| Objective To investigate the effect of glucose, tumor necrosis factor alpha and interleukin-1 beta on the expression of EPCR mRNA of human umbilical vein endothelial cells(ECV304), as well as the effect of troglitazone, a thiazolidinedione compound, as a potential therapeutic agent.Methods The human umbilical vein endothelial cells(ECV304) were cultured at 370C , 5% CO2 humidified environment and changed the culture medium every 2 days. We harvested the cells when the cells had grown to 80%-90% confluence, and confirmed the expression of EPCR in ECV304 in protein and mRNA levels by flow cytometry (FM) technique and reverse transcription-polymerase chain reaction (RT-PCR) technique respectively. ECV304 were incubated with or without high concentrate glucose ,TNF-α, IL-1βand troglitazone, to observe the effect on the expression of EPCR mRNA of ECV304.Group 1: The cells were cultured in the RPMI-1640 medium for 48h to 90% confluence, with a change of the medium after 24h. The cells were cultured with medium without newborn calf serum for 24h. The cells were then cultured with starvation medium (with 3% newborn calf serum) continually and stimulated with glucose (5mmol/L,10mmol/L,30mmol/L,50mmol/L) at 370C for 24h. The cells were incubated to different time points(0.5h, 3h, 6h, 12h, 24h) with starvation medium with 50mmol/L glucose. The cells of control group was received only from the fresh starvation medium without glucose. All samples were performed in triplicate.Group 2: The cells were cultured in the RPMI-1640 medium for 48h to 90% confluence, with a change of the medium after 24h. The cells were cultured with medium without newborn calf serum for 24h. The cells were then cultured with starvation medium (with 3% newborn calf serum) continually and stimulated with recombinant human TNF-α(2ng/ml, 10ng/ml, 25ng/ml) at 370C for 12h. The cells...
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