The Preliminary Research On The Role Of Renin-angiotensin System In Oxidizing LDL In Raw264.7 Cells

Objective: Atherosclerosis(AS)is the leading cause of high morbidity and mortality in people with cardiovascular disease(CVD).AS is a chronic inflammatory disease caused by the accumulation of low density lipoprotein(LDL)under the endothelium and oxidized to ox-LDL,causing an inflammatory reaction,and then taken up by macrophages and developed into foam cells.The Renin-angiotensin system(RAS)plays an important role in the pathogenesis of CVD.Blocking the RAS pathway reduces AS plaque progression and ischemic events.This experiment was carried out in three parts,aiming to explore the role of RAS in the active oxidation of LDL in Raw264.7 cells,thus providing a new research direction for the oxidation of LDL,and providing new ideas for the treatment of clinical diseases.Method:1.Culture Raw264.7 cells,LDL was stimulated at different times: ACE activity assay kit to detect changes in intracellular ACE activity;Ang II kit to measure Ang II expression;Real-Time PCR to detect ACE m RNA expression;Real-Time PCR and Western Blot to measure the expression of AT2;the purpose was to investigate the changes of RAS during the oxidation of LDL by macrophages Raw264.7.2.Raw264.7 cells were cultured,and Ang II and Ang IV were stimulated at different times: Western Blot was used to detect the expression of FABP5.The purpose of this study was to investigate the effect of RAS on the expression of FABP5 in macrophages Raw264.7.3.After stimulating the cells with ACE inhibitor(Captopril,captopril)for different time,it was divided into three groups: captopril stimulation,LDL stimulation,LDL+captopril stimulation : PLA2 Total activity photometric quantitative detection kit was used to determine the change of PLA2 activity in cells;Western Blot was used to detect the expression of FABP5;and whether RAS involved in oxidized LDL was involved in Raw264.7 macrophages.4.Stimulate cells with LDL,LDL+captopril,captopril,Ang II,Ang IV for 24 h,then collected the cell culture supernatant and cells,determine the amount of Malonaldehyde(MDA)contained in them,and determine the treatment methods.The degree of lipid peroxidation of Raw264.7 cells.Results:1.After LDL stimulated Raw264.7 cells,the activity of ACE increased,the production of Ang II increased,and the expression of ACE m RNA and AT2 m RNA and protein increased(P<0.05).2.After stimulated Raw264.7 cells by Ang II and Ang IV,FABP5's expression was increased.In the LDL-stimulated group,the activity of PLA2 was increased at 12 min,and the expression of FABP5 increased at 3 h and 5 h.3.In the LDL and captopril-stimulated group,the activity of PLA2's activity was decreased at 12 min,and FABP5' expression was decreased at 5 h;in the captopril-stimulated group,there was no change in the activity of PLA2 and the expression of FABP5 compared with the control group.4.In the cells,compared with the control group,LDA,Ang II,Ang IV treated Raw264.7 cells for 24 h,MDA content increased(P<0.05),LDL+captopril,captopril treated cells for 24 h,There was no significant change in MDA content;compared with LDL group,MDA content decreased after LDL+captopril treatment(P<0.05).In the cell culture supernatant,compared with the control group,LDA,LDL+captopril,Ang II,Ang IV treated Raw264.7 cells for 24 h,MDA content increased(P<0.05),and MDA after captopril treatment There was no change in the content;compared with the LDL group,the LDA+captopril treatment decreased the MDA content(P<0.05).Conclusion: LDL can activate RAS,RAS can improve FABP5's expression level in Raw264.7 cells,and the increase of FABP5 expression during LDL oxidation is achieved by RAS.The subsequent step may be through the binding of heterodimers formed by PPARs and RXR?,and then to PPRE on the target gene to stimulate the oxidative stress pathway and promote the oxidation of LDL.