European Journal of Nutrition

, Volume 55, Issue 3, pp 1041–1057 | Cite as

Modulation of cholesterol-related gene expression by ergosterol and ergosterol-enriched extracts obtained from Agaricus bisporus

  • Alicia Gil-Ramírez
  • Víctor Caz
  • Roberto Martin-Hernandez
  • Francisco R. Marín
  • Carlota Largo
  • Arantxa Rodríguez-Casado
  • María Tabernero
  • Alejandro Ruiz-Rodríguez
  • Guillermo Reglero
  • Cristina Soler-Rivas
Original Contribution



To investigate the effect of two extracts obtained from Agaricus bisporus on the mRNA expression of cholesterol-related genes. One of the extracts contained ergosterol and other fungal sterols (SFE) and the other contained β-glucans and fungal sterols (EβG).


Firstly, the dietary mixed micelles (DMMs) generated after in vitro digestion of standards and SFE were applied to Caco2 cells. Then, the lower compartment after a Caco2-transport experiment was applied to HepG2 cells. The mRNA expression was assessed in both cell lines by low-density arrays (LDA). Mice received the extracts, ergosterol or control drugs after 4 weeks of a high-cholesterol diet. The lipid profile of plasma, liver and feces was determined. LDA assays were performed in liver and intestines.


The DMM fraction of SFE up-regulated the LDLR mRNA expression in Caco2 cells. The lower compartment after Caco2-transport experiments up-regulated LDLR and modulated several other lipid-related genes in HepG2 cells. In mice, SFE decreased TC/HDL ratio and reduced hepatic triglycerides paralleled with down-regulation of Dgat1 expression, while EβG did it without transcriptional changes. Addition of SFE or ergosterol induced in jejunum a similar transcriptional response to simvastatin and ezetimibe; they all down-regulated Srebf2 and Nr1h4 (FXR) genes.


Ergosterol-containing extracts from A. bisporus lowered hepatic triglyceride and modify the mRNA expression of cholesterol-related genes although the transcriptional regulation was unrelated to changes in plasma lipid profile. These extracts may be useful limiting hepatic steatosis and as bioactive ingredients to design novel functional foods preventing lifestyle-related diseases such as non-alcoholic fatty liver disease.


Ergosterol Cholesterol Supercritical CO2 extraction White button mushroom Low-density array (LDA) Gene expression 



Lard enriched with cholesterol and ergosterol


Lard enriched with cholesterol


Lard enriched with cholesterol and the SFE extract


Dietary mixed micelles

EβG extract

Extract containing ergosterol and β-glucans


Farnesoid X receptor


High-density lipoprotein


Low-density array


Low-density lipoprotein


Liver X receptor

SFE extract

Extract obtained by supercritical fluid extraction


Sterol regulatory element-binding protein



The research was supported by AGL2010-21537 national R+D program from the Spanish Ministry of Science and Innovation and ALIBIRD-C M S2009/AGR-1469 regional program from the Community of Madrid (Spain). CTICH (Centro Tecnológico de Investigación del Champiñón de La Rioja, Autol, Spain) is also acknowledged for the cultivation and supplying of the mushrooms fruiting bodies and Maria Navarro-Rubio for her technical assistance.

Conflict of interest



  1. 1.
    Gil-Ramirez A, Soler-Rivas C (2014) The use of edible mushroom extracts as bioactive ingredients to design novel functional foods with hypocholesterolemic activities. In: Pesti G (ed) Cultivation, antioxidant properties and health benefits. Nova Publishers, New York, pp 43–73Google Scholar
  2. 2.
    Jeong SC, Jeong YT, Yang BK, Islam R, Koyyalamudi SR, Pang G, Cho KY, Song CH (2010) White button mushroom (Agaricus bisporus) lowers blood glucose and cholesterol levels in diabetic and hypercholesterolemic rats. Nutr Res 30(1):49–56CrossRefGoogle Scholar
  3. 3.
    Calpe-Berdiel L, Escolà-Gil JC, Blanco-Vaca F (2009) New insights into the molecular actions of plant sterols and stanols in cholesterol metabolism. Atherosclerosis 203:18–31CrossRefGoogle Scholar
  4. 4.
    Lammert F, Wang DQH (2005) New insights into the genetic regulation of intestinal cholesterol absorption. Gastroenterol J 129:718–734CrossRefGoogle Scholar
  5. 5.
    Gil-Ramírez A, Ruiz-Rodríguez A, Marín FR, Reglero G, Soler-Rivas C (2014) Effect of ergosterol-enriched extracts obtained from Agaricus bisporus on cholesterol absorption using an in vitro digestion model. J Funct Foods 11:589–597CrossRefGoogle Scholar
  6. 6.
    Kaneko E, Matsuda M, Yamada Y, Tachibana Y, Shimomura I, Makishima M (2003) Induction of intestinal ATP-binding cassette transporters by a phytosterol-derived liver X receptor agonist. J Biol Chem 278(19):36098Google Scholar
  7. 7.
    Matsubara T, Li F, Gonzalez FJ (2013) FXR signaling in the enterohepatic system. Moll Cell Endocrinol 368(1–2):17–29CrossRefGoogle Scholar
  8. 8.
    Miyata M, Hata T, Yamazoe Y, Yoshinari K (2014) SREBP-2 negatively regulates FXR-dependent transcription of FGF19 in human intestinal cells. Biochem Biophys Res Commun 443(2):477–482CrossRefGoogle Scholar
  9. 9.
    Jesch ED, Seo JM, Carr TP, Lee JY (2009) Sitosterol reduces messenger RNA and protein expression levels of Niemann-Pick C1-like 1 in FHs 74 Int cells. Nutr Res 29:859–866CrossRefGoogle Scholar
  10. 10.
    Gil-Ramírez A, Aldars-García L, Palanisamy M, Jiverdeanu RM, Ruiz-Rodríguez A, Marín FR, Reglero G, Soler-Rivas C (2013) Sterol enriched fractions obtained from Agaricus bisporus fruiting bodies and by-products by compressed fluid technologies (PLE and SFE). Innov Food Sci Emerg Technol 18:101–107CrossRefGoogle Scholar
  11. 11.
    Van de Steeg E, Kleemann R, Jansen HT, van Duyvenvoorde W, Offerman EH, Wortelboer HM, Degroot J (2013) Combined analysis of pharmacokinetic and efficacy data of preclinical studies with statins markedly improves translation of drug efficacy to human trials. J Pharmacol Exp Ther 347:635–644CrossRefGoogle Scholar
  12. 12.
    Lei YF, Chen JL, Wei H, Xiong CM, Zhang YH, Ruan JL (2011) Hypolipidemic and anti-inflammatory properties of Abacopterin A from Abacopteris penangiana in high-fat diet-induced hyperlipidemia mice. Food Chem Toxicol 49:3206–3210CrossRefGoogle Scholar
  13. 13.
    Qin YW, Ye P, He JQ, Sheng L, Wang LY, Du J (2010) Simvastatin inhibited cardiac hypertrophy and fibrosis in apolipoprotein E-deficient mice fed a “Western-style diet” by increasing PPAR α and γ expression and reducing TC, MMP-9, and Cat S levels. Acta Pharmacol Sin 31:1350–1358CrossRefGoogle Scholar
  14. 14.
    Zúñiga S, Molina H, Azocar L, Amigo L, Nervi F, Pimentel F, Jarufe N, Arrese M, Lammert F, Miquel JF (2008) Ezetimibe prevents cholesterol gallstone formation in mice. Liver Int 28:935–947CrossRefGoogle Scholar
  15. 15.
    Wang HH, Portincasa P, Mendez-Sanchez N, Uribe M, Wang DQ (2008) Effect of ezetimibe on the prevention and dissolution of cholesterol gallstones. Gastroenterology 134:2101–2110CrossRefGoogle Scholar
  16. 16.
    Muraoka T, Aoki K, Iwasaki T, Shinoda K, Nakamura A, Aburatani H, Mori S, Tokuyama K, Kubota N, Kadowaki T, Terauchi Y (2011) Ezetimibe decreases SREBP-1c expression in liver and reverses hepatic insulin resistance in mice fed a high-fat diet. Metabolism 60:617–628CrossRefGoogle Scholar
  17. 17.
    Palanisamy M, Aldars-García L, Gil-Ramírez A, Ruiz-Rodríguez A, Marín FR, Reglero G, Soler-Rivas C (2014) Pressurized water extraction of β-glucan enriched fractions with bile acids-binding capacities obtained from edible mushrooms. Biotechnol Progr 30:391–400CrossRefGoogle Scholar
  18. 18.
    Nitschke J, Altenbach HJ, Malolepszy T, Mölleken H (2011) A new method for the quantification of chitin and chitosan in edible mushrooms. Carbohydr Res 346:1307–1310CrossRefGoogle Scholar
  19. 19.
    Vandesompele J, De Preter K, Pattyn F, Poppe B, Van Roy N, De Paepe A, Speleman F (2002) Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. Genome Biol 3(7):1–12CrossRefGoogle Scholar
  20. 20.
    Pfaffl MW (2001) A new mathematical model for relative quantification in real-time RT–PCR. Nucleic Acids Res 29(9):e45CrossRefGoogle Scholar
  21. 21.
    Fukushima M, Nakano M, Morii Y, Ohashi T, Fujiwara Y, Sonoyama K (2010) Hepatic LDL receptor mRNA in rats is increased by dietary mushroom (Agaricus bisporus) fiber and sugar beet fiber. J Nutr 130:2151–2156Google Scholar
  22. 22.
    Ikemoto S, Takahashi M, Tsunoda N, Maruyama K, Itakura H, Kawanaka K, Tabata I, Higuchi M, Tange T, Yamamoto TT, Ezaki O (1997) Cholate inhibits high-fat diet-induced hyperglycemia and obesity with acyl-CoA synthetase mRNA decrease. Am J Physiol 273:E37–E45Google Scholar
  23. 23.
    Duan L-P, Wang HH, Wang DQ-H (2004) Cholesterol absorption is mainly regulated by the jejunal and ileal ATP-binding cassette sterol efflux transporters Abcg5 and Abcg8 in mice. J Lipid Res 45(7):1312–1323CrossRefGoogle Scholar
  24. 24.
    Field FJ, Born E, Mathur SN (1997) Effect of micellar β-sitosterol on cholesterol metabolism in CaCo-2 cells. J Lipid Res 38:348–360Google Scholar
  25. 25.
    Plat J, Mensink RP (2002) Increased intestinal ABCA1 expression contributes to the decrease in cholesterol absorption after plant stanol consumption. FASEB J 16(10):1248–1253CrossRefGoogle Scholar
  26. 26.
    Field FJ, Born E, Mathur SN (2004) LXR/RXR ligand activation enhances basolateral efflux of β-sitosterol in CaCo-2 cells. J Lipid Res 45:905–913CrossRefGoogle Scholar
  27. 27.
    Petruzzelli M, Groen AK, van Erpecum KJ, Vrins C, van der Velde AE, Portincasa P, Palasciano G, van Berge Henegouwen GP, Lo Sasso G, Morgano A, Moschetta A (2009) Micellar lipid composition profoundly affects LXR-dependent cholesterol transport across CaCo2 cells. FEBS Lett 583:1274–1280CrossRefGoogle Scholar
  28. 28.
    EFSA Panel on Dietetic Products, Nutrition and Allergies (NDA) (2010) Scientific Opinion on the substantiation of health claims related to plant sterols and plant stanols and maintenance of normal blood cholesterol concentrations (ID 549, 550, 567, 713, 1234, 1235, 1466, 1634, 1984, 2909, 3140), and maintenance of normal prostate size and normal urination (ID 714, 1467, 1635) pursuant to Article 13(1) of Regulation (EC) No 1924/2006. EFSA J 8:1813–1834. Available online:
  29. 29.
    Reagan-Shaw S, Nihal M, Ahmad N (2008) Dose translation from animal to human studies revisited. FASEB J 22:659–661CrossRefGoogle Scholar
  30. 30.
    Verbraecken J, Van de Heyning P, De Backer W, Van Gaal L (2006) Body surface area in normal-weight, overweight, and obese adults. A comparison study. Metabolism 5:515–524CrossRefGoogle Scholar
  31. 31.
    EFSA Panel on Dietetic Products, Nutrition and Allergies (NDA) (2008) Scientific opinion of the panel on dietetic products nutrition and allergies on a request from Unilever PLC/NV on plant sterols and lower/reduced blood cholesterol, reduced the risk of (coronary) heart disease. EFSA J 781:1–12. Available online:
  32. 32.
    Plat J, Mensink RP (2001) Effects of plant stanol esters on LDL receptor protein expression and on LDL receptor and HMG-CoA reductase mRNA expression in mononuclear blood cells of healthy men and women. FASEB J 16(2):258–260Google Scholar
  33. 33.
    Calpe-Berdiel L, Escolΰ-Gil JC, Ribas V, Navarro-Sastre A, Garcιs-Garcιs J, Blanco-Vaca F (2005) Changes in intestinal and liver global gene expression in response to a phytosterol-enriched diet. Atherosclerosis 181(1):75–85CrossRefGoogle Scholar
  34. 34.
    Fukushima M, Ohashi T, Fujiwara Y, Sonoyama K, Nakano M (2001) Cholesterol-lowering effects of maitake (Grifola frondosa) fiber, shiitake (Lentinus edodes) fiber, and enokitake (Flammulina velutipes) fiber in rats. Exp Biol Med 226:758–765Google Scholar
  35. 35.
    Gil-Ramirez A, Soler-Rivas C (2014) The use of edible mushroom extracts as bioactive ingredients to design novel functional foods with hypocholesterolemic activities. In: GrégoirePesti (ed) Mushrooms: cultivation, antioxidant properties and health benefits. Nova Science Publishers, Inc. Hauppauge, NY, pp 43–73Google Scholar
  36. 36.
    Bobek P, Ozdín L, Kuniak L (1997) Effect of oyster mushroom and isolated β-glucan on lipid peroxidation and on the activities of antioxidative enzymes in rats fed the cholesterol diet. J Nut Biochem 8:469–471CrossRefGoogle Scholar
  37. 37.
    Carter SJ, Roberts MB, Salter J, Eaton CB (2010) Relationship between Mediterranean Diet Score and atherothrombotic risk: findings from the Third National Health and Nutrition Examination Survey (NHANES III), 1988–1994. Atherosclerosis 210:630–636CrossRefGoogle Scholar
  38. 38.
    Carter B, Taylor O, Prendergast D, Zimmerman T, Von Furstenberg R, Moore D, Karpen S (2007) Stigmasterol, a soy lipid–derived phytosterol, is an antagonist of the bile acid Nnuclear receptor FXR. Pediatr Res 62(3):301–306CrossRefGoogle Scholar
  39. 39.
    Yang C, Yu L, Li W, Xu F, Cohen JC, Hobbs HH (2004) Disruption of cholesterol homeostasis by plant sterols. J Clin Invest 114(6):813–822CrossRefGoogle Scholar
  40. 40.
    Yang C, McDonald JG, Patel A, Zhang Y, Umetani M, Xu F, Westover EJ, Covey DF, Mangelsdorf DJ, Cohen JC, Hobbs HH (2006) Sterol intermediates from cholesterol biosynthetic pathway as liver X receptor ligands. J BiolChem 281(38):27816–27826Google Scholar
  41. 41.
    Cui J, Huang L, Zhao A, Lew J-L, Yu J, Sahoo S, Meinke PT, Royo I, Pelαez F, Wright SD (2003) Guggulsterone is a farnesoid X receptor antagonist in coactivator association assays but acts to enhance transcription of bile salt export pump. J BiolChem 278(12):10214–10220Google Scholar
  42. 42.
    Garcia-Calvo M, Lisnock J, Bull HG, Hawes BE, Burnett DA, Braun MP, Crona JH, Davis HR, Dean DC, Detmers PA, Graziano MP, Hughes M, MacIntyre DE, Ogawa A, O’Neill KA, Iyer SPN, Shevell DE, Smith MM, Tang YS, Makarewicz AM, Ujjainwalla F, Altmann SW, Chapman KT, Thornberry NA (2005) The target of ezetimibe is Niemann-Pick C1-Like 1 (NPC1L1). Proc Natl Acad Sci USA 102(23):8132–8137CrossRefGoogle Scholar
  43. 43.
    Watanabe M, Houten SM, Wang L, Moschetta A, Mangelsdorf DJ, Heyman RA, Moore DD, Auwerx J (2004) Bile acids lower triglyceride levels via a pathway involving FXR, SHP, and SREBP-1c. J Clin Invest 113:1408–1418CrossRefGoogle Scholar
  44. 44.
    Kanaya N, Kubo M, Liu Z, Chu P, Wang C, Yuan YC, Chen S (2011) Protective effects of white button mushroom (Agaricus bisporus) against hepatic steatosis in ovariectomized mice as a model of postmenopausal women. PLoS ONE 6:e26654CrossRefGoogle Scholar
  45. 45.
    Inoue N, Inafuku M, Shirouchi B, Nagao K, Yanagita T (2013) Effect of Mukitake mushroom (Panellus serotinus) on the pathogenesis of lipid abnormalities in obese, diabetic ob/ob mice. Lipids Health Dis 12:18CrossRefGoogle Scholar
  46. 46.
    Inafuku M, Nagao K, Nomura S, Shirouchi B, Inoue N, Nagamori N, Nakayama H, Toda T, Yanagita T (2012) Protective effects of fractional extracts from Panellus serotinus on non-alcoholic fatty liver disease in obese, diabetic db/db mice. Br J Nutr 107:639–646CrossRefGoogle Scholar
  47. 47.
    Volger OL, van der Boom H, de Wit ECM, van Duyvenvoorde W, Hornstra G, Plat J, Havekes LM, Mensink RP, Princen HMG (2001) Dietary plant stanol esters reduce VLDL cholesterol secretion and bile saturation in apolipoprotein E*3-Leiden transgenic mice. Arterioscler Thromb Vasc Biol 21(6):1046–1052CrossRefGoogle Scholar
  48. 48.
    Igel M, Giesa U, Lόtjohann D, von Bergmann K (2003) Comparison of the intestinal uptake of cholesterol, plant sterols, and stanols in mice. J Lipid Res 44(3):533–538CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Alicia Gil-Ramírez
    • 1
  • Víctor Caz
    • 2
  • Roberto Martin-Hernandez
    • 3
  • Francisco R. Marín
    • 1
  • Carlota Largo
    • 2
  • Arantxa Rodríguez-Casado
    • 3
  • María Tabernero
    • 2
  • Alejandro Ruiz-Rodríguez
    • 1
  • Guillermo Reglero
    • 1
    • 3
  • Cristina Soler-Rivas
    • 1
  1. 1.Department of Production and Characterization of Novel Foods, CIAL – Research Institute in Food Science (UAM+CSIC)Universidad Autónoma de MadridMadridSpain
  2. 2.Department of Experimental SurgeryResearch Institute Hospital La Paz (IdiPAZ)MadridSpain
  3. 3.IMDEA Food InstitutePabellón Central del Antiguo Hospital de Cantoblanco (Edificio no 7)MadridSpain

Personalised recommendations