| Objective:To study the effects of specifically blocking the binding of transforming growth factor-β1(TGF-β1)downstream molecule β1-catenin/TCF in ApoE-/-mice on macrophage transformation and plaque stability in mouse carotid plaques influences.Methods:Take 40 6-8 week old ApoE-/-C57BL/6J male mice(20~25g)and feed them with a high-fat diet for 6 weeks.Carotid artery plaque model was created by tandem ligation of the right carotid artery and continued high-fat diet.They were fed for 5 weeks and were randomly divided into four groups(A,B,C,D,n=10),and were treated with corresponding exogenous drugs.ApoE-/-C57BL/6J mice in group A were treated with saline(5mg/kg)+ TGF-β1(50ug/kg/d);ApoE-/-C57BL/6J mice in group B were treated with ICG-001(5 mg/kg)kg)+TGF-β1(50ug/kg)treatment;ApoE-/-C57BL/6J mice in group C were treated with saline(50ug/kg)+ICG-001(5mg/kg);group D ApoE-/-C57BL/6J mice were treated with normal saline(5.05mg/kg);the corresponding reagents were injected intraperitoneally every day for 14 consecutive days.Evaluate the general condition of the four groups of mice;carefully separate the mouse carotid tissues and perform hematoxylin-eosin(HE)staining to assess the stability and vulnerability of carotid plaques;use immunohistochemistry(IHC)techniques CD86(M1 macrophages)and CD206(M2 macrophages)in each group of specimens were detected to assess the phenotype,location and level of macrophages in each tissue;the enzyme-linked immunosorbent assay(ELISA)was used to determine the size of each group.The contents of interleukin 6(IL-6),low-density lipoprotein cholesterol(LDL-c),interleukin10(IL-10)and total cholesterol(TC)in rat serum.Image plus6.0,SPSS25.0 statistical software was used for analysis.Results:1.General information:1.1 Body weight: Before the experiment,the weight measurement data of the 4groups of mice had no statistical difference between each group(P>0.05);after the experiment,the weight measurement data of the 4 groups of mice had no statistical difference between each group(P>0.05).1.2 There was no significant difference in the variance analysis of serum total cholesterol(TC)levels in ApoE-/-C57BL/6J mice in each group(F=1.496,P>0.05);serum low density in ApoE-/-C57BL/6J mice in each group The results of variance analysis of lipoprotein cholesterol lipid(LDL-c)levels were not statistically significant(F=1.382,P>0.05).2.HE staining: Compared with the normal saline group ApoE-/-C57BL/6J mice,the carotid artery section HE of the ICG-001+TGF-β1 and normal saline+TGF-β1 group ApoE-/-C57BL/6J mice The staining results show that the lumen is narrow,the fiber cap is thick,the lipid core is small,and the number of lipid cores is small.3.Immunohistochemistry: Compared with the normal saline group,the anti-inflammatory M2 macrophage cell surface expression molecule mannose receptor(CD206)expressed more in the ICG-001+TGF-β1 and normal saline+TGF-β1 groups,The staining becomes darker,while the positive expression of the cell surface expression molecule CD86 secreted by M1 type macrophages is less,and the staining becomes lighter.4.ELISA: The variance analysis of serum interleukin 10(IL-10)levels in each group was not statistically significant(F=1.531,P>0.05);serum interleukin 6(IL-6)in each group The difference in horizontal analysis of variance was statistically significant(F=51.324,P<0.05),and the difference between groups was analyzed by LSD analysis method,and it was found that the normal saline + TGF-β1 group,the ICG-001 + TGF-β1group,and the normal saline + ICG There were significant differences in IL-6 levels between the-001 group and the normal saline group(P<0.05),with the ICG-001+TGF-β1 group having the lowest level,followed by normal saline + TGF-β1group,normal saline +ICG-001 group,normal saline group.Conclusion:1 Exogenous administration of TGF-β1 can affect the phenotype of macrophages in mouse carotid atherosclerotic plaques;2 After ICG-001 specifically blocks β-catenin/TCF downstream of TGF-β,it has a positive effect on reducing the content of pro-inflammatory M1 macrophages and increasing plaque stability;3 Exogenous addition of TGF-β1+ICG-001 can enhance the plaque stabilization effect of TGF-β1,which may be achieved by enhancing the anti-inflammatory effect ofβ1-catenin/FOXO1 downstream of TGF-β1. |
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