AFAP-1L2表达可增强胰腺癌细胞对吉西他滨化疗的敏感性,促进对胰腺癌细胞的杀伤作用,可作为胰腺癌治疗的靶向候选基因。

Objective To observe the influence of down-regulation of AFAP-IL2 expression on the chemotherapy sensitivity of pancreatic cancer cells to gemcitabine and its killing effect on pancreatic cancer cells. Methods Target siAFAP-IL2 interfering plasmid was constructed to down-regulate the AFAP-IL2 expression and transfect into MiaPaCa-2 cells. Interfering effect was detected by Western blot and （qRT）-PCR. The cell viability after AFAP-IL2 down-regulation was detected by MTT assay. The transfected siRNA cells was interfered with gemcitabine,and the proliferation and apoptosis of cells was detected by MTT assay and flow cytometry,respectively. Results Compared with siNegative control group, the cell viability was lower in siAFAP-IL2 group when different concentration grade of siAFAP-IL2 was given （ P 〈 0. 05 ） ; MiaPaCa-2 cell viability rate was lower in siAFAP-IL2 group after culture for 48 h （P 〈0. 05 ）, which was correlated with transfected concentration and time of siAFAP-IL2 （r- -0. 921, r- -0. 907）. Gemcitabine 20 nmol/L was given into MiaPaCa-2 cells which were transfected with different concentration grade of siAFAP-IL2 for 48 h,the cell viability in siAFAP-IL2 group was lower than that in siNegative group （P 〈 0. 05） ;when Gem 20 nmol/L was given into MiaPaCa-2 cells which were transfected with siAFAP-IL2 100 nmol/L, the cells were cultured for 24,48,72 and 96 h,the cell viability was lower in siAFAP-IL2 group than that in siNegative control group after the cells were cultured for 72 h and 96 h;the cell viability interfered with Gem was correlated with transfected concentration and time of siAFAP-IL2 （ r - - 0. 967, r - - 0. 984 ）. The apoptosis rate in siAFAP-IL2 ＋ Gem group was higher than that in siAFAP-I L2 group （ 37. 28 % vs. 11.65 % , P 〈 0. 05 ）. Conclusion AFAP-I L2 down-regulation can increase the sensitivity of pancreatic cancer cells to gemcitabine,and attenuate the killing effect on pancreatic cancer cells, which may be a target candidate for pancreatic cancer therapy.