Qin C-F, Zhao F-L
Hernia and Abdominal Surgery Department, Beijing Chao-Yang Hospital, Capital Medical University, Beijing, China.
Eur Rev Med Pharmacol Sci. 2017 May;21(10):2377-2384.
This paper aimed to investigate the effect of long non-coding RNA TUG1 (lncRNA TUG1) on cell proliferation, as well as cell migration in pancreatic cancer.
The mRNA levels of Taurine-up-regulated gene 1 (TUG1) in three kinds of pancreatic cancer cells BxPC3, PaTu8988 and SW1990 was detected by RT-qPCR. Meantime, RT-qPCR was used to examine the mRNA levels of TUG1 in 20 cases of human pancreatic cancer tissues and its para-carcinoma tissues. pCDH-TUG1 plasmid and its empty plasmid pCDH were transfected into BxPC3 and PaTu8988 cells to up-regulate TUG1 expression. siRNA targeting TUG1 and the control siRNA were transfected into SW1990 cells to down-regulate TUG1 expression. Cell clone formation and CCK-8 assay were used to detect the cell proliferation capacity. Transwell assay was used to evaluate cell migration capacity. Western blot was applied to examine the protein expressions of MMP2, MMP9, E-cadherin, Smad 2, Smad 3, p-Smad 2, p-Smad 3, TGF-β and TGF-βR. RT-qPCR was used to detect the levels of MMP2 and MMP9.
The results showed that TUG1 was differentially expressed in the three kinds of pancreatic cancer cells, among which the expression level of SW1990 was relatively high, and the expression levels of BxPC3 and PaTu8988 were relatively low. TUG1 had more expression in pancreatic cancer tissues than that in para-carcinoma tissues. After the up-regulation of TUG1, cell proliferation and migration capacities were increased, protein levels of MMP2 and MMP9 were increased and protein level of E-cadherin was declined. Conversely, after down-regulation of TUG1 expression, cell proliferation and migration capacities were weakened, protein levels of MMP2 and MMP9 were decreased and protein level of E-cadherin was increased. In addition, over-expressed TUG1 could promote Smad2 and Smad3 phosphorylation, but Smad2 and Smad3 phosphorylation were weakened after down-regulated expression of TUG1. The protein expression of TGF-β and TGF-β receptor were more in the TUG1 overexpression group than that in the control group, while the result was just opposite after TUG1 expression was down-regulated.
These data suggest that lncRNA TUG1 may enhance the proliferation and migration of pancreatic cancer cells through EMT pathway.
本文旨在研究长链非编码RNA TUG1(lncRNA TUG1)对胰腺癌细胞增殖及细胞迁移的影响。
采用RT-qPCR检测三种胰腺癌细胞BxPC3、PaTu8988和SW1990中牛磺酸上调基因1(TUG1)的mRNA水平。同时,运用RT-qPCR检测20例人胰腺癌组织及其癌旁组织中TUG1的mRNA水平。将pCDH-TUG1质粒及其空质粒pCDH转染至BxPC3和PaTu8988细胞,以上调TUG1表达。将靶向TUG1的siRNA及对照siRNA转染至SW1990细胞,以下调TUG1表达。采用细胞克隆形成实验和CCK-8实验检测细胞增殖能力。运用Transwell实验评估细胞迁移能力。通过蛋白质印迹法检测基质金属蛋白酶2(MMP2)、基质金属蛋白酶9(MMP9)、E-钙黏蛋白、Smad 2、Smad 3、磷酸化Smad 2(p-Smad 2)、磷酸化Smad 3(p-Smad 3)、转化生长因子-β(TGF-β)及转化生长因子-β受体(TGF-βR)的蛋白表达。采用RT-qPCR检测MMP2和MMP9的水平。
结果显示,TUG1在三种胰腺癌细胞中表达存在差异,其中SW1990表达水平相对较高,BxPC3和PaTu8988表达水平相对较低。TUG1在胰腺癌组织中的表达高于癌旁组织。上调TUG1后,细胞增殖及迁移能力增强,MMP2和MMP9蛋白水平升高,E-钙黏蛋白蛋白水平降低。相反,下调TUG1表达后,细胞增殖及迁移能力减弱,MMP2和MMP9蛋白水平降低,E-钙黏蛋白蛋白水平升高。此外,过表达TUG1可促进Smad2和Smad3磷酸化,但下调TUG1表达后,Smad2和Smad3磷酸化减弱。TUG1过表达组中TGF-β及TGF-β受体的蛋白表达高于对照组,而下调TUG1表达后结果相反。
这些数据表明lncRNA TUG1可能通过上皮-间质转化(EMT)途径增强胰腺癌细胞的增殖和迁移。