Chronic venous insufficiency (CVI) is a common disease characterized by structural and functional abnormalities of the venous system. Until recently, the pathogenesis of CVI remains largely unknown. MicroRNAs (miRNAs) are a family of endogenous small non-coding RNAs emerged as post-transcriptional gene repressors and play essential roles in diverse pathological processes including vascular disease. However, their roles in CVI have not been elucidated. In this study, we employed oligonucleotide microarrays to perform a genome-wide miRNAs profiling in the great saphenous vein (GSV) tissues of patients with CVI. Our results revealed a total of 14 miRNAs that are expressed differentially in GSV tissues. Among them nine miRNAs were found significantly up-regulated, while five miRNAs were down-regulated significantly. Real-time RT-PCR verified statistically consistent expression of three selected miRNAs (miR-34a, miR-155 and miR-202) with microarrays analysis. These three miRNAs, which were described as crucial regulators in many biological processes and vascular diseases, might also play important roles in CVI. Functional annotation of target genes of differentially expressed miRNAs via bioinformatics approaches revealed that these predicted targets were significantly enriched and involved in several key signaling pathways important for CVI, including mitogen-activated protein kinase pathways, pathways in cancer, apoptosis, and cell cycle, and p53 signaling pathways. In summary, miRNAs might involve in multiple signaling pathways contributing to the pathological processes of CVI. These data may provide fundamental insights into the molecular basis of CVI, which may aid in designing novel approaches for prevention and treatment of this complex disease.