【摘要】 目的STAT5信号转导途径在慢性粒细胞白血病的发病中有十分重要作用,本研究应用诱骗寡核苷酸抑制STAT5信号转导途径,诱导白血病K562细胞凋亡,并初步探讨其中的分子机制。方法体外设计合成的STAT5诱骗寡核苷酸,在脂质体的介导下转染K562细胞,以Annexin-V/PI双染实验和电镜检查细胞凋亡,用RT-PCR和Westernblot方法检测STAT5下游基因bcl-xL、c-myc的表达。结果Annexin-V/PI双染实验检测诱骗寡核苷酸作用24h后,早期凋亡细胞即有14.57%±1.08%;电镜检查观察到诱骗寡核苷酸作用引起细胞凋亡的超微结构形态学改变;RT-PCR实验和Westernblot实验证实诱骗寡核苷酸作用引起STAT5下游基因bcl-xL和c-mycmRNA和蛋白表达下调。结论STAT5诱骗寡核苷酸能诱导白血病K562细胞凋亡,其作用机制可能与诱骗寡核苷酸下调STAT5下游凋亡相关基因的表达相关。更多还原

【Abstract】 Objective STAT5 is critical for CML transformation. We try to use a decoy oligodeoxynucleotid(ODN) inhibiting STAT5 signal pathway of K562, then observe the effect of the decoy ODNs on apoptosis and investigate a possible related molecular mechanism. Methods A decoy ODN against STAT5 was designed and synthesized in vitro , was transfected into K562 cells mediated by cationic lipid. Annexin-V/PI staining method calculated apoptotic cells by FCM. Electron microscope observed the apoptosis cells. Then,RT-PCR method was used to observe the effects of STAT5 decoying ODNs, on the expression of bcl-xL and c-myc mRNA. Western blot technique was used to determine the effect of STAT5, decoy ODNs, on the expression of bcl-xL and c-myc protein. Results It was treated with decoy ODNs for 24 h, FCM-Annexin-V/PI dual staining method determined the percentage of early apoptotic cells was 14. 57±1. 08%. Electron microscope also determined the microcosmic changes of apoptotic cells. RT-PCR test indicated that STAT5, decoy ODNs, had decreased the mRNA expression of bcl-xL and c-myc. Western blot test also determined decrease of protein expressions of both bcl-xL and c-myc. Conclusion STAT5 decoying ODNs could induce the apoptosis of K562 cells. The expressions of of both bcl-xL and c-myc mRNAs as well as the proteins are all decreased after treatment with decoying ODN, and it may be the cause of apoptosis.更多还原