AIM: To investigate the reciprocal modulation between microRNA (miRNA) and DNA methylation via exploring the correlation between miR-373 and methyl-CpG-binding domain protein (MBD)2. METHODS: MiR-373 expression was examined using the TaqMan miRNA assay. Methylation of miR-373 was investigated using methylation-specific polymerase chain reaction, and recruitment of methyl binding proteins was studied using the chromatin immunoprecipitation assay. Mutation analysis was conducted using the QuikChange (TM) Site-Directed Mutagenesis kit. The activity of miR-373 gene promoter constructs and targeting at MBD2-three prime untranslated region (3'UTR) by miR-373 were evaluated by a dual-luciferase reporter gene assay. RESULTS: In hilar cholangiocarcinoma, miR-373 decreased and was closely associated with poor cell differentiation, advanced clinical stage, and shorter survival. The promoter-associated CpG island of miR-373 gene was hypermethylated and inhibited expression of miR-373. MBD2 was up-regulated and enriched at the promoter-associated CpG island of miR-373. Methylation-mediated suppression of miR-373 required MBD2 enrichment at the promoter-associated CpG island, and miR-373 negatively regulated MBD2 expression through targeting the 3'UTR. CONCLUSION: MiR-373 behaves as a direct transcriptional target and negative regulator of MBD2 activity through a feedback loop of CpG island methylation. (c) 2012 Baishideng. All rights reserved.