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RNAi 抑制GSK-3β 基因表达增强卵巢癌 SKOV3细胞对紫杉醇敏感度的研究[J]. 肿瘤防治研究, 2011, 38(03): 247-249. DOI: 10.3971/j.issn.1000-8578.2011.03.002
引用本文: RNAi 抑制GSK-3β 基因表达增强卵巢癌 SKOV3细胞对紫杉醇敏感度的研究[J]. 肿瘤防治研究, 2011, 38(03): 247-249. DOI: 10.3971/j.issn.1000-8578.2011.03.002
Enhancing Sensibility of Ovarian Cancer Cell Line SKOV3 to Paclitaxel by Inhibiting Gene GSK-3β with RNA Interference[J]. Cancer Research on Prevention and Treatment, 2011, 38(03): 247-249. DOI: 10.3971/j.issn.1000-8578.2011.03.002
Citation: Enhancing Sensibility of Ovarian Cancer Cell Line SKOV3 to Paclitaxel by Inhibiting Gene GSK-3β with RNA Interference[J]. Cancer Research on Prevention and Treatment, 2011, 38(03): 247-249. DOI: 10.3971/j.issn.1000-8578.2011.03.002

RNAi 抑制GSK-3β 基因表达增强卵巢癌 SKOV3细胞对紫杉醇敏感度的研究

Enhancing Sensibility of Ovarian Cancer Cell Line SKOV3 to Paclitaxel by Inhibiting Gene GSK-3β with RNA Interference

  • 摘要: 目的构建GSK-3β基因特异性短发卡状(shRNA)干扰载体,观察RNA干扰技术对卵巢癌SKOV3细胞GSK-3β基因表达的抑制作用,并且探讨促进紫杉醇药物敏感度的机制。 方法设计合成并构建针对GSK-3 β mRNA的特异性shRNA干扰质粒,将干扰质粒转染SKOV3细胞。筛选稳定转染的单细胞扩大培养,应用RT-PCR、Western blot方法检测各组细胞GSK-3β表达抑制情况。流式细胞学检测法(FACS)检测细胞凋亡率。比较抑制GSK-3β 基因前后卵巢癌SKOV3细胞抵抗紫杉醇药物的差异。结果成功构建pGSK3β-shRNA 干扰载体。SKOV3 细胞转染pGSK3β-shRNA干扰载体后,RT-PCR、Western blot方法显示,转染后GSK-3βmRNA 和蛋白表达均显著下降。FACS检测结果显示,转染pGSK3β-shRNA干扰载体,不同浓度紫杉醇作用24h后,卵巢癌SKOV3细胞对紫杉醇敏感度都有显著性提高。细胞凋亡率显著增加(P<0.05)。结论运用RNAi 技术构建的pGSK3β-shRNA干扰载体能特异、高效抑制靶基因GSK-3β表达;降低卵巢癌细胞中GSK-3 β基因表达水平,能增强卵巢癌细胞对紫杉醇的敏感度,促进肿瘤细胞凋亡。

     

    Abstract: ObjectiveTo consrtuct the specific short hairpin interfering vector of gene GSK-3β and study its suppressing effect to understand mechanisms of enhancing sensibility of ovarian cancer cell line SKOV3 to taxol. MethodsDesign and construct specific short hairpin interfering vector of gene GSK-3β and transfect it into SKOV3 cell line for screening stable cell lines. Detect RNAi suppressing efficacy using RT-PCR and Western blot.Compare the different apoptosis ratio and cell sensibility to taxol before and after interfering gene GSK-3β by FACS. ResultsThe interfering vector pGSK3β-shRNA was constructed successfully.The expression of mRNA and protein of gene GSK-3β in stable cell line transfected with vector pGSK3β-shRNA was obviously lower than that in cells before transfected by RT-PCR and Western blot.Moreover, FACS results demonstrated the apoptosis ratio after 24h treatment of taxol was obviously increased and sensitivity was better in cells after transfected than before with significant difference(P<0.05). ConclusionThe constructed vector pGSK3β-shRNA had a specific and high effective inhibiting effect on gene GSK-3β. It could enhance the sensibility of ovarian cancer cell line SKOV3 to taxol and promote tumor cell apoptosis by inhibiting gene GSK-3β.

     

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