MiR-224 Executes a Tumor Accelerative Role during Hepatocellular Carcinoma Malignancy by Targeting Cytoplasmic Polyadenylation Element-Binding Protein 3

Abstract

Purpose: The purpose of our study was to probe the mechanism of how miR-224/cytoplasmic polyadenylation element-binding protein 3 (CPEB3) axis is concerned with hepatocellular carcinoma (HCC).

Methods: The expressions and prognostic values of miR-224 and CPEB3 in HCC patients were analyzed based on the data acquired from the TCGA and GEO databases. qRT-PCR was conducted to test the mRNA expression levels of miR-224 and CPEB3. The expression level of miR-224 in SMMC-7721/HuH-7 cells was up-/downregulated by miR-224 mimic/inhibitor to explore its influence on HCC cell proliferation and motility by utilizing CCK8 and transwell assays, respectively. Luciferase activity assay was applied for verifying the target of miR-224. The relationship between miR-224 and CPEB3 was analyzed utilizing Pearson's correlation coefficient. The protein level of CPEB3 was tested by Western blotting. Rescue assay was performed to determine whether CPEB3 involved in the process of HCC cell phenotype changes caused by miR-224 alteration.

Results: MiR-224 was highly expressed and CPEB3 was lowly expressed in HCC tissues. Besides, the high expression of miR-224 and low expression of CPEB3 were correlated with worse prognosis in HCC patients. Up-/downregulation of miR-224 accelerated/restrained SMMC-7721/HuH-7 cell proliferation and motility. CPEB3 was predicted and proofed as a target gene of miR-224. We discovered that CPEB3 was negatively modulated by miR-224. We also found a sharply negative correlation between CPEB3 and miR-224. Using rescue assay, we showed that overexpression of CPEB3 suppressed the proliferation and motility of SMMC-7721 cells with overexpressed miR-224, while knockdown of CPEB3 facilitated the proliferation and motility of HuH-7 cells with downregulated miR-224.

Conclusion: Our data provided evidences that miR-224 is implicated in HCC cell proliferation and motility via targeting CPEB3. The relationship between miR-224 and CPEB3 might be a novel finding, and miR-224/CPEB3 axis might be markers for providing therapeutic and prognostic information in HCC.

Keywords: Biomarker; Invasion; Migration; Proliferation; Rescue assay; Target.