The Apoptosis Effectiveness Of Combined Use Of High Glucose And High Palmitic Acid On HUVECs

Objective To evaluate the apoptosis effectiveness and the mechanism of combined use of high glucose with high palmitic acid on human vessel endothelial cells.Methods High glucose and high palmitic acid were applied, alone or together, to human umbilical vein endothelial cells ECV-304. Flow cytometer (FCM) and fluorescent staining with hoechst33342 were used to detect apoptosis effectiveness of EVC304. HPLCMS (High Performance Liquid Chromatography / Mass Spectrometry) was used to detect the amounts changes of intracellular ceramide. At the same time, the changes of Caspase-3 activity were detected by microplate spectrophotometer.Results Cell apoptosis could be induced no matter high glucose and high palmitic acid were used alone or together. The apoptosis rate was higher in the group of high glucose+high palmitic acid than that in the group of high glucose or high palmitic acid alone (p<0.05) . The amounts of intracellular ceramide were higher in the group of high glucose+ high palmitic acid and group of high glucose or high palmitic acid used alone than in the control group, most obviously in the group of combined use (P<0.05) . The inhibitor of ceramide synthetase myriocin could obviously decrease the apoptosis rate induced by high glucose+ high palmitic acid (P<0.05) . The Caspase-3 activity of EVC304 was higher in the group of high glucose+ high palmitic acid and group of high glucose or high palmitic acid used alone than in the control group, most obviously in the group of combined use (P<0.05) . The inhibitor of Caspase-3 zDEVD-fmk could obviously decrease the apoptosis rate induced by high glucose+ high palmitic acid (P<0.05 ) . Myriocin could obviously inhibit the change of Caspase-3 activity induced by high glucose+ high palmitic acid (P<0.05) .Conclusion The combined use of high glucose and high palmitic acid can promote cell apoptosis synergistically. This process of synergistic apoptosis includes ceramide and caspase-3. Caspase-3 appeared to function downstream of ceramide activation in this process.