阻断胰腺癌细胞膜hENTs对氟尿嘧啶细胞毒性的影响

The effect of human equilibrative nucleoside transport (hENTs) in pancreatic cancer cell membrane on the cytotoxicity of 5-fluorouracil

目的 研究胰腺癌细胞株中核苷载体（NTs）的表达，以及应用潘生丁（dipyridamole，DP）阻断平衡型核苷转运载体（hENTs）后，5-氟尿嘧啶（5-Fluorouraci，5-FU）对胰腺癌细胞株毒性作用的影响。方法 选择胰腺癌细胞株Panc-1、Sw1990和Aspc-1进行实验。RT-PCR法检测细胞株hENT1、hENT2、hCNT1、hCNT2和hCNT3的mRNA表达。根据DP浓度将各细胞株分为4个实验组，即DP空白组，DP浓度为5μmol·L^-1组、10μm01·L^-1组和40μmol·L^-1组，每组细胞分别在含有一定浓度（0-2×10^8ng·L^-1）的5-FU的培养液中培养48h。MTT法检测每组3种细胞株的增殖，并计算各组5-Fu的半数抑制浓度（IC50）。结果hENT1的mRNA在Panc-1和Sw1990中表达较在Aspc-1中高（P〈0．05），hENT2在Aspc-1中未见表达；hCNTs在3种细胞株中均有表达，且无明显差异。DP本身对3种细胞株无毒性作用，而5-FU与其联合后能明显提高5-FU对高表达hENTs细胞（Panc-1和Sw1990）的毒性作用；在DP 10μmol·L^-1时，对Panc-1细胞的毒性作用增强7倍（P〈0．01），对Sw1990细胞增强10倍（P〈0．01）。而在hENT1低表达、hENT2未见表达的细胞（Aspc-1）中，DP对5-FU的毒性作用没有影响。结论在胰腺癌细胞株中，DP阻断细胞膜上hENTs后能显著增强5-FU的作用效果，提示在临床上可能需要依据hENTs的表达情况来选择联合应用hENTs阻断剂增强核苷类抗癌药物作用。

Objective To investigate subtype expressions of nucleoside transporters in pancreatic cancer cell lines, and the effects of dipyridamole （DP）, one of the hENTs blockers, on 5- Fluorouracil （5-FU） cytotoxicity to the cell lines. Methods Three pancreatic cancer cell lines（Panc-1 ,Sw 1990,Aspc-1 ） were chosen to detect the mRNA expressions of hENT1, hENT2, hCNT1, hCNT2 and hCNT3 with RT-PCR. According to the DP concentration in the study, the cell lines were divided into four groups, the zero DP concentration group, 5μmol·L^-1 group,10 μmol·L^-1 group and 40 μmol·L^-1group. The cells in each group were incubated in the medium with a serial concentrations of S-FU from 0 to 2×10^8ng L^-1 for 48 h. Then the cell proliferation in each group was measured with MTT assay and the IC50 were evaluated. Results The expression of hENT1 was higher in Panc-1 and Sw1990 than that in Aspc-1 （P〈0. 05） ,while the hENT2 was not to be detected in Aspc-1. All the three cell lines expressed hCNTs but no difference was found. DP itself did not present any toxic effects on the three cell lines. However, combined with 5-FU, the cytotoxicity to the cells with high expressions of hENTs （Panc-1 and Sw1990） was significantly increased,7 times increased in Panc-1 and 10 times increased in Sw1990 in 10μmol·L^-1 of DP （P〈0.01 ）. In contrast,in the Aspc-1 cells with low hENT1 expression and no bENT2 expression, the 5-FU cytotoxicity was not influenced by DP. Conclusions DP can significantly enhance the 5- FU cytotoxic effect on the pancreatic cancer cell by blocking hENTs transporters on the cell membrane. It is implicated that the hENTs expression may be the clue for the choice of hENTs blockers with nucleoside anticancer drugs in clinic.