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Background: Red cell distribution width (RDW) is a predicter of infections, cancer and diabetes. However, the relationship between RDW and β-cell function and insulin resistance remains unclear in patients with type 2 diabetes mellitus (T2DM). The aim of the study was to explore the relationship between RDW and β-cell function in patients with T2DM. Methods: A total of 559 T2DM patients were enrolled in this cross-sectional study. Patients were divided into three groups according to RDW tertiles. Clinical and biochemical characteristics such as age, duration of diabetes, blood pressure, RDW, glycosylated hemoglobin A1c (HbA1c), C-peptide and lipid profiles were collected. Homeostasis model assessment of insulin resistance (HOMA2IR) and homeostasis model assessment of β-cell function (HOMA2%B) were assessed using homeostasis model assessment (HOMA) based on fasting blood glucose (FBG) and fasting C-peptide index (FCPI). Correlations and multiple linear regressions were performed to explore the association between RDW and islet function indexes in total population and in different gender subgroups. Results: The HOMA2%B gradually increased according to RDW tertiles (lowest, second, highest RDW tertiles; 47.1(32.9–75.4), 54.05(34.1–81), and 57.9(38.65–95.4), respectively; P = 0.036). Correlation analysis indicated that there were significant correlations between RDW and age, diabetes duration, diastolic blood pressure (DBP), triglycerides (TG), aspartate transaminase (AST), FBG, HbA1c and HOMA2%B in all subjects. In male subjects, RDW correlated positively with age, high-density lipoprotein cholesterol (HDL) and AST, and it correlated negatively with body mass index (BMI), DBP and TG. In female subjects, RDW correlated positively with age, duration, serum creatinine (Cr), FCPI and HOMA2%B, and it correlated negatively with alanine transaminase (ALT), FBG and HbA1c. Multiple linear regressions indicated that RDW was significantly correlated with HOMA2%B and HbA1c in the total population in both unadjusted and adjusted analysis. This finding could be reproduced in the subgroup of men for HOMA2%B only and in women for HbA1c only. Conclusions: RDW is associated with β-cell function assessed by HOMA2%B after adjusting for covariates in male T2DM patients. [ABSTRACT FROM AUTHOR]