Aim To observe the influence of glucose on the polyol pathway in human vascular endothelial cells and to investigate its potential mechanisms. Methods Human umbilical vein endothelial cells were cultured in vitro with glucose at different concentrations or for different cultural time. The level of nitric oxide (NO) and sorbitol, aldose reductase (AR) AR activity and the expression of AR mRNA level were detected with high performance liquid chromatography (HPLC), nitrate reductase method,biochemical assay and reverse transcription polymerase chain reaction (RT PCR). Results The levels of sorbitol in high glucose groups were higher than in control group (p<0.05 or p<0.01), but the levels of NO in high glucose groups were lower than in control group (p<0.05 or p<0.01). Both AR mRNA expression and its activity were dependent on glucose concentrations or cultural times (p<0.05 or p<0.01) AR mRNA expression and its activity with glucose at 22 mmol/L for 48 h or at 44 mmol/L for 24 h did not increase accordingly, but began to decrease(p<0.05). Conclusion Glucose can affect the function of endothelial cells such as the increasing prodction of sorbitol and the decreasing systhesis of NO. The mechanisms may be that glucose can upregulate AR gene expression and enhance its activity, then activates the polyol pathway in endothelial cells.