摘要
目前,已发现的靶向抑制肠道脂质摄取的降脂药物作用大多局限于胆固醇,达不到预期的降脂效果,且具有一定的副作用.因此,寻找能同时有效抑制肠道脂肪酸和胆固醇摄取的新药物及新靶点是可行的新策略.为了寻找和筛选对肠道脂质摄取具有抑制效应的天然化合物,本文构建了基于脂质摄取和转化的荧光高通量体外双评价体系.经过对104个天然化合物的筛选发现,盐酸小檗碱(BBR)的效应最为明显,且对肠道胆固醇和脂肪酸摄取的双重抑制作用均呈现时间-剂量依赖性.此外,随着剂量和处理时间的增加,BBR可以显著抑制酰基辅酶A:胆固醇酰基转移酶2(SOAT2)的表达.进一步通过SOAT2干扰实验证实,SOAT2特异性介导了BBR对脂质摄取的抑制作用,并发现SOAT2可能是同时调控胆固醇与脂肪酸摄取的关键靶点.因此,本研究通过利用高通量筛选体系,为降脂药物筛选提供了新策略,并提出肠道SOAT2可能是一种潜在的降脂新靶点,可用于筛选和寻找更安全有效的治疗药物,同时也为BBR的临床应用提供了新的证据支持.
Most of the lipid-lowering drugs used in clinical practice target the liver and achieve lipid-lowering effects by promoting lipid clearance and metabolism.Reducing intestinal uptake of lipids may be an important new strategy for the development of lipid-lowering drugs.It is necessary to find new lipid-lowering drug targets that can effectively inhibit intestinal lipid uptake.Acyl-CoA:cholesterol acyltransferase 2(SOAT2,also known as ACAT2)is a key enzyme in the synthesis of into cholesteryl esters from free cholesterol and fatty acids;it is expressed only in the liver and intestine.SOAT2 is involved in intestinal cholesterol uptake,which suggests that it is a promising target for intestinal lipid-lowering drugs.We developed a fluorescent high-throughput in vitro evaluation system based on lipid uptake and transformation to identify natural compounds with inhibitory effects on intestinal free fatty acid and cholesterol uptake.Subset 1 of natural compounds that inhibit cholesterol uptake was obtained by detecting the uptake and esterification of fluorescently labelled cholesterol(NBD-cholesterol)in Caco2 cells after exposure to natural compounds through high-throughput screening.Subset 2 of natural compounds inhibiting the uptake of free fatty acids was obtained by indicating the uptake of fluorescently labelled fatty acids(BODIPYTM FL C16).Based on the screening results of the two subsets,candidate natural compounds with inhibitory effects on cholesterol and fatty acid uptake were obtained.After screening 104 natural compounds,berberine hydrochloride(BBR)was the most potent natural chemical that had inhibitory effects on both intestinal cholesterol and fatty acid uptake.The inhibitory efficiencies of fatty acid and cholesterol uptake were 16.3%and 28.53%,respectively.We found that SOAT2 was a key protein in mediating the BBR-induced inhibition of intestinal cholesterol uptake.When SOAT2 expression resumed,the inhibitory effect of BBR on cellular cholesterol uptake was mitigated.Our results provide evidence that the reduction of hepatic cholesterol caused by BBR may be resulted from decrease in cholesterol input through the inhibition of intestinal cholesterol absorption,which was mediated by SOAT2.In addition,we demonstrated that the inhibitory effect of BBR on intestinal fatty acid uptake was regulated by SOAT2.This study is the first to identify the role of SOAT2 in the regulation of intestinal fatty acid uptake and provides the novel evidences for the application of intestinal SOAT2 inhibition in lipid-lowering.The expression of SOAT2 and the uptake of lipids(cholesterol and fatty acids)decreased in a dose and time dependent manner after BBR treatment.The inhibitory effect of BBR on intestinal cholesterol and even fatty acid uptake was restored in cells overexpressing SOAT2.Therefore,SOAT2 would be an important target for mediating intestinal uptake,and the inhibition of SOAT2 to reduce intestinal lipid uptake is a new approach for future lipid-lowering drug development.This study provides a new high-throughput method for screening intestinal lipid-lowering drugs.Intestinal SOAT2 inhibition may be a potential lipid-lowering strategy.
作者
梁静佳
邵文涛
顾爱华
Jingjia Liang;Wentao Shao;Aihua Gu(State Key Laboratory of Reproductive Medicine,Nanjing Medical University,Nanjing 211166,China)
出处
《科学通报》
EI
CAS
CSCD
北大核心
2023年第16期2124-2132,共9页
Chinese Science Bulletin
基金
国家自然科学基金-广西壮族自治区联合基金(U21A20340)资助。
