shRNA 干扰长链非编码RNA PVT1 对甲状腺乳头状癌IHH-4细胞增殖、凋亡和周期的影响

目的:探究沉默长链非编码RNA PVT1 对甲状腺乳头癌IHH-4 细胞增殖、凋亡和周期的影响。方法:慢病毒转染PVT1 shRNA(sh-PVT1),RT鄄PCR 检测PVT1 的表达,CCK8 检测细胞增殖倍数,流式检测细胞凋亡及细胞周期,Westernblot 检测细胞Ki67、Caspase-3 和Cyclin A 的表达;复制裸鼠甲状腺乳头状癌肿瘤移植模型,记录肿瘤生长情况,Western blot 检测肿瘤组织Ki67、Caspase-3 和Cyclin A 的表达。结果:sh-PVT1 转染细胞24 h 后,IHH-4 细胞PVT1 的表达水平明显降低(P<0.01);转染细胞4 d 后,细胞增殖倍数显著降低,增殖标志蛋白Ki67 表达明显被抑制(P<0.05,P<0.01);同时,sh-PVT1 能显著升高IHH-4 细胞凋亡率,诱导凋亡标志蛋白Caspase-3 表达(P<0.01);此外,sh-PVT1 还可诱导细胞G2/ M 期阻滞,抑制周期蛋白Cyclin A 的表达(P<0.05,P<0.01)。干扰PVT1 表达能显著抑制甲状腺乳头瘤生长(P<0.01),下调肿瘤组织Ki67 和Cyclin A 的表达水平(P<0.01),诱导Caspase-3 表达(P<0.01)。结论:沉默PVT1 表达能显著抑制甲状腺乳头癌IHH-4 细胞增殖且诱导细胞凋亡和周期阻滞。

Effect of silencing long non-coding RNA PVT1 with shRNA on cell proliferation,apoptosis and cycle of papillary thyroid carcinoma cell line IHH-4

Objective:To investigate the effect of silencing long non-coding RNA PVT1 on cell proliferation,apoptosis andcycle of papillary thyroid carcinoma cell line IHH-4.Methods: Cells were transferred with PVT1 shRNA(sh-PVT1) by lentiviruses,and the expression of PVT1 in IHH-4 cell was measured by RT-PCR.CCK8 assay was performed for proliferation.Apoptosis and cellcycle were determined by flow cymetory.Western blot was employed to detect protein levels of proliferation,apoptosis and cell cycle-related proteins.Established a papillary thyroid carcinoma nude model and observing tumor growth.The protein levels of Ki67,Caspase-3 and Cyclin A were determined by Western blot.Results: The expression of PVT1 was inhibited after cells transferred with sh-PVT1for 24 h(P<0.01).Cell proliferation folds was decreased by sh-PVT1 after cells transferred with shRNA for 4 d and the expression ofKi67 also was inhibited by sh-PVT1 significantly(P<0.05,P<0.01). Meanwhile,sh-PVT 1 increased cell apoptosis rate and proteinlevel of Caspase-3(P<0.01).In addition,sh-PVT1 arrested cell cycle at G2/ M stage and makedly decreased Cyclin A expression(P<0.05,P<0.01).sh-PVT1 inhibited the growth of papillary thyroid tumor(P<0.01),down-regulated the expression levels of Ki67 andCyclin A and increased the protein level of Caspase-3(P<0.01). Conclusion: Silencing PVT1 inhibits proliferation,induces apoptosisand cell cycle arrest of papillary thyroid carcinoma cell line IHH-4.