Hax1 siRNA对黑素瘤A375细胞化疗敏感性的影响.doc

Hax1 siRNA对黑素瘤A375细胞化疗敏感性的影响
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作者:李文彬冯捷, 闫小宁, 张彩晴
【摘要】目的: 探讨Hax1 siRNA对黑素瘤A375细胞化疗敏感性的影响。方法: MTT法、 AnnexinVFITC/PI染色、 Hoechst 33258染色、 Western blot检测Hax1 siRNA联合顺铂或苦参碱对细胞增殖、凋亡和caspase3活性形式表达的影响。结果: 相同药物浓度作用下, Hax1 siRNA转染组细胞抑制率、凋亡率及cleaved caspase3表达明显高于未转染组和转染无关载体组(P ); 荧光显微镜下观察, 相同药物浓度作用下, Hax1 siRNA转染组细胞凋亡现象较明显。结论: Hax1 siRNA增强顺铂或苦参碱对黑素瘤A375细胞的增殖抑制和诱导凋亡作用, 该作用可能和caspase3的活性形式表达增强相关。
【关键词】 siRNA; Hax1; 黑素瘤; 化疗敏感性
[Abstract] AIM: To investigate the effects of Hax1 siRNA on sensitivity to chemotherapy of melanoma A375 cells. METHODS: The effects of Hax1 bined with cisplatin(DDP) or Matrine(Mat) on proliferation, apoptosis of A375 cells and active forms of caspase
3 were detected with MTT assay, Annexin VFITC/PI staining with FCM, Hoechst 33258 staining, and Western blot respectively. RESULTS: The inhibition of proliferation , induction of apoptosis and the expression of active caspase3 forms in DDP or Mat treated A375 cells transfected with Hax1 siRNA were all significantly higher than those in untransfected A375 cells or A375 cells transfected with control siRNA (). The distinctive apoptotic morphology was observed in Mat or DDP treated A375 cells transfected with Hax1 siRNA. CONCLUSION: The data from our current study indicate that Hax1 siRNA could enhance the effects of DDP or Mat on inhibition of melanoma A375 cell growth and induction apoptosis, which may be caused by the increased expression of active forms of caspase3.
[Keywords]siRNA; Hax1; melanoma; chemotherapy sensitivity
黑素瘤病程中晚期或具有高危因素的早期病变应予以全身辅助化学治疗, 但临床资料显示黑素瘤细胞对化疗药物相对不敏感, 且化疗的毒副作用大, 患者难以耐受, 常常导致治疗的失败。凋亡抑制是肿瘤对化疗药物产生耐药的原因之一, 增加肿瘤细胞对化疗药物诱导凋亡的敏感性从而逆转肿瘤化疗耐药, 是目前逆转肿瘤耐药的手段之一。Hax
1是近年发现的一个凋亡抑制蛋白[1]。我们前期研究已证实, 通过RNAi技术下调基因Hax1能明显诱导A375细胞发生凋亡, 并抑制细胞的增殖[2]。本研究探讨Hax1是否和肿瘤对化疗药物的耐药有关, 观察了Hax1 siRNA转染后, 顺铂、苦参碱对A375细胞作用的影响。
1 材料和方法
材料恶性黑素瘤细胞株A375细胞由西安交通大学第二医院皮肤科保存。DMEM培养基购自美国Invitrogen公司; 胎牛血清(FBS)购自杭州四季青生物工程公司; 载体pSU