低氧条件下Stat3反义寡核苷酸对喉癌细胞Hep-2的化疗增敏作用

摘要

Objective To investigate changes in apoptosis and cell cycle progression in Hep-2 cells after treating them with cisplatin and antisense oligonucleotides targeting the signal transducer and activator of transcription 3 (Stat3). Methods Hep-2 cells were transfected with different concentrations of liposome-encapsulated Stat3 sense or antisense oligonucleotides.The cells were then exposed to different cisplatin doses (0-5μg/ml)in normoxic and hypoxic environments,and their sensitivity was measured in the MTT survival assay.Cell cycle progression and apoptosis were examined by flow cytometry. Results Apoptosis increased with increasing doses of cisplatin in both hypoxic and normoxic conditions.Rate of apoptosis was similar between cells treated with 1μg/ml DDP under hypoxic conditions and untreated control cells under normoxic conditions(P>0.05).However,cells treated with 3 or 5 μg/ml cisplatin under hypoxic conditions showed a lower apoptosis rate than cells under normoxic conditions(P<0.01).Transfecting cells with Stat3 antisense oligonucleotides decreased Hep-2 survival under hypoxic conditions in a dose-dependent manner (100-400 nmol/L)and led to cell cycle blockade at G0/G1 phase.The rate of apoptosis in cultures treated with Stat3 antisense oligonucleotides and cisplatin together was significantly different from the rates in control cultures and in cultures treated with sense oligonucleotides and cisplatin together, antisense nucleotides on their own,or cisplatin on its own(P<0.01). Conclusions Treating Hep-2 cells with Stat3 antisense oligonucleotides and cisplatin together can induce apoptosis under hypoxic conditions and increase sensitivity to chemotherapy.Gene therapy involving RNA interference targeting Stat3 may be a new approach to adjuvant chemotherapy for laryngeal carcinoma.%目的：探讨低氧条件下信号转导子和转录激活子3（signal transducer and activator of transcription 3，Stat3）反义寡核苷酸（antisense oligonucleotide，AS-ON）联合化疗对Hep-2细胞凋亡及细胞周期的影响。方法以脂质体法将Stat3 AS-ON转染Hep-2细胞，应用MTT法检测不同浓度的AS-ON转染后Hep-2细胞存活率的变化以及低氧对Hep-2细胞化疗敏感性的影响；应用流式细胞术检测低氧条件下Stat3 AS-ON联合化疗对Hep-2细胞凋亡及细胞周期的影响。结果Hep-2细胞凋亡率在顺铂浓度为0~5μg/ml范围内随药物浓度上升而增加，以1μg/ml顺铂作用低氧组与常氧组Hep-2细胞凋亡率无差异（P>0.05），3μg/ml、5μg/ml顺铂同等浓度下低氧组Hep-2细胞凋亡率低于常氧组（P<0.01）。细胞存活率随转染Stat3 AS-ON浓度的增加而下降，但不能完全抑制细胞增殖。低氧条件下Stat3 AS-ON联合化疗组的细胞周期与对照组相比G0/G1增加；低氧条件下Stat3 AS-ON+DDP组细胞凋亡率与空白对照组、Stat3 S-ON+DDP组、Stat3 AS-ON组、DDP组比较差异有统计学意义（P<0.01）。结论 Stat3 AS-ON转染Hep-2细胞联合化疗在低氧条件下可诱导肿瘤细胞凋亡，增加化疗敏感性。靶向Stat3的干扰技术有望成为喉癌基因治疗的新策略，为临床辅助化疗探索一条新途径。