Effects Of High Glucose On Necroptosis And IL-6 Expression In Cultured Human Umbilical Vein Endothelial Cells

Objective: To investigate the expression of RIP1, RIP3, IL-6 in the cultured human umbilical vein endothelial cells(HUVEC) through the experim in vitro. To explore the role and mechanism of necroptosis in the injury of HUVEC.Methods: Human umbilical vein endothelial cells(HUVEC) in culture were randomly divided into 3 groups according to different managements: normal concentration of glucose as controlled group(DMEM containing 5.5mmol/L glucose), mannitol group(DMEM containing 5.5mmol/L glucose + 24.5mmol/L mannitol), high glucose group(DMEM containing 30mmol/L glucose). The expression of VIII in the HUVECs was detected by immunofluorescence. The expression of m RNA of RIP, RIP3 and IL-6 were respectively detected by Real-Time PCR, immunohistochemical and Western Blot.Results: 1.immunofluorescence: The VIII were mainly deposited on the cell cytoplasm of that the cell is identified as HUVECs. 2.RT-PCR: The expression of RIP, RIP3 and IL-6 m RNA in high glucose group were apparently increased than those in normal group and mannitol group(P<0.05). However their expression had no significant difference between the normal glucose group and mannitol group(P>0.05). There were significantly Positive correlation between the expression of IL-6、RIP1 and RIP3 respectively(r=0.961, P<0.001; r=0.801,P=0.010). 3.immunohistochemical: The RIP, RIP3 and IL-6 were mainly deposited on the cell cytoplasm of HUVECs. Compared with the normal glucose group, the expression of RIP, RIP3 and IL-6 in high glucose group were apparently increased than those in normal group and mannitol group(P<0.05). But their depositions had no significant difference between the normal glucose group and mannitol group(P>0.05). There were significantly Positive correlation between the expression of IL-6、RIP1 and RIP3 respectively(r=0.755, P=0.019; r=0.810, P=0.008). 4.Western Blot: The expression of RIP, RIP3 and IL-6 in high glucose group were apparently increased than those innormal group and mannitol group(P<0.05). However their expression had no significant difference between the normal glucose group and mannitol group(P>0.05). There were significantly Positive correlation between the expression of IL-6, RIP1 and RIP3 respectively(r=0.908, P<0.001; r=0.966, P<0.001).Conclusion: High concentration of glucose increased the expression of RIP1 and RIP3 suggesting that the increase of necroptosis in the cultured HUVECs. The interaction of necroptosis and IL-6 may promote the inflammatory reaction and the injury of HUVECs.