LncRNA DLEU2通过自噬机制调节甲状腺乳头状癌细胞增殖与凋亡

LncRNA DLEU2 regulates the proliferation and apoptosis of papillary thyroid carcinoma cells through autophagy mechanism

目的:探讨LncRNA DLEU2是否通过自噬影响甲状腺乳头状癌(papillary thyroid carcinoma,PTC)细胞增殖和凋亡。方法:免疫组织化学和实时荧光定量聚合酶链反应(quantitative real-time PCR,qRT-PCR)技术方法检测17对甲状腺乳头状癌及癌旁组织Beclin-1和ATG7的表达;qRT-PCR检测DLEU2在TPC-1细胞与正常甲状腺细胞中的表达;以Lipofectamine TM 3000转染试剂分别将DLEU2沉默载体(shDLEU2)及对照载体(shNC)转染至甲状腺乳头状癌TPC-1细胞系,实验分为:空白对照组(无转染的TPC-1细胞)、DLEU2沉默组(TPC-1细胞转染shDLEU2)、阴性对照组(转染照载体shNC)。利用生长曲线、划痕实验及Transwell检测TPC-1细胞的增殖、迁移和侵袭能力;流式细胞术检测shDLEU2对TPC-1凋亡的影响;Western blot和免疫细胞化学染色检测DLEU2对TPC-1细胞Beclin-1、ATG7和LC3表达的影响。结果:免疫组织化学染色结果显示:ATG7与Beclin-1在甲状腺乳头状癌高表达(P<0.05),shDLEU2组TPC-1细胞增殖、迁移、侵袭能力下降(P<0.05),Western blot显示沉默DLEU2基因,Beclin-1与ATG7的表达量增加,LC3 A/B比值减小(P<0.05)。结论:沉默DLEU2基因,可能通过自噬促进TPC-1的凋亡,抑制其增殖、迁移和侵袭。

Objective:To explore whether LncRNA DLEU2 affects the proliferation and apoptosis of thyroid papillary carcinoma cells through autophagy.Methods:Detecting expression of Beclin-1 and ATG7 in 17 pairs of papillary thyroid carcinomas and paracancerous tissues by immunohistochemistry and quantitative real-time PCR(qRT-PCR).DLEU2 expression in TPC-1 cells and normal thyroid cells were detected by qRT-PCR.The DLEU2 silencing vector(shDLEU2)and control vector(shNC)were transfected into the thyroid papillary carcinoma TPC-1 cell line by Lipofectamine TM 3000 transfection reagent.The experiment was divided into a blank control group(non transfected TPC-1 cells),a DLEU2 silencing group(shDLEU2 transfected TPC-1 cells),and a negative control groups(shCN transfected TPC-1 cells).Using growth curves,scratch experiments,and Transwell to detect the proliferation,migration,and invasion abilities of TPC-1 cells.Flow cytometry was used to detect the effect of silencing DLEU2 on the apoptosis of TPC-1.Western blot and immunocytochemistry were used to detect the effect of DLEU2 on the expression of Beclin-1,ATG7,and LC3 in TPC-1 cells.Results:IHC results showed that ATG7 and Beclin-1 were highly expressed in papillary thyroid carcinoma(P<0.05).In silencing DLEU2 group,the proliferation,migration,and invasion ability of TPC-1 cells decreased(P<0.05).Western blot showed silencing of DLEU2 gene,increased expression of Beclin-1 and ATG7,and decreased LC3 A/B ratio(P<0.05).Conclusion:Silencing the DLEU2 gene may promote apoptosis of TPC-1 and inhibit its proliferation,migration and invasion through autophagy.