Effect And Preliminary Mechanism Of Midkine On Atherosclerotic Lesions And Abca1 Expression In ApoE^-/- Mice

Background:Atherosclerosis?As?is one of the main pathological bases of coronary heart disease,cerebral infarction,and peripheral vascular disease and other diseases.Currently,it is an important research filed to study the pathogenesis of As and explore effective treatment strategies.Studies have shown that midkine?MK?is a new growth factor with multiple functions in the human body,and its reduction can slow the course of As.In healthy conditions,the expression level of MK in the vascular tissues of arteries and veins is low,but in the animal models of rabbit and rat endothelial injury,the expression level of MK is significantly up-regulated.In addition,it has been found that macrophages infiltrated from the damaged blood vessel site are the main source of MK production.Previous studies have shown that the role of MK in promoting As is mainly manifested in pro-inflammatory cell infiltration,promoting the propagation of endothelial cells and smooth muscle cells,and intimal hyperplasia,etc.However,these mechanisms cannot fully explain all pathological phenomena of As.Therefore,The role and molecular mechanism of MK in the occurrence and development of As remains to be further studied.Purpose:In our study,ApoE^-/-mice were fed with high-fat diets to replicate the animal model of As,to observe the effect of MK on atherosclerotic lesions.At the same time,the expression of ABCA1,the key protein of cholesterol efflux,was studied to explore the role of MK in the occurrence and development of As and its mechanism.Methods:Thirty-two 8-week-old male ApoE^-/-mice were randomly divided into 5 groups:the control group?n=4?,the vehicle group?n=4?,the MK group?n=8?,the SP600125?a JNK inhibitor?group?n=8?,and the MK+SP600125 group?n=8?.After 1 week of adaptive feeding,mice were fed on a high-fat diet for 4 weeks.Then the MK group was injected intraperitoneally daily with recombinant human midkine?rhMK??20ug/kg/d?,the SP600125 group was injected intraperitoneally daily with SP600125?16mg/kg/d?,the MK+SP600125 group was injected intraperitoneally daily with rhMK and SP60012,while the control group was injected intraperitoneally with the same volume of corn oil and The control group did nothing.Mice were fed on a high-fat diet while administrating the drug.The mice were sacrificed after 2 weeks of treatment.Blood was taken from the eyeball and serum was collected after centrifugation.Serum TC,TG,LDL and HDL were tested by automatic biochemical analyzer.The frozen heart sections of the aortic sinus tissue were made in the mouse heart,and the frozen liver sections were made in the mouse liver.Lipid accumulation and atherosclerotic lesions were observed by HE,Masson and oil red O staining.The expressions of p-JNK,AP1 and ABCA1 were detected by immunohistochemistry and immunofluorescence staining.Results:1.The serum lipid levels of ApoE^-/-mice showed that compared with the control group,TG,TC,and LDL levels of the MK group were increased?P<0.05?;However,compared with the MK group,TG,TC,and LDL levels in the MK+SP600125 group were reduced?P<0.05?.2.Compared with the control group,the aortic sinus area of the MK group has increased by 8.08%?P<0.05?.The lipid accumulation was significantly increased by oil red O staining,and the degree of fibrosis was also aggravated by Masson staining.Compared with the MK group,the aortic sinus plaque area of the MK+SP600125 group has decreased by 3.88%?P<0.05?,lipid accumulation and fibrosis degree were reduced.3.HE staining and oil red O staining of frozen sections of mouse liver tissue suggested that the structure of the hepatic lobules of the control group was normal with clear morphology,and the morphology and the structure of the hepatocytes were normal.The liver structure of the MK group was damaged,the structure of hepatic lobule was disordered,and the liver cells were oedematous.Compared with MK group,the degree of hepatocellular lesion in MK+SP600125 group was significantly reduced,and the damage degree of hepatic lobule structure was less.4.Immunohistochemistry and immunofluorescence staining of the aortic sinus and liver tissues of the mice showed that compared with the control group,the expression of ABCA1 was down-regulated and the expression of AP1 and p-JNK were up-regulated.Compared with the MK group,the expression of ABCA1 of the MK+SP600125 group was up-regulated,while the expression of AP1and p-JNK were down-regulated.Conclusion:MK can promote the increase of blood lipid level of ApoE^-/-mice,promote the progression of atherosclerotic lesions,down-regulate the expression of ABCA1 in the atherosclerotic lesions,which may be achieved by regulating the JNK/AP1 pathway.