Abstract: Objective To explore high concentrations of urea-induced human brain microvascular endothelial cells (HBMECs) to produce inflammatory cytokines and possible mechanism. Methods HBMECs were incubated in high concentrations of urea or mannitol(as osmotic control) for 3，6，12 and 24 hours. Expressions of TNF-a and iNOS were observed by Immunofluorescence. Western blot analysis was employed to assess the protein expressions of TNF-a, iNOS, COX-2, NF-κB/P65 and p-P65. NO concentration was determined by a commercial NO assay kit. Results Immunofluorescence showed high positive immunostaining of TNF-a and iNOS after high concentrations of urea stimulued compared with control group .The protein expressions of TNF-a, COX-2 and p-P65 were significantly increased at 3 and 6 hours after high urea treatment (p<0.01), and iNOS were continued to increase from 3 to 24 hours (p<0.01).Moreover, NO content was increased at 3 hours after high urea treatment (p<0.05). Conclusion High concentrations of urea can induce HBMECs to produce inflammatory cytokines.

Key words: urea, HBMECs, TNF-a, NO, NF-κB