热休克蛋白70反义寡核苷酸诱导PC-3m凋亡及抑制生长的研究

The study of HSP70 antisense oligomer inducing apoptosis and inhibiting proliferation of PC-3m

目的 探讨热休克蛋白 70 (HSP70 )与雄激素不敏感前列腺癌细胞生长的关系。 方法 用形态学、流式细胞术、琼脂糖凝胶DNA电泳及Westernblot方法观察 0 16 μmol/L人工合成的HSP70反义寡核苷酸作用 0 10 0h后 ,对体外培养的人雄激素不敏感前列腺癌细胞株PC 3mHSP70表达的抑制及其对生长和凋亡的影响。 结果 10 μmol/LHSP70反义寡核苷酸作用 48h后 ,West ernblot显示PC 3m无HSP70表达且琼脂糖凝胶DNA电泳可见阶梯状DNA片段 ;HE染色出现凋亡细胞 ;台盼蓝拒染测定的生长抑制率和流式细胞术分析的凋亡率与HSP70反义寡核苷酸呈剂量、时间依赖性关系 ;细胞周期分析表明生长抑制和凋亡主要发生在G1期和S期。 结论 HSP70为正常状况下雄激素不敏感前列腺癌细胞生长增殖所必需 ;HSP70反义寡核苷酸通过抑制HSP70表达而诱导PC 3m凋亡并抑制其生长 ;HSP70可能是一种调控前列腺癌细胞凋亡的新基因 ,可作为基因及药物治疗前列腺癌的新靶向。

Objective To investigate the relation of HSP70 with growth and survival of the androgen-insensitive prostate cancer cell. Methods The effects of HSP70 antisense oligomer (0-16μmol/L treatment for0-100h) on HSP70 expression levels,the proliferation and apoptosis of human androgen-insensitive prostate cancer cell line PC-3m in vitro were studied by Western blot,morphology,agarose gel DNA electrophoresis and flow cytometry. Results A ladder-like pattern of DNA fragments on agarose gel electrophoresis and no HSP70 expression in Western blot were observed at 10μmol/L for 48h.Occurrence of apoptotic cells observed on HE staining, the in hibiting proliferation rate on trypan blue dye exclusion test and the induced apoptosis rate through flow cytometry analyses revealed the apoptosis and inhibition of PC-3m accurred mainly at G 1 and S phase which and were dose-and-time dependent. Conclusions The results indicate that HSP70 is required for androgen-insensitive prostate cancer cell to proliferate and survive under normal condition, and HSP70 antisense oligomer can induce apoptosis and inhibit the growth of PC-3m by abrogating its HSP70 expression,moreover,HSP70 might be a new gene modulating the apoptosis of prostatic cancer cells and may be used as a novel target for the gene and drug therapy of prostatic carcinoma.