Study On The Role Of Nrf2 In Myeloid Cell In The Development Of Atherosclerosis

Objective:Atherosclerosis is the major reason of myocardial infarction and stroke,which poses a huge threat to people life and health.Atherosclerosis occurs when vascular endothelial cells are damaged.Monocytes enter the endothelial cells and differentiate into macrophages,which phagocyte oxidized low density lipoprotein（LDL）and form foam cells that accumulate in the intima,forming lipid lines.Macrophages play a vital role in atherosclerosis not only in the uptake of oxLDL,but also in the formation of foam cells.Nuclear transcription factor NF-E2 related factor 2（Nrf2）regulates transcription expression of antioxidant enzymes and detoxifies enzymes against oxidative stress.Previous studies have shown that in mouse model of atherosclerosis,the degree of atherosclerosis was reduced after Nrf2 systemic knockout.Atherosclerosis is aggravated by Nrf2-specific knockout in mononuclear cells which obtained from bone marrow transplantation,but the specific regulatory mechanism of Nrf2 is not clear.Therefore,in this study,a mouse model of atherosclerosis with Nrf2 specific deletion in myeloid monocytes was established to study the role and mechanism of Nrf2 in the occurrence and development of atherosclerosis.Methods:1.Establishment of a mouse model of atherosclerosis with specific deletion of Nrf2in myeloid monocytesUsing Nrf2-LoxP;ApoE^-/-and Lysozyme2-Cre;ApoE^-/-tool mice to obtain myeloid cell Nrf2 knockout(Nrf2（M）-KO;ApoE^-/-)mice and littermate-born controls(Nrf2-LoxP;ApoE^-/-)mice.Male and female mice were given normal diet for 26-28weeks,and body weight,blood glucose,serum,plasma and tissues samples of various important organs were collected at the end of the diet.2.Mice sample treatmentThe content of lipoprotein in plasma were detected by the kit.The aorta fixed with 4%paraformaldehyde was stained with oil red,and the aortic root was embedded with paraffin and cryo-embedment.H&E staining,immunohistochemical staining and oil red staining were performed.3.Extraction and detection of primary peritoneal macrophages from mice with Nrf2-deficient myeloid cellsPrimary peritoneal macrophages were extracted from homologous and homologous Nrf2（M）-KO and control mice.Oxidative low density lipoprotein（Ox LDL）cells were given for a certain time and dose to detect the mRNA expression levels of Nrf2 and related inflammatory factors and lipoprotein receptors.The chemotaxis of primary peritoneal macrophages by CCl2 was detected by Transwell.Results:1.In the context of ApoE gene deletion,Nrf2 specific deletion in myeloid monocytes had no effect on body weight,blood glucose,blood pressure,blood lipid and other basic indexes of mice.2.In the background of ApoE gene deletion,Aortic oil red staining showed increased plaque area（P<0.05）,aortic root H&E staining showed intimal thickening,and immunohistochemical staining showed increased macrophage infiltration in mice with Nrf2-specific deletion of myeloid mononuclear cells.3.The primary peritoneal macrophages treated with oxLDL increased the inflammatory factor Il-6,Il-1β,Tnf-αmRNA levels and the levels of Ccl2 compared with the control group（P<0.05）.In basal level,the lipoprotein receptor Cd36,as the downstream gene of Nrf2,decreased with the deletion of Nrf2 mRNA level（P<0.05）,but the mRNA levels of Abca1,Abcg1,Cd36 and Sr-a increased after oxLDL treatment compared with the control group（P<0.05）.4.After chemotaxis treatment of primary peritoneal macrophages in culture medium containing CCL2,Nrf2 deletion promoted the migration of macrophages through Transwell（P<0.05）.Conclusion:1.In the context of ApoE gene deletion,the degree of atherosclerotic lesions in mice with Nrf2-specific deletion of myeloid monocytes was increased.2.Nrf2 deficiency in myeloid monocytes increases the expression levels of inflammatory cytokines and chemokines,also promotes lipid uptake by macrophages.3.Nrf2 deficiency in macrophages increases the ability of macrophages to be chemotaxed by chemokines.