Objective To explore the chronic effects of excess glucose on INS-1E cells and isolated mouse islets. Methods Islets were isolated by the collagenase digestion from adult female NMRI mice (6-10 weeks) after induction of anesthesia by phenpbarbital. INS-1E cells and isolated mouse islets were cultured in RPMI 1640 supplemented with 11.1 or 25.0 mmol/L glucose for 72 h, respectively. Then the medium were changed to Krebs-Ringer buffer containing 3.3 or 16.7 mmol/L glucose and incubated for 1 h. Correspondingly upper medium was collected for insulin assay. Total RNA was extracted from INS-1E cells after 72 h incubation with different concentration of glucose, then cDNA was synthesized and mRNA expressions of Pdx1, insulin 1, insulin 2, and GLUT2 were determined by RT-PCR. Results After incubation with excess glucose, basal insulin secretion was increased in both INS-1E cells and isolated mouse islets [INS-1E cells: (10.47±0.78) vs (7.71±0.59) ng/10000 cells, P＜0.01; isolated mouse islets: (3.85±0.26) vs (2.18±0.21) mg/L, P＜0.001], and glucose-stimulated insulin secretion was impaired [INS-1E cells: (17.11±1.98) vs (30.76±2.20) ng/10000 cells, P＜0.001; isolated mouse islets: (14.78±1.03) vs (20.46±1.49)mg/L, P＜0.01]. Pdx1, insulin 1, insulin 2, and GLUT2 mRNA levels were reduced in INS-1E cells after treatment of excess glucose. Conclusion Long-term exposure to supraphysiologic glucose concentrations may cause dysfunction of pancreatic islet cells.