基于N-糖基化修饰组学探究人参皂苷Rb1改善高脂血症大鼠脂代谢紊乱机制

Mechanism of Ginsenoside Rb1 in Improving Lipid Metabolism Disorder in Hyperlipidemia Rats Based on N-glycoproteomic Analysis

目的通过研究人参皂苷Rb1对高脂血症大鼠血浆N-糖基化修饰组学的影响,在蛋白质修饰水平探讨人参皂苷Rb1对脂代谢紊乱的作用机制。方法SPF级SD大鼠24只,随机分为正常对照组、高脂组与Rb1组,每组8只。高脂组与Rb1组饲喂高脂饲料4周。Rb1组从第5~12周给予人参皂苷Rb1200 mg/(kg·d)灌胃,正常对照组与高脂组同期给予等体积生理盐水灌胃,给药期间高脂组与Rb1组继续给予高脂喂养。12周后,全自动生化分析仪检测血清血脂[总胆固醇(TC)、总甘油三酯(TG)、低密度脂蛋白胆固醇(LDL-C)、高密度脂蛋白胆固醇(HDL-C)]水平,HE与油红O染色观测肝组织形态学变化与脂质沉积情况。N-糖基化修饰组学探究人参皂苷Rb1改善高脂血症大鼠脂代谢紊乱的作用机制。结果与正常对照组比较,高脂组血清TC、TG、LDL-C水平升高,HDL-C水平降低(P<0.01);与高脂组比较,Rb1组血清TC、TG降低(P<0.01,P<0.05),HDL-C水平升高(P<0.05)。形态学结果显示人参皂苷Rb1能够显著改善高脂血症大鼠肝脏组织结构与脂质沉积。N-糖基化修饰组学结果表明,53个糖基化修饰蛋白的98个位点在两个比较组中(高脂组与正常对照组比较及Rb1组与高脂组比较)发生共同改变,白蛋白(Alb)与Serpinc1是修饰位点改变最多的蛋白;基因本体(GO)功能分析发现差异修饰糖蛋白主要参与到炎症反应(Serpinc1)、细胞铁离子稳态[铜蓝蛋白(Cp)]和胆固醇流出的正向调节[对氧磷酶1(PON1)和低密度脂蛋白受体相关蛋白1(lrp1)]等生物学过程中;京都基因与基因组百科全书(KEGG)分析发现补体和凝血级联反应是富集差异修饰蛋白最多的通路。结论人参皂苷Rb1可能通过调节胆固醇流出、细胞铁离子稳态、炎症反应以及补体和凝血级联反应等过程中相关蛋白的N-糖基化修饰水平,调节血脂紊乱,改善肝脏脂质沉积。

Objective To investigate the effect of ginsenoside Rb1 on N-glycosylation of plasma proteins in hyperlipidemia rats and to explore the mechanism of ginsenoside Rb1 on lipid metabolism disorder at protein modification level.Methods Totally 24 SPF SD rats were randomly divided into normal control group,hyperlipidemia group and Rb1 group,with 8 rats in each group.The hyperlipidemia group and Rb1 group were fed high fat diet for 4 weeks.Rb1 group was given ginsenoside Rb1200 mg·kg-1·d-1 in gavage from week 5 to week 12,and normal control group and hyperlipidemia group were given equal volume of normal saline in gavage at the same time.The hyperlipidemia group and Rb1 group continued high fat diet during administration.After 12 weeks,serum lipid levels[total cholesterol(TG),total triglycerides(TC),low density lipoprotein cholesterol(LDL-C),high density lipoprotein cholesterol(HDL-C)]were detected by automatic biochemical analyzer,liver histomorphology and lipid deposition were observed by HE and oil red O staining.The mechanism of ginsenoside Rb1 in improving lipid metabolism disorder in hyperlipidemia rats was investigated by N-glycoproteomic analysis.Results Compared with normal control group,serum TC,TG and LDL-C level in hyperlipidemia group increased,while HDL-C level decreased(P<0.01).Compared with hyperlipidemia group,serum TC and TG in Rb1 group decreased(P<0.01,P<0.05),and HDL-C level increased(P<0.05).Morphological results showed that ginsenoside Rb1 could significantly improve liver tissue structure and lipid deposition in hyperlipidemia rats.Analyses of N-glycosylation sites distribution found that 98 differential N-glycosylation sites on 53 glycoproteins between 2 comparison groups(hyperlipidemia group vs.normal control group,Rb1 group vs.hyperlipidemia group)were identified,and albumin(Alb)and Serpinc1 were most heavily modified.Gene Ontology enrichment analysis showed that differential modified glycoproteins were involved in inflammatory response(Serpinc1),cellular iron ion homeostasis(ceruloplasmin)and positive regulation of cholesterol efflux(paraoxonase 1 and LDL receptor-related protein 1)etc.biological process.Complement and coagulation cascades was the most significant enriched in the KEGG pathway enrichment analysis.Conclusion Ginsenoside Rb1 may regulate blood lipid disorders and improve liver lipid deposition by regulating cholesterol effluence,cell iron homeostasis,inflammatory response,complement and coagulation cascade and other related protein N-glycosylation modification levels.