RNA干扰下调YAP基因对甲状腺癌细胞TPC-1功能的影响

Effect of RNA interference targeting YAP gene on thyroid cancer TPC-1 cells

关键词:  甲状腺肿瘤  Yes相关蛋白  siRNA  细胞功能

Key words:  thyroid cancer  Yes associated protein  siRNA  cellular function

摘要:

目的 检测siRNA沉默Yes相关蛋白(Yes associated protein,YAP)后的人甲状腺癌TPC-1细胞功能变化。方法 应用阳离子脂质体转染试剂LipofectamineTM2000将靶向沉默YAP基因的siRNA序列转染至甲状腺癌TPC-1细胞,应用qRT-PCR、Western印迹法检测转染后TPC-1细胞YAP基因及蛋白表达,MTT、平板克隆实验、Transwell小室、划痕实验和流式细胞术分别检测转染对细胞增殖、侵袭、迁移和细胞周期及凋亡的影响。结果转染siRNA后,YAP mRNA和蛋白表达明显下降(P<0.05)；TPC-1细胞的增殖、侵袭、迁移能力受到抑制,细胞周期G₀/G₁阻滞,细胞凋亡未明显上升。结论 抑制甲状腺癌细胞YAP表达可有效降低细胞的增殖、迁移和侵袭能力,改变细胞周期分布,但对细胞凋亡无明显影响。

Abstract:

Objective To investigate the effect of small interfering RNA (siRNA) targeting Yes associated protein (YAP) gene on thyroid cancer TPC-1 cells. Methods siRNA targeting YAP gene was transfected in thyroid cancer TPC-1 cells with LipofectamineTM2000. qRT-PCR and Western blotting were conducted to determine YAP suppression at mRNA and protein levels by siRNA. MTT assay, colony formation assay, Transwell migration assay wound healing assay and flow cytometry were used to study the effects of siRNA on cell proliferation, migration, cell cycle and apoptosis, respectively. Results Expression of YAP was suppressed after interference of YAP siRNA (P<0.05), proliferation, migration and invasion of TPC-1 cells were inhibited; cells were arrested at G₀/G₁ phase, but cellular apoptosis rate was not significantly changed. Conclusion The results indicate that YAP gene exerts a vital role in biological behaviors of thyroid cancer TPC-1 cells.