唑来膦酸对胃癌耐药细胞株SGC一7901／ADR的生物学作用

Biological effects of zoledronic acid on gastric cancer drug-resistant cell line SGC-7901/ADR

目的观察唑来膦酸（ZOL）对人胃癌耐阿霉素（ADM）细胞株SGC一7901／ADR侵袭和黏附的影响，探讨ZOL是否具有逆转该细胞株对ADM的耐药作用及其机制。方法采用噻唑蓝（MTT）比色法检测1×10-4mol/L的ZOL联合0．5mg／L的ADM对SGC-7901／ADR细胞黏附作用的影响；Transwell实验分析上述浓度的ZOL和ADM对细胞侵袭力的影响；酶联免疫吸附试验（ELISA）法检测细胞上清液基质金属蛋白酶（MMP）-2、细胞问黏附分子一1（ICAM-1）蛋白表达水平的变化；实时定量聚合酶链反应（Real—timePCR）检测细胞MMP-2、基质金属蛋白酶抑制因子（TIMP）．2、MMP-9、ICAM一1、CD44mRNA表达水平的改变。结果（1）1×10。mol／L的ZOL单药及联合0．5mg／L的ADM与细胞共同孵育6h，黏附率分别为：对照组（C组）44．32％，ZOL单药组（Z组）34．73％，ZOL＋ADM组（z＋A组）31．30％，与C组比较，单药能够抑制细胞的黏附力（P〈0．05），z＋A组细胞黏附能力进一步下降（P〈0．01）；（2）在研究侵袭能力的实验中，C组、z组、z＋A组每个高倍视野中细胞数（个）分别为：103．73±7．84、97．64±9．62、71．58±10．57，与对照组比较，联合用药时抑制作用显著（P〈0．05）；（3）ELISA结果显示，与C组比较、z组及z＋A组均能够显著降低细胞上清液中的MMP-2和ICAM-1蛋白表达水平（P〈0．05或P〈0．01），与z组比较，Z＋A组差异统计学-意义（P〈0．01）。（4）在Real—timePCR实验中，以C组作为参考标准，药物和细胞共同孵育后，上清液CD44mRNA表达在C组、Z组、A组、z＋A组分别为1．0000±0．1083、0．2601±0．0041、0．46814-0．0314、0．0950±0．0052，同时ICAM-1、MMP-2、MMP-9mRNA的表达在各组中也表现出类似结果。结论ZOL对人胃癌耐ADM细胞株SGC-7901／ADR具有抑制降低细胞黏附和侵袭能力的作用，一定程度上逆转该细胞株对ADM的耐药，这些作用和下调ICAM-1、MMP-2、MMP-9、CD44mRNA及上调TIMP-2mRNA的表达水平相关。

Objective To detect effects of zoledronic acid on the invasion and adhesion of adria- mycin resistant gastric cancer cell line SGC-7901/ADR, and to investigate whether zoledronic acid could restore sensitivity to adriamycin and possible mechanism. Methods Methyl thiazol tetrazolium （MTT） as- say was used to investigate the effect of 1 x 10-4 tool/L zoledronic acid and 0. 5 mg/L doxorubicin on the adhesion of SGC7901/ADR cells. Transwell was used to investigate cellular invasive ability. Enzyme linked immunosorbent assay （ELISA） was used to quantify supernatant matrix metalloproteinase （MMP）-2 and intercellular adhesion molecule-1 （ICAM-1）. MMP-2, tissue inhibitorof metalloproteinase （TIMP）-2, MMP-9, ICAM-1 and CD44 levels were quantified by using real-time polymerase chain reaction （PCR）. Results （ 1 ） After SGC-7901/ADR cells were incubated with 1 x 10 4 mol/L zoledronie acid （Z） alone or in combined with O. 5 mg/L adriamycin （ Z ＋ A） for 6 h, the adhesion rate was 44. 32% for control （ C ） group, 34.73% for Z group, and 31.30% for Z ＋ A group. The adhesion rate was reduced in treated groups as compared with untreated group （ P 〈 0. 05 ） ; （ 2 ） The number of invasive cells per high power field in C group, Z group and Z ＋ A group was 103.73 ±7.84, 97.64 ±9. 62, and 71.58_＋10.57 re- spectively. As compared with control group, the invasive ability was reduced significantly in Z ＋ A group （P 〈 0. 05 ） ; （3） ELISA results indicated that as compared with control group, the MMP-2 and ICAM-1 in the supernatant of Z group and Z ＋ A group were significantly reduced （P 〈0. 05 or P 〈0. 01 ）. As corn-pared with Z group, both MMP-2 and /CAM-1 were significantly reduced in Z ＋ A group （ P 〈 0. 01 ） ; （4） CIM4 mRNA expression in C group, Z group, and Z ＋ A group was 1. 0000 ＋ 0. 1083, 0. 2601 -＋ 0. 0041,0. 4681 -＋ 0. 0314, and 0. 0950 ＋- 0. 0052 respectively. Similar results were observed in the ex- pression of ICAM-1, MMP-2 and MMP-9. Conclusion Zoledronie acid could inhibit the adhesion and in- vasion of SGC-7901/ADR cells, and restore the sensitivity of SGC-7901/ADR cells to doxorubicin. These effects may be related to the decreased expression of ICAM-1, MMP-2, MMP-9 and CD44, and the in- creased TIMP-2 expression.