Zhuyu Pills(茱萸丸)Regulate TMA/FMO3/TMAO Pathway and Promote Cholesterol Reverse Transport and Anti-Atherosclerosis
Objective This article aims to investigate the effect of Zhuyu Pills(茱萸丸)on atherosclerosis and the underlying mechanism.The mouse model of atherosclerosis was induced by a high-fat diet,and the total modeling period was 12 weeks.Methods A total of 47 ApoE-mice successfully modeled were randomized into 5 groups,including 10 in the model group,9 in each of low,medium and high dose(130.54,261.08 and 522.16 mg·kg-1,respectively)Zhuyu Pills groups,and 10 in the ator-vastatin calcium(10.40 mg·kg-1)group.In addition,10 C57BL/6J mice were included as the normal group.The mice in the normal group and model group were administrated with an equal volume of sterile distilled water,and those in the other groups were administrated with the corresponding agents by gavage once a day for 12 weeks.The pathological changes of aorta and liver were observed by hematoxylin-eosin(HE)staining after administration.The levels of oxidized low-density lipoprotein(ox-LDL),monocyte chemotactic egg-1(MCP-1),vascular cell adhesion-1(VCAM-1)and intercellular adhesion molecule-1(ICAM-1)in serum were detected by enzyme-linked immunosorbent assay(ELISA).The levels of total cholesterol(TC),tri-glyceride(TG)and low density lipoprotein cholesterol(LDL-C)in liver were detected by biochemical method.Ultra high per-formance liquid chromatography-mass spectrometry(UHPLC-MS/MS)was used to detect the plasma trimethylamine(TMA)and trimethylamine oxide(TM AO).The expression level of flavin containing monooxygenase 3(FMO3)protein in mouse liver was detected by immunofluorescence.Real-time fluorescence quantitative polymerase chain reaction(real-time PCR)was em-ployed to measure the mRNA levels of FMO3,adenosine triphosphate binding cassette transporter A1(ABCA1),scavenger recep-tor B1(SR-B1),ATP binding cassette transporter G5(ABCG5),adenosine triphosphate-binding cassette transporter G8(AB-CG8)and cytochrome P450 7A1(CYP7A1)in the liver tissues.Results Compared with the control group,the model group showed large area thickening of the aortic intima and severe hepatic steatosis and inflammatory infiltration.The levels of ox-LDL,MCP-1,VCAM-1 and ICAM-1 in serum were increased.The levels of TC,TG and LDL-C in liver increased.The levels of plasma TMA and TMAO increased.The mRNA and protein expression levels of FMO3 in liver were increased,and the mRNA expression levels of ABCA1,SR-B1,ABCG5,ABCG8 and CYP7A1 were increased(P＜0.01).Compared with that of the model group,HE staining of low,medium and high dose Zhuyu Pills groups and atorvastatin calcium group showed that with the concentration increase of Zhuyu Pills,the plaque enrichment area was gradually narrowed,liver steatosis and inflammatory in-filtration decreased.The levels of ox-LDL,MCP-1,VCAM-1 and ICAM-1 in serum were decreased.The levels of TC,TG and LDL-C in liver decreased.The levels of TMA and TMAO in plasma decreased.The mRNA and protein expression levels of FMO3 in liver were decreased,and the mRNA expression levels of ABCA1,SR-B1,ABCG5,ABCG8 and CYP7A1 were de-creased(P＜0.01 or P＜0.05).Conclusion Zhuyu Pills have a good therapeutic effect on atherosclerotic plaque in mice,and its mechanism may be related to regulating the lipid metabolism pathway of TMA/FMO3/TMAO and promoting the reverse transport and decomposition of cholesterol.
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