乙酰泽泻醇降低胆固醇分子机制研究

Study on Molecular Mechanism of Alisol Acetates in Lowering Cholesterol

目的从分子水平探讨乙酰泽泻醇降低胆固醇(TC)的作用机理。方法利用试剂盒法测定了调脂中药泽泻主要有效成分23-乙酰泽泻醇B和24-乙酰泽泻醇A对高脂小鼠TC的影响,利用试剂盒法、Western blotting技术结合分子模拟技术研究两者对TC代谢关键酶3-羟基-3-甲基戊二酸单酰辅酶A(HMG-Co A)还原酶作用的分子机理。结果乙酰泽泻醇能显著降低高脂小鼠TC(P<0.01,P<0.05),23-乙酰泽泻醇B作用强度高于24-乙酰泽泻醇A(P<0.05),同时在体内、体外均可剂量依赖性下调HMG-Co A还原酶活性(P<0.01),且23-乙酰泽泻醇B对该酶作用强于24-乙酰泽泻醇A(P<0.05)。两者均未显著下调HMG-Co A还原酶的蛋白表达(P>0.05)。两者与HMG-Co A还原酶结合的关键氨基酸残基可能为Lys691、Asp767、Asn658。结论乙酰泽泻醇降低TC的机制可能是其原型药物通过抑制HMG-Co A还原酶活性来达到,且可能是通过直接竞争性与HMG-Co A还原酶结合抑制其作用,乙酰泽泻醇的舵手基团为其侧链,侧链与母环折叠结合弱,打开结合强。

Objective To explore the mechanism of alisol acetates in regulating total cholesterol（TC）at the molecular level.Methods We observed the impact of alisol B 23-acetate and alisol A 24-acetate,the main active ingredients of the lipid-regulating Chinese herbal medicine Rhizoma Alismatis,on TC of hyperlipidemic mice by using the reagent kits.The interaction of alisol B 23-acetate and alisol A 24-acetate to 3-hydroxy-3-methylglutary-coenzyme A（HMGCo A） reductase,a key enzyme involving in the metabolism of TC,was studied using the reagent kit method and the Western blotting technique combined with molecular simulation technique.Results The results showed that alisol acetates significantly lowered TC concentrations of hyperlipidemic mice（P〈0.01,P〈0.05）.The effect of alisol B23-acetate on HMG-Co A reductase was stronger than that of alisol A 24-acetate（P〈0.05）.Meanwhile,alisol acetates lowered HMG-Co A reductase activity in a dose-dependent manner,and the enzyme inhibition of alisol B 23-acetate was superior to that of alisol A 24-acetate,both in vivo and in vitro（P〈0.01）.Neither of the two alisol acetates significantly lowered the protein expression of HMG-Co A（P〈0.05）.The key amino acid residues binding to HMG-Co A reductase may be Lys691,Asp767,and Asn658.Conclusion The mechanism of alisol acetates lowering the TC level is probably through inhibiting the activity of HMG-Co A reductase by its prototype drug,which may exhibit an inhibition effect via directly and competitively binding to HMG-Co A.The side chain of the alisol acetate is the steering group.A folded side chain or parent ring bound to HMG-Co A reductase less weakly than an open side chain or parent ring.