| Background and Objective:Atherosclerosis is a progressive arterial disease,which is mainly caused by the long-term accumulation of vascular occlusive plaque in the subendothelial intima layer of large and medium-sized arteries,and eventually led to myocardial infarction(MI)and stroke.More and more evidences confirm that atherosclerosis is an autoimmune disease caused by multiple factors.In our previous experiments,we had screened a new autoantigen associated with atherosclerosis,collagen Ⅵ,and successfully expressed its humanized full-length antibody,collagen Ⅵ antibody.When immunized fed apoE-/-mice fed with high-fat-diet with collagen Ⅵ antibody,we initially found that collagen Ⅵ antibody could effectively regress the plaque area of aorta more than 40%,and promote the polarization of macrophages from M1 toward M2,as well as increase ABCA1 expression and cholesterol efflux.However,the mechanism of collagen Ⅵ antibody in regression atherosclerotic plaque area is still unclear.This project intends to further clarify the therapeutic effect of collagen Ⅵ antibody on atherosclerosis on the basis of previous research,and explore the role of hepatocyte cholesterol reverse transport pathway in the elimination of plaque area by collagen Ⅵ antibody and its regulatory mechanism.Methods:1.Oil red O staining was used to analyze the plaque area of the aorta,and serum was separated to determine serum and liver total cholesterol,triglycerides,HDL-C,LDL-C content.2.The ultra-high resolution small animal ultrasound imaging system vevo2100 was used to measure the abdominal aorta inner diameter and blood flow velocity before and after collagen Ⅵ antibody treatment.3.Western blot and RT-qPCR were applied to detect the expression of SR-BI,ApoA-Ⅰ,ApoA-Ⅱ,CYP7A1 and CYP27A1 both in the liver and HepG2 cell,liver tissue was evaluated with H&E staining,and the content of total bile acid in mouse feces was determined.4.siRNA knockeddown the expression of FcRn,Western blot was used to detect the ERK1/2-MAPK signaling pathway and the expression of SR-BI,ApoA-Ⅰ,ApoA-Ⅱ,CYP7A1 and CYP27A1.5.Construct SR-BI luciferase reporter gene,and SR-BI luciferase activity was detected by truncation and site mutation.6.Flow cytometry and fluorescence detection of Dil-HDL uptake by HepG2 cells.Results:1.Collagen Ⅵ antibody reduces the aortic plaque area of apoE-/-mice fed by high fat diet,but has no significant effect on serum total cholesterol,triglycerides,HDL-C,and LDL-C.2.Collagen Ⅵ antibody delays the progression of atherosclerosis and improves liver damage.3.Collagen Ⅵ antibody up-regulates the expression of SR-BI,ApoA-Ⅰ,ApoA-Ⅱ,CYP7A1 and CYP27A1 in mouse liver,and promotes the excretion of bile acids in mouse feces.4.Collagen Ⅵ antibody up-regulates the expression of SR-BI,ApoA-Ⅰ,and ApoA-Ⅱ proteins in HepG2 cells,and increases the transcription of CYP7A1 and CYP27A1 mRNA.5.Collagen Ⅵ antibody activates the ERK1/2-MAPK signaling pathway and up-regulates the expression of SR-BI,ApoA-Ⅰ,and ApoA-Ⅱ proteins.6.The expression of SR-BI depends on PPARa and is regulated by ERK1/2-MAPK.7.Collagen Ⅵ antibody up-regulates the expression of SR-BI,ApoA-Ⅰ and ApoA-Ⅱ proteins in HepG2 cells through FcRn receptor.8.Collagen Ⅵ antibody promotes the uptake of Dil-HDL by HepG2 cells.Conclusions:1.Collagen Ⅵ antibody reduces the area of atherosclerotic plaque by 40%,delays the progression of abdominal aortic artery stenosis effectively.2.Collagen Ⅵ antibody improves liver steatosis and promotes the excretion of bile acids.3.Collagen Ⅵ regulates the expression of SR-BI in hepatocytes through FcRnERKl/2-PPARα/RXR signal pathway. |
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