摘要
目的通过体外实验探讨人端粒酶反转录酶（hTERT）反义寡核苷酸（ASODN）能否增强胰腺癌细胞对吉西他滨（健择）的敏感性。方法采用实时荧光定量逆转录聚合酶链反应（RT—PCR）法检测hTERTmRNA表达情况；端粒重复序列扩增法（TRAP）、聚丙烯酰胺凝胶电泳和银染法检测端粒酶活性；四甲基偶氮唑盐（MTT）法检测细胞增殖能力；膜联蛋白V-异硫氰酸荧光素（Annexin V-FITC）和碘化丙啶（PI）双染色法检测早期凋亡。结果瞬时转染hTERTASODN呈浓度依赖性下调细胞hTERTmRNA表达。0．2μmol／LhTERTASODN可显著下调端粒酶活性至0．47。健择在ASODN转染组中的IC50值为（0．8±0．2）μmol／L，而其在单纯脂质体转染对照组中为（7．3±0．9）μmol／L，差异具有统计学意义。ASODN可显著增加健择对肿瘤细胞的凋亡诱导作用，健择诱导的早期凋亡率在ASODN转染组中为60．28％，而其在脂质体对照组中为17．34％。结论hTERT ASODN可增强胰腺癌细胞对健择的敏感性，其机制可能与hTERTmRNA和端粒酶活性下调相关。
Objective To evaluate whether antisense oligonucleotides （ASODN） targeting hTERT mRNA could sensitize human pancreatic cancer cell to gemcitabine in vitro. Methods hTERT mRNA expression was measured by real-time quantitative reverse transcription polymerase chain reaction （RT-PCR） assay. Telomerase activity was examined by TRAP, polyacrylamide gel electrophoresis and silver staining. The proliferation capacity and the apoptosis of cancer cells were evaluated by MTT assay and double staining with both Annexin V-FITC and PI. Results Transient transfection in human pancreatic cancer cell with hTERT ASODN diminished the abundance of hTERT mRNA in a concentration-dependent fashion. And 0.2 μmol/L ASODN decreased the telomerase activity by 0. 47-fold in cancer cell. The IC50 value of gemcitabine in ASODN-transfected cell was （ 0. 8 ± 0. 2 ） μmol/L while that in oligofectamine-transfected control cell （7.3 ± 0. 9） μmol/L with a statistically significant difference, hTERT ASODN significantlly increased the gemcitabine-induced apoptosis rate in pancreatic cancer cell. The gemeitabine-induced apoptosis rate in ASODN-transfected cell was 60. 28% while that in oligofectamine-transfeeted control cell 17.34%. Conclusion hTERT antisense oligodeoxynucleotide can increase the sensitivity of pancreatic cancer cell to gemeitabine. The mechanism may be due to the down-regulated expression of hTERT mRNA and telomerase activity.
出处
《中华医学杂志》
CAS
CSCD
北大核心
2009年第34期2391-2394,共4页
National Medical Journal of China
基金
江苏省常州市科技计划（CS2004214）
