Inhibition of cholesterol biosynthesis by Delta22-unsaturated phytosterols via competitive inhibition of sterol Delta24-reductase in mammalian cells.
Dietary phytosterols are cholesterol-lowering agents that interfere with the intestinal absorption of cholesterol. In the present study, we have studied their effects on cholesterol biosynthesis in human cells, particularly in the sterol-conversion pathway. For this, both Caco-2 (intestinal mucosa) and HL-60 (promyelocytic) human cell lines were incubated with [(14)C]acetate, and the incorporation of radioactivity into sterols was determined using HPLC and radioactivity detection online. Sterols containing a double bond at C-22 in the side chain (stigmasterol, brassicasterol and ergosterol) dramatically inhibited the activity of sterol Delta(24)-reductase, as indicated by the decrease in radioactivity incorporation into cholesterol and the accumulation of its precursors (mainly desmosterol). Phytosterols with the saturated side chain (beta-sitosterol and campesterol) were inactive in this regard. The inhibition of sterol (24)-reductase was confirmed in rat liver microsomes by using (14)C-labelled desmosterol as the substrate. The (22)-unsaturated phytosterols acted as competitive inhibitors of sterol (24)-reductase, with K(i) values (41.1, 42.7 and 36.8 microM for stigmasterol, brassicasterol and ergosterol respectively) similar to the estimated K(m) for desmosterol (26.3 microM). The sterol 5,22-cholestedien-3beta-ol, an unusual desmosterol isomer that lacks the alkyl groups characteristic of phytosterols, acted as a much stronger inhibitor of (24)-reductase (K(i)=3.34 microM). The usually low intracellular concentrations of the physiological substrates of (24)-reductase explains the strong inhibition of cholesterol biosynthesis that these compounds exert in cells. Given that inhibition of sterol (24)-reductase was achieved at physiologically relevant concentrations, it may represent an additional mechanism for the cholesterol-lowering action of phytosterols, and opens up the possibility of using certain (22)-unsaturated sterols as effective hypocholesterolaemic agents.
Fernández
Carlos
C
Servicio de Bioqui;mica-Investigación, Hospital Ramón y Cajal, 28034 Madrid, Spain.
Suárez
Yajaira
Y
Ferruelo
Antonio J
AJ
Gómez-Coronado
Diego
D
Lasunción
Miguel A
MA
D000818:Animals; D001667:Binding, Competitive; D018938:Caco-2 Cells; D002784:Cholesterol / Q000096:biosynthesis* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D004305:Dose-Response Relationship, Drug; D018922:HL-60 Cells; D006801:Humans; D000960:Hypolipidemic Agents / Q000494:pharmacology; D007700:Kinetics; D008264:Macrophages / Q000378:metabolism; D008297:Male; D051379:Mice; D008862:Microsomes, Liver / Q000378:metabolism; D008956:Models, Chemical; D010088:Oxidoreductases / Q000737:chemistry / Q000378:metabolism*; D010840:Phytosterols / Q000737:chemistry*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013265:Stigmasterol / Q000494:pharmacology; D013997:Time Factors; D014407:Tumor Cells, Cultured
D000960:Hypolipidemic Agents; D010840:Phytosterols; D002784:Cholesterol; D013265:Stigmasterol; D010088:Oxidoreductases; C022673:delta 24-sterol reductase