Roles Of MiR-26a And MiR-125b In Regulating Gender Disparity Of Hepatocellular Carcinoma

As a major form of liver cancer,Hepatocellular carcinoma(HCC)presents a significant male-dominant feature on both cancer incidence and mortality.MicroRNAs(miRNAs)are a class of small non-coding RNA molecules contain:ing approximately 22 nucleotides and play important roles in regulating genes at their post-transcriptional levels.It has been well known that miRNAs are involved in HCC initiation,progression and metastasis.However,little is known about their roles in regulating HCC gender disparity.We first screened miRNAs with differential expression between female and male non-tumors as well as between tumor and non-tumors from HCC patients,and miRNAs related to HCC overall survival.Our analysis revealed that two candidates,i.e.,miR-26a and miR-125b,were gender-,tumor-,and HCC survival-related.In HCC non-tumor tissues,miR-26a and miR-125b expressed higher levels in female compared to male.Both miRNAs were significantly down-regulated in tumors compared to non-tumor tissues from HCC patients.Moreover,HCC patients with higher levels of miR-26a or miR-125b in tumors had better overall survival and lower recurrence rates.We further validated these results in mice model.Consistently,in livers of C57BL/6 mice,miR-26a and miR-125b had higher levels in female's liver compared to male's.In the N-diethylnitrosamine(DEN)-induced HCC mouse model,miR-26a and miR-125b also showed female-dominant expression in non-tumor tissues,and expressed lower levels in DEN-induced HCC tumors compared to non-tumor liver tissuesFurther survival analysis revealed that HCC patients with high expression of miR-26a and miR-125b had the longest overall survival and lowest recurrence rates,in comparison to patients with low miR-26a and miR-125b,and patients with low level of either one.We then performed gene expression profiling comparison between HCC patients with low levels of miR-26a/miR-125b and patients with high levels of miR-26a/miR-125b.Between these two groups,a significant difference was noticed on their gene expression profiles,indicating the difference on their tumor biology.Cellular biology assays further revealed that co-expression of miR-26a and miR-125b in HCC cells could significantly inhibit colony formation,wound healing and proliferation.TargetScan and MICRORNA.ORG were conducted to predict the target genes of miR-26a and miR-125b.In 3' untranslated regions(UTRs)of IL-6 and IL6ST,there were binding sites for miR-26a.The binding sites of miR-125b were found in 3'UTR of IL-6R and STAT3.Through dual luciferase assay,we found that IL-6 was a bona fide target of miR-26a in HCC cells.However,over-expressed miR-26a and miR-125b did not seem to affect the activation of IL-6/STAT3 pathway.We'll continue to explore the roles of miR-26a and miR-125b in the IL-6 signaling pathway in futureWe also found that the expression of miR-26a and miR-125b was significantly correlated in HCC-related tissues from either mice or human.The promoter region analysis of miR-26a gene revealed that miR-26a had the distinct proximal promoter from its host gene.Both miRNA array data and qPCR data in HCC tissues and HCC cell lines showed the discordant expression of miR-26s and their host genes.Experimentally,we found that miR-26 family members were independently transcribed from their host genes,via their independent promoter regions.Furthermore,transcriptome integration analysis and bioinformatics analysis discovered that discordant expression of intronic miRNAs and their host genes was relatively ubiquitous in HCC.These results suggest that female-biased miR-26a and miR-125b could inhibit HCC malignant features,providing new insights into HCC gender-disparity.The transcription of miR-26 family members was independent from their host genes,which improved our understanding on miRNA transcriptional mechanisms.