A Study About The Protective Effect Of Proglumide-ADT-OH Condensation On Injury Of Human Umbilical Vein Endothelial Cells Induced By Ox-LDL

Objective:The aim of this study was to investigate whether the Proglumide-ADT-OH condensation has a protective effect on HUVEC damage model and whether the protective mechanism is related to the regulation of JAK/STAT pathway and NF-кB pathway.Methods:Proglumide-ADT-OH condensation as H₂S donor was synthesized by condensation of Proglumide and ADT-OH.HUVECs were cultured to detect release of H₂S from Proglumide-ADT-OH.The HUVEC damage cell model was established by inducing human umbilical vein endothelial cells（HUEVC）with 80μg/ml oxygenized low density lipoprotein（ox-LDL）for 24h.The content of total cholesterol（TC）and free cholesterol（Fch）levels were detected by the total cholesterol（TC）and free cholesterol（Fch）kits CCK-8 kits were used to detect the effect of the essential oil at different concentration and time on the cell proliferation of HUVECs.The effect of the Proglumide-ADT-OH condensation on the secretion of IL-6,IL-10 and TNF-αin HUVEC damage model was measured by ELISA kits of IL-6,IL-10 and TNF-α.SOD kit was used to detect the activity of SOD intracellular.Western blotting was used to detect the expression of p-NF-кB p65 protein,p-JAK2 protein and p-STAT3 protein.Results:The Proglumide-ADT-OH condensation was synthesized successfully by MS and NMR.Proglumide-ADT-OH release the max of H₂S is at 100μmol/L after acted on HUVEC for 24h.The oil red O staining showed that a large amount of red staining granules appeared in the cells after HUVECs were induced with 80μg/ml ox-LDL for 24 hours and Fch/TC was not less than 50%,which indicated that the HUVEC damage model was successful.The cell proliferation was best after the Proglumide-ADT-OH at 100μmol/L acted on HUVECs for 12hours.The ELISA assay showed that the Proglumide-ADT-OH could significantly reduce the level of IL-6 and TNF-α（P<0.01）by regulating the NF-кB signaling pathway and JAK/STAT signaling pathway,and promote the production of IL-10（P<0.05）in the HUVEC damage model by regulating the NF-кB signaling pathway,as well as increase the intracellular superoxide dismutase activity of SOD（P<0.05）.Western blotting results showed that Proglumide-ADT-OH condensation can down regulate the expression of p-NF-кB p65 protein by regulating the NF-кB signaling pathway（P<0.05）and down regulate the expression of p-JAK2protein by regulating the JAK/STAT signaling pathway（P<0.05）.Conclusion:The Proglumide-ADT-OH condensation has shown the protective effect against ox-LDL induced HUVEC damage model,and the protective mechanism is related to the regulation of JAK/STAT pathway and NF-кB pathway.