Guo X-F, Wang A-Y, Liu J
Department of Infectious Liver Diseases, Zaozhuang Municipal Hospital, Zaozhuang, Shandong, China.
Eur Rev Med Pharmacol Sci. 2016 Jul;20(14):3011-6.
In this study, we investigated whether miR-33a downregulation in HCC is a result of hypoxia-inducible factors (HIFs) overexpression. Then, we further studied the regulative effects of miR-33a on Twist1 and their regulation in HCC cell invasiveness.
Human hepatocellular cancer (HCC) cell lines (HepG2 and BEL-7402) were transfected with miR-33a mimics, HIFs siRNA or Twist1 siRNA. MiR-33a level was measured using QRT-PCR. The binding between miR-33a and Twist1 3'UTR was verified using Western blot analysis and dual luciferase assay. E-cadherin and N-cadherin expression levels were detected by western blot analysis. Tumor cell invasion was assessed using transwell assay.
MiR-33a downregulation in HCC cells is hypoxia-induced and is a result of HIFs upregulation. HIF-1α and HIF-2α suppression partly rescued miR-33a expression under hypoxia. Both HepG2 and BEL-7402 cells with miR-33a overexpression had significantly decreased E-cadherin expression and increased N-cadherin level. Transwell analysis confirmed that miR-33a overexpression significantly suppressed the tumor cell invasion capability. Twist1 is a direct target of miR-33a in HCC. HepG2 cells with Twist1 knockdown had significantly increased E-cadherin, decreased N-cadherin and suppressed invasion capability.
MiR-33a downregulation in HCC cells is hypoxia-induced and is a result of HIFs upregulation. MiR-33a can modulate EMT and invasion of hepatocellular cancer cells at least partly via downregulating Twist1.
在本研究中，我们调查了肝癌中miR-33a表达下调是否是缺氧诱导因子（HIFs）过表达的结果。然后，我们进一步研究了miR-33a对Twist1的调控作用及其在肝癌细胞侵袭中的调节作用。
用miR-33a模拟物、HIFs siRNA或Twist1 siRNA转染人肝癌（HCC）细胞系（HepG2和BEL-7402）。使用QRT-PCR检测miR-33a水平。通过蛋白质印迹分析和双荧光素酶测定验证miR-33a与Twist1 3'UTR之间的结合。通过蛋白质印迹分析检测E-钙黏蛋白和N-钙黏蛋白的表达水平。使用Transwell试验评估肿瘤细胞侵袭。
肝癌细胞中miR-33a表达下调是缺氧诱导的，是HIFs上调的结果。在缺氧条件下，HIF-1α和HIF-2α的抑制部分挽救了miR-33a的表达。miR-33a过表达的HepG2和BEL-7402细胞均有E-钙黏蛋白表达显著降低和N-钙黏蛋白水平升高。Transwell分析证实，miR-33a过表达显著抑制肿瘤细胞侵袭能力。Twist1是肝癌中miR-33a的直接靶标。Twist1敲低的HepG2细胞E-钙黏蛋白显著增加，N-钙黏蛋白减少，侵袭能力受到抑制。
肝癌细胞中miR-33a表达下调是缺氧诱导的，是HIFs上调的结果。miR-33a至少部分通过下调Twist1来调节肝癌细胞的上皮-间质转化和侵袭。
