MiR-760 Mediated The Proliferation And Metastasis Of Hepatocellular Carcinoma Cells By Regulating HMGA2

Objective This study aimed to detect the expression and biological significance of miR760 in HCC cells,to identify the target gene of miR-760,and to provide a new way for exploring the roles of miR-760 in HCC carcinogenesis.It offers new insight into selecting diagnostic and prognostic biomarkers as potential therapeutic targets.Methods(1)Funrich software was used to determine that miR-760 plays an important role in the occurrence and development of HCC.(2)miR-760 expression in LO2 and SMMC-7721,HepG2,HCCLM3,and Huh7 cells was determined by real-time quantitative PCR(qRT-PCR).miR-760 mimics were transiently transfected with Lippo2000 to make it highly expressed,and the corresponding negative control(NC mimics)was transfected as the negative control.The effects of miR-760 overexpression on the proliferation,apoptosis,migration,and invasion of HCC cells were analyzed by in vitro experiments.The effect of miR-760 overexpression on the growth of xenograft tumors was evaluated by in vivo tumorigenesis experiment in nude mice.(3)HMGA2 was used as the predictive target gene for miR-760 by TargetScan、miRDB and StarBase databases.The targeting relationship between miR-760 and HMGA2 was analyzed using luciferase activity reporter assay,qRTPCR,and Western blot.The effect of PIRH2 on the tumor-suppressive effects of miR-320a on HCC cells was analyzed by in vitro experiments,such as MTT,Edu,colony formation assay,and transwell cell assay.(4)The methylated level of CpG sites of miR-760 was evaluated by MSP in HCC tissues and adjacent tissues.(5)The effect of SP1 on the expression level of miR-760 was verified by qRT-PCR.The binding of SP1 to miR-760 promoter was verified by ChIP assays and dual-luciferase assay.Results(1)DEmiRNAs in patients with LIHC were identified from the TCGA-LIHC dataset.And miR-760 is closely related to hepatocellular carcinoma.(2)Compared with LO2 cells,we found miR-760 expression was reduced in SMMC-7721,HepG2,HCCLM3,and Huh7 cells.Up-regulation of miR-760 observably inhibits cell proliferation,migration,invasion,and the growth of xenograft tumors in HepG2 and Huh7 cells.(3)Dual-luciferase reporter assay exhibited HMGA2 is a direct target gene of miR-760.Experimental validation displayed that HMGA2 expression was suppressed by miR-760 on protein and mRNA levels.(4)miR-760 suppresses proliferation and migration by targeting HMGA2.(5)DNA methylation may result in the downregulation of miR-760.(6)Transcription factor SP1 could inhibit the transcription of miR-760.Conclusions Our study displayed that miR-760 was markedly reduced in HCC cells,and determined the tumor-suppressive role of miR-760 in dramatically suppressing proliferation,migration,invasion,and the growth of xenograft tumor in HCC cells.We confirmed that miR-760 inhibited proliferation and migration by HMGA2.Besides,DNA methylation and SP1 could be resulted in decreased miR-760 expression.In summary,our study demonstrated that SP1/miR-760/HMGA2 may serve as a molecular regulatory axis for HCC treatment.