Study On The Function Of MicroRNA-125b

BackgroundMicroRNAs (miRNAs) belong to a large family of non-coding, single-stranded RNAs,18-25nucleotides long. There is growing evidence that miRNAs regulate the expression ofprotein-coding genes, either at the level of messenger RNA degradation or translation, bypairing with their target mRNA's3'UTR. In1993, the first miRNA lin-4was discovered.They are involved in many biological processes including cell cycle regulation,development, differentiation, metabolism, and ageing. MiRNAs' dysregulation has beenobserved in many human malignancies and there is some evidence for their involvement inthe progression of tumors, either as oncogenes or tumor suppressors.HCC is one of the most prevalent cancers and the leading cause of cancer-related deathsworldwide. Several miRNAs are up-regulated or down-regulated in the tumor, some actingas modulators of cell growth, apoptosis, migration or invasion. However, their exactbiological role in HCC remains unclear.AimsMiR-125b is one of the recently identified tumor-associated miRNAs. Existing datasuggest that dysregulation of miR-125b is involved in tumorigenesis and progression ofmany types of cancer, including HCC. Previous research showed downregulation ofmiR-125b in human HCC, so we studied the function of miR-125b in the HCC cells byoverexpressing it.Methods1. We examined the expression of miR-125b in9paired samples of clinical HCC tumor,adjacent normal liver tissues and6HCC cell lines using quantitative R-T PCR analysis.2. We constructed a eukaryotic expression vector of miR-125b and obtained HCC cellswith overexpression miR-125b by lentivirus packaging and cell infection.3. Methods including MTT, colony formation assays, tumor formation in nude mice, flowcytometry and staining with hoechst33342were used to measure cells proliferation and apoptosis.4. We investigated the direct target gene of miR-125b by a dual-luciferase reporter system,Western blot and immunohistochemical assay.Results1. We found that the expression of miR-125b is substantially lower in5out of the6HCCcell lines and in7out of the9paired samples of clinical HCC tumors.2. MiR-125b inhibits HCC cell proliferation in vitro and in vivo, and miR-125b inducesHCC cell apoptosis.3. Western blot results show decreased expression of Bcl-2protein in MHCC97H cellswith overexpression of miR-125b than control, this is confirmed by immunohistochemicalassay data.4. We confirm Bcl-2is a direct target of miR-125b using a dual-luciferase reporter system.In conclusion, we report the altered miR-125b expression pattern in HCC, investigatethe potential role of miR-125b in tumorigenesis, and demonstrate that Bcl-2is a targetgene of miR-125b. Our data demonstrate an important role for miR-125b in the molecularetiology of cancer and suggest its potential application in cancer therapy.