MiR-144Target ROCK1to Inhibit Invasion And Metastasis In Hepatocellular Carcinoma

Objective:To study the relationship between miR-144target ROCK1(Rho-associated, coiled-coil-containing protein kinase1) and invasion and metastasis in Hepatocellular carcinoma (HCC). To investigate the feasibility of using bioinformatics to predict target miRNA.Methods:The expression of ROCK1in specimens of42of HCC,28of normal hepatic tissue beside HCC were detected by immunohistochemistry. Taeget miRNAs of ROCK1were predicted by informatics methods. And to obtain the miRNA which have the best stability and specificity. The binding between miR-144and ROCK1were detected by Quantificational real-time polymerase chain reaction (qRT-PCR). And the relationship between miR-144and metastasis of HCC were also analysized by qRT-PCR. The expression of ROCK1were detected by western blot after miR-144target to ROCK1. After down regulation of ROCK1by miR-144, the ability of invasion and metastasis were analysized by Transwell in HCC.Results:(1)17target miRNAs of ROCK1were obtained by bioinformatics methods, and miR-144was the one which target ROCK1with the best stability and specificity.(2) The positive ratio of ROCK1in HCC and normal normal hepatic tissue beside HCC were76.2%(32/42),28.6%(8/28), respectively. There were significance between them (P<0.05).(3) There were low expression of ROCK1and high expression of miR-144in HCC cell line with low metastasis potentiality. And there high expression of ROCK1and low expression of miR-144in HCC cell line with high metastasis potentiality. The data indicate that miR-144downregulate expression of ROCK1and cause metastasis of HCC.(4) There were high expression of miR-144and down regulation of ROCK1in HepG2-miR-144mimics. And there were no difference among HepG2, HepG2-miR-Scramble, HepG2-miR-144-PM about the expression of miR-144and ROCK1. The data showed that miR-144could downregulate expression of ROCK1by target to ROCK1mRNA.(5) There were significantly lower cells counts by Transwell cave experiment in HepG2-miR-144mimics than others(P<0.05). And there were no significantly difference among HepG2, HepG2-miR-Scramble, HepG2-miR-144-PM (P>0.05). the data showed that enhance the level of miR-144could down regulate expression of ROCK1and decrease the metastasis ability of HCC cells in vitro.Conclusion:(1) There were a lot of miRNAs could binding to ROCK1. And miR-144was the one which target ROCKl with the best stability and specificity(2) miR-144target to the3’UTR of ROCK1and cause down regulation of ROCK1and decrease the invasion and metastasis of HCC.