Epigenetics may affect the susceptibility for type 2 diabetes mellitus (T2DM). Previously, our studies have shown that the hypomethylation of human miR-375 promoter may contribute to the pathogenesis of T2DM. However, the methylation pattern of miR-375 promoter in T2DM is not yet fully understood. In this study, the DNA methylation status of the different region of miR-375 promoter in Chinese Han population with T2DM were explored. 100 Han patients with T2DM and 100 Han healthy controls with normal glucose tolerance (NGT) were collected. Then the transcription level of pre-miR-375 and mature miR-375 were examined using quantitative real-time PCR and the methylation status of 27 CpG sites in the miR-375 promoter was determined by MassARRAY Spectrometry. The relative expression of mature miR-375 was shown as fold difference relative to miR-16 (3.0-fold, P=0.0260) and pre-miR-375 was markedly unregulated (2.6-fold, P=0.0415) in Han T2DM samples. Aberrant methylation was significantly higher within the amplicon of the miR-375 promoter in T2DMs than in NGTs, an average of 10.27% and 7.24% (P=0.0004; Figure 3A), respectively. Further, one CpG unit (CpG_26.27) was significantly hypermethylated in T2DM samples compared with NGT. Together, our results highlights for the first time that aberrant hypermethylation is a common event in Han T2DM, suggesting that the aberrant methylation of the CpG sites within miR-375 promoter may serve as a potential candidate biomarker for T2DM in the Chinese Han population.