Effects Of Chrysin On Human Aortic Endothelial Cell Injury Induced By High Glucose And High Fat

Objective:1.In order to establish HAEC cell injury model induced by high glucose and high fat,HAEC were cultured in high glucose and high fat medium with suitable concentration of palmitic acid.2.To observe the protective effects and mechanism of Chrysin on HAEC cell injury induced by high glucose and high fat.Methods:1.First,the HAECs were incubated with different concentrations of SPA(0.1-0.4mmol·L^-1)and GLU(30mmol·L^-1)for 24 hours.The survival rate of HAECs was detected by MTT method,and the appropriate concentration of SPA was selected.Then the cytotoxicity of Chrysin to HAECs was determined by incubation for 4 hours with different concentrations of CHR(1,10,20?mol·L^-1).The HAECs were pretreated with CHR-L(1?mol·L^-1)and CHR-H(10?mol·L^-1)for 4 hours,and then incubated with 30mmol·L^-1HG+0.1mmol·L^-1 SPA for 24 hours to induce HAEC cell injury,in order to observe the protective effects of Chrysin.2.The HAECs were divided into CON group,HG+SPA group,CHR-L(1?mol·L^-1)group and CHR-H(10?mol·L^-1)group.The LDH release rate,NO concentration in culture medium and intracellular ROS production of HAECs were measured.The expressions of endoplasmic reticulum stress-related proteins GRP78,IRE1?,PERK,ATF6,and inflammation,adhesion related proteins were detected by Western Blot.Results:1.The high glucose concentration was 30 mmol·L^-1,and the survival rate of HAECs decreased with the increase of SPA concentration.When the SPA concentration was 0.1mmol·L^-1,the survival rate of HAECs was?45.79±5.16?%,then we selected 0.1mmol·L^-1as the concentration of SPA for modeling.The MTT results showed that the survival rate of HAEC decreased significantly after incubation with 20?mol·L^-1 CHR for 4 hours.Then the subsequent experiments were divided into CHR-L(1?mol·L^-1)group and CHR-H(10?mol·L^-1)group.Pre-incubation of CHR(1?mol·L^-11 and 10?mol·L^-1)could significantly inhibit the decrease of survival rate of HAECs induced by high glucose and high fat,which indicated that CHR could protect the HAECs from injury induced by high glucose and high fat.2.HG+SPA incubation resulted in increased LDH release,decreased NO release,increased intracellular ROS production in HAECs.The expression of GRP78,IRE1,PERK and ATF6,TXNIP/NLRP3,and ICAM-1 were significantly up-regulated by high glucose and high fat.Pre-incubation of CHR(1?mol·L^-11 and 10?mol·L^-1)decreased LDH release,increased NO release,decreased intracellular ROS production,and significantly reduced the expression levels of ERS marker protein,TXNIP/NLRP3 and ICAM-1 in HAECs.Conclusion:High glucose and high fat could induce HAEC cell injury.In this experiment,30mmol·L^-1 GLU+0.1mmol·L^-1 SPA was used as the model condition.Chrysin(1?mol·L^-1 and 10?mol·L^-1)pre-incubated for 4 hours could reduce HAEC cell damage caused by high glucose and high fat,could increase cell survival rate,decrease LDH release and increase NO release.Pre-incubation of CHR decreased expression of adhesion molecules.Chrysin protects HAEC from injury induced by high glucose and high fat via inhibiting endoplasmic reticulum stress,reducing ROS production,and then down-regulating the expression of TXNIP/NLRP3 induced by oxidative stress.