Role Of LncRNA SNHG11in HBV-related HCC

BackgroundHepatocellular carcinoma (HCC) is one of the most common malignant tumor ofAsian people, with high malignant degree and poor prognosis. Hepatitis B virus (HBV)infection could significantly increase the risk of suffering from HCC. By regulatingspecific long non-coding RNA, HBV infection may affect the tumorigenesis ofHBV-related HCC. However, little is known about the mechanism.ObjectiveWe intend to investigate the mechanism through which HBV infection regulatesspecific long non-coding RNA, subsequently affects the incidence of HCC and theprognosis of patients with HBV-related HCC.Methods1、Analysis of long non-coding RNA (LncRNA) microarray in HCC tissuesTo investigate transcription of LncRNA and related mRNA in HCC tissues, LncRNAand mRNA microarray analyses were performed using ten HHC specimens whichcontained five HBV-related HCC and five HCC without HBV. Bioinformatics Impactanalysis was performed to select mRNAs which had a biologically significant and relevantrole in pathway level, using Pathway Express software online. The LncRNAs correlatedwith selected mRNAs were selected for further analysis.2、Verification of Gene microarray result and analysis of clinical outcomesTo verify the reliability of LncRNA/mRNA microarray results, qRT-PCR wasapplied to detect expression of LncRNA SNHG11and CLDN1in both carcinoma andpara-carcinoma tissues. By tissue microarray and immunohistochemical staining, weinvestigated the expression change of Claudin-1(CLDN1coding protein) of carcinomaand para-carcinoma tissues in218patients. And then survival analysis was appliedlooking for the relationship of Claudin-1and HBV-related HCC.3、Analysis of lncRNA SNHG11’s function in HBV-related HCCTo analysis the function of LncRNA SNHG11in HBV-related HCC, over-expression plasmid and siRNA were applied to regulate transcription of LncRNA SNHG11in HCC cell line. Then we investigated cell proliferation, migration andinvasion by CCK-8assay, wound healing assay and transwell assay. Real-time PCR,western blot was applied to analyse regulation of CLDN1by LncRNA SNHG11.Results1、22LncRNAs and40mRNAs were found specially expressed in HCC tissuescompared with corresponding noncarcinoma tissues. Combining with bioinformaticsanalysis, we found that5of these40mRNAs belong to the Claudin family（includingCLDN1, CLDN2, CLDN10, CLDN14and CLDN19）, and the biological informationimpact factors of them were significantly higher than other mRNAs. Besides, thetranscription trend of lncRNA SNHG11was negative correlatd with the trend of CLDN1.2、By qRT-PCR, We found that Gene microarray results were with high reliability.Through tissue microarray and immunohistochemical staining, we affirmed the ubiquity ofdifferential expression of Claudin-1in HCC. In further analysis we found that in patientsof HBV-related HCC，the expression of Claudin-1was associated with the size of tumor.Claudin-1was identified as an independent prognostic factor (OS, hazard ratio (HR)=0.625,95%confidence interval (CI):1.001-3.484, P=0.050; DFS, HR=0.874,95%CI:1.370-4.192, P=0.002) in HBV-related HCC patients with HBV replication.3、We selected HCC cell line HepG2.2.15which was stably transfected HBV gene.Transcription of LncRNA SNHG11in HepG2.2.15was analyzed by qRT-PCR.Down-regulation of LncRNA SNHG11by siRNA resulted in elevation of CLDN1expression. Cell proliferation rate, migration ability were both enhanced after LncRNASNHG11down-regulation. And up-regulation of LncRNA SNHG11by plasmid couldreduce cell migration ability.ConclusionVariational transcription of lncRNAs in HCC tissues indicated their crucial roles inHBV-related HCC. Our research showed that LncRNA SNHG11could alter HepG2.2.15cells proliferation and migration by regulating CLDN1.