European Journal of Nutrition

, Volume 56, Issue 1, pp 225–236 | Cite as

Food supplementation with rice bran enzymatic extract prevents vascular apoptosis and atherogenesis in ApoE−/− mice

  • C. Perez-TerneroEmail author
  • M. D. Herrera
  • U. Laufs
  • M. Alvarez de Sotomayor
  • C. Werner
Original Contribution



Atherosclerosis is associated with reduced mononuclear cell (MNC) telomere length, and senescent cells have been detected in atherosclerotic plaques. Rice bran is a source of γ-oryzanol, phytosterols and tocols with potential lipid-lowering, antioxidant and anti-inflammatory activities. Here, we tested the hypothesis that rice bran enzymatic extract (RBEE) impacts on apoptosis, telomere length and atherogenesis in mice.


Seven-week-old male ApoE−/− mice were fed high-fat diet (HFD) or isocaloric HFD supplemented with 5 % (w/w) RBEE for 23 weeks. Wild-type mice of the same age were kept under standard diet as controls.


RBEE treatment reduced total cholesterol (19.24 ± 1.63 vs 24.49 ± 1.71 mmol/L) and triglycerides (1.13 ± 0.18 vs 1.75 ± 0.22 mmol/L) and augmented HDL-cholesterol (1.86 ± 0.20 vs 1.07 ± 0.20 mmol/L). RBEE attenuated macrophage infiltration by 56.69 ± 4.65 % and plaque development (7737 ± 836 vs 12,040 ± 1001 μm2) in the aortic sinus. In the aorta, RBEE treatment reduced expression of the apoptosis pathway components p16, p53 and bax/bcl-2 ratio. RBEE prevented apoptosis of aortic endothelial cells (2.81 ± 0.71–1.14 ± 0.35 apoptotic nuclei/ring for ApoE−/− HFD and ApoE−/− HFD 5 % RBEE, respectively). In contrast, MNC of RBEE-fed mice exhibited enhanced apoptosis marker expression with increased p53 and bax/bcl-2 protein levels. Compared to WT, ApoE−/− mice on HFD were characterized by significant telomere shortening in aorta (11 ± 2 %) and MNC (73 ± 7 %), which was reduced by supplementation with RBEE (aorta: 40 ± 7 %; MNC: 105 ± 10 %). Expression of telomere repeat-binding factor 2 was increased in RBEE-fed mice.


Long-term food supplementation with RBEE lowers cholesterol and prevents atherosclerotic plaque development in ApoE−/− mice. Differential regulation of vascular and MNC apoptosis and senescence were identified as potential mechanisms.


ApoE−/− Rice bran enzymatic extract Atherosclerosis Inflammation Apoptosis Telomeres 



The authors thank Dr. Juan Parrado (University of Seville) for supplying the rice bran extract (RBEE) and Jennifer Kieffer and Ellen Becker for excellent technical assistance. The study was funded by the Corona foundation via the Stifterverband für die Deutsche Wissenschaft and the Spanish Government. Cristina Perez-Ternero is a recipient of a FPU fellowship from the Spanish Government.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical standards

Approval for all experiments was provided by the Ethic Committee for Animal Experimentation of the University of Seville (Spain) (AGL2013-407791-P). The manuscript does not contain clinical studies or patient data.

Supplementary material

394_2015_1074_MOESM1_ESM.docx (22 kb)
Supplementary material 1 (DOCX 21 kb)


  1. 1.
    Minamino T, Komuro I (2007) Vascular cell senescence: contribution to atherosclerosis. Circ Res 100(1):15–26CrossRefGoogle Scholar
  2. 2.
    Lopez-Otin C, Blasco MA, Partridge L, Serrano M, Kroemer G (2013) The hallmarks of aging. Cell 153(6):1194–1217CrossRefGoogle Scholar
  3. 3.
    Fyhrquist F, Saijonmaa O, Strandberg T (2013) The roles of senescence and telomere shortening in cardiovascular disease. Nat Rev Cardiol 10(5):274–283CrossRefGoogle Scholar
  4. 4.
    Aviv A, Levy D (2012) Telomeres, atherosclerosis, and the hemothelium: the longer view. Annu Rev Med 63:293–301CrossRefGoogle Scholar
  5. 5.
    Werner C, Fürster T, Widmann T, Pöss J, Roggia C, Hanhoun M, Scharhag J, Büchner N, Meyer T, Kindermann W et al (2009) Physical exercise prevents cellular senescence in circulating leukocytes and in the vessel wall. Circulation 120(24):2438–2447CrossRefGoogle Scholar
  6. 6.
    Werner C, Gensch C, Pöss J, Haendeler J, Böhm M, Laufs U (2011) Pioglitazone activates aortic telomerase and prevents stress-induced endothelial apoptosis. Atherosclerosis 216(1):23–34CrossRefGoogle Scholar
  7. 7.
    Chen S, Lin J, Matsuguchi T, Blackburn E, Yeh F, Best LG, Devereux RB, Lee ET, Howard BV, Roman MJ et al (2014) Short leukocyte telomere length predicts incidence and progression of carotid atherosclerosis in American Indians: the strong heart family study. Aging (Albany NY) 6(5):414–427CrossRefGoogle Scholar
  8. 8.
    Huzen J, Peeters W, de Boer RA, Moll FL, Wong LS, Codd V, de Kleijn DP, de Smet BJ, van Veldhuisen DJ, Samani NJ et al (2011) Circulating leukocyte and carotid atherosclerotic plaque telomere length: interrelation, association with plaque characteristics, and restenosis after endarterectomy. Arterioscler Thromb Vasc Biol 31(5):1219–1225CrossRefGoogle Scholar
  9. 9.
    Brouilette SW, Moore JS, McMahon AD, Thompson JR, Ford I, Shepherd J, Packard CJ, Samani NJ (2007) Telomere length, risk of coronary heart disease, and statin treatment in the West of Scotland Primary Prevention Study: a nested case-control study. Lancet 369(9556):107–114CrossRefGoogle Scholar
  10. 10.
    O’Donnell CJ, Demissie S, Kimura M, Levy D, Gardner JP, White C, D’Agostino RB, Wolf PA, Polak J, Cupples LA et al (2008) Leukocyte telomere length and carotid artery intimal medial thickness: the Framingham Heart Study. Arterioscler Thromb Vasc Biol 28(6):1165–1171CrossRefGoogle Scholar
  11. 11.
    Willeit P, Willeit J, Brandstatter A, Ehrlenbach S, Mayr A, Gasperi A, Weger S, Oberhollenzer F, Reindl M, Kronenberg F et al (2010) Cellular aging reflected by leukocyte telomere length predicts advanced atherosclerosis and cardiovascular disease risk. Arterioscler Thromb Vasc Biol 30(8):1649–1656CrossRefGoogle Scholar
  12. 12.
    Baragetti A, Palmen J, Garlaschelli K, Grigore L, Pellegatta F, Tragni E, Catapano AL, Humphries SE, Norata GD, Talmud PJ (2015) Telomere shortening over 6 years is associated with increased subclinical carotid vascular damage and worse cardiovascular prognosis in the general population. J Intern Med 277(4):478–487CrossRefGoogle Scholar
  13. 13.
    Butt HZ, Atturu G, London NJ, Sayers RD, Bown MJ (2010) Telomere length dynamics in vascular disease: a review. Eur J Vasc Endovasc Surg 40(1):17–26CrossRefGoogle Scholar
  14. 14.
    Poch E, Carbonell P, Franco S, Diez-Juan A, Blasco MA, Andres V (2004) Short telomeres protect from diet-induced atherosclerosis in apolipoprotein E-null mice. FASEB J 18(2):418–420Google Scholar
  15. 15.
    Soory M (2012) Nutritional antioxidants and their applications in cardiometabolic diseases. Infect Disord Drug Targets 12(5):388–401CrossRefGoogle Scholar
  16. 16.
    Riccioni G, Speranza L, Pesce M, Cusenza S, D’Orazio N, Glade MJ (2012) Novel phytonutrient contributors to antioxidant protection against cardiovascular disease. Nutrition 28(6):605–610CrossRefGoogle Scholar
  17. 17.
    Chen G, Wang H, Zhang X, Yang ST (2014) Nutraceuticals and functional foods in the management of hyperlipidemia. Crit Rev Food Sci Nutr 54(9):1180–1201CrossRefGoogle Scholar
  18. 18.
    Goufo P, Trindade H (2014) Rice antioxidants: phenolic acids, flavonoids, anthocyanins, proanthocyanidins, tocopherols, tocotrienols, gamma-oryzanol, and phytic acid. Food Sci Nutr 2(2):75–104CrossRefGoogle Scholar
  19. 19.
    Rukmini C, Raghuram TC (1991) Nutritional and biochemical aspects of the hypolipidemic action of rice bran oil: a review. J Am Coll Nutr 10(6):593–601CrossRefGoogle Scholar
  20. 20.
    Islam MS, Nagasaka R, Ohara K, Hosoya T, Ozaki H, Ushio H, Hori M (2011) Biological abilities of rice bran-derived antioxidant phytochemicals for medical therapy. Curr Top Med Chem 11(14):1847–1853CrossRefGoogle Scholar
  21. 21.
    Friedman M (2013) Rice brans, rice bran oils, and rice hulls: composition, food and industrial uses, and bioactivities in humans, animals, and cells. J Agric Food Chem 61(45):10626–10641CrossRefGoogle Scholar
  22. 22.
    Parrado J, Miramontes E, Jover M, Gutierrez JF, de Teran L, Bautista J (2006) Preparation of a rice bran enzymatic extract with potential use as functional food. Food Chem 98(4):742–748CrossRefGoogle Scholar
  23. 23.
    Cawthon RM, Smith KR, O’Brien E, Sivatchenko A, Kerber RA (2003) Association between telomere length in blood and mortality in people aged 60 years or older. Lancet 361(9355):393–395CrossRefGoogle Scholar
  24. 24.
    Kim NW, Wu F (1997) Advances in quantification and characterization of telomerase activity by the telomeric repeat amplification protocol (TRAP). Nucleic Acids Res 25(13):2595–2597CrossRefGoogle Scholar
  25. 25.
    Werner C, Hanhoun M, Widmann T, Kazakov A, Semenov A, Pöss J, Bauersachs J, Thum T, Pfreundschuh M, Müller P et al (2008) Effects of physical exercise on myocardial telomere-regulating proteins, survival pathways, and apoptosis. J Am Coll Cardiol 52(6):470–482CrossRefGoogle Scholar
  26. 26.
    Martinet W, Kockx MM (2001) Apoptosis in atherosclerosis: focus on oxidized lipids and inflammation. Curr Opin Lipidol 12(5):535–541CrossRefGoogle Scholar
  27. 27.
    Revilla E, Santa-María C, Miramontes E, Bautista Lorite J, García-Martínez A, Cremades O, Cert R, Parrado J (2009) Nutraceutical composition, antioxidant activity and hypocholesterolemic effect of a water-soluble enzymatic extract from rice bran. Food Res Int 42:387–393CrossRefGoogle Scholar
  28. 28.
    Justo ML, Rodriguez-Rodriguez R, Claro CM, Alvarez de Sotomayor M, Parrado J, Herrera MD (2013) Water-soluble rice bran enzymatic extract attenuates dyslipidemia, hypertension and insulin resistance in obese Zucker rats. Eur J Nutr 52(2):789–797CrossRefGoogle Scholar
  29. 29.
    Justo ML, Candiracci M, Dantas AP, de Sotomayor MA, Parrado J, Vila E, Herrera MD, Rodriguez-Rodriguez R (2013) Rice bran enzymatic extract restores endothelial function and vascular contractility in obese rats by reducing vascular inflammation and oxidative stress. J Nutr Biochem 24(8):1453–1461CrossRefGoogle Scholar
  30. 30.
    Justo ML, Claro C, Vila E, Herrera MD, Rodriguez-Rodriguez R (2014) Microvascular disorders in obese Zucker rats are restored by a rice bran diet. Nutr Metab Cardiovasc Dis 24(5):524–531CrossRefGoogle Scholar
  31. 31.
    Candiracci M, Justo ML, Castano A, Rodriguez-Rodriguez R, Herrera MD (2014) Rice bran enzymatic extract-supplemented diets modulate adipose tissue inflammation markers in Zucker rats. Nutrition 30(4):466–472CrossRefGoogle Scholar
  32. 32.
    Ausman LM, Rong N, Nicolosi RJ (2005) Hypocholesterolemic effect of physically refined rice bran oil: studies of cholesterol metabolism and early atherosclerosis in hypercholesterolemic hamsters. J Nutr Biochem 16(9):521–529CrossRefGoogle Scholar
  33. 33.
    Wang YX, Li Y, Sun AM, Wang FJ, Yu GP (2014) Hypolipidemic and antioxidative effects of aqueous enzymatic extract from rice bran in rats fed a high-fat and-cholesterol diet. Nutrients 6(9):3696–3710CrossRefGoogle Scholar
  34. 34.
    Berger A, Rein D, Schafer A, Monnard I, Gremaud G, Lambelet P, Bertoli C (2005) Similar cholesterol-lowering properties of rice bran oil, with varied gamma-oryzanol, in mildly hypercholesterolemic men. Eur J Nutr 44(3):163–173CrossRefGoogle Scholar
  35. 35.
    Most MM, Tulley R, Morales S, Lefevre M (2005) Rice bran oil, not fiber, lowers cholesterol in humans. Am J Clin Nutr 81(1):64–68Google Scholar
  36. 36.
    Chen CW, Cheng HH (2006) A rice bran oil diet increases LDL-receptor and HMG-CoA reductase mRNA expressions and insulin sensitivity in rats with streptozotocin/nicotinamide-induced type 2 diabetes. J Nutr 136(6):1472–1476Google Scholar
  37. 37.
    Lakatta EG, Levy D (2003) Arterial and cardiac aging: major shareholders in cardiovascular disease enterprises: part I: aging arteries: a “set up” for vascular disease. Circulation 107(1):139–146CrossRefGoogle Scholar
  38. 38.
    Zhang W, Hui R, Yang S (2014) Telomeres, cardiovascular aging, and potential intervention for cellular senescence. Sci China Life Sci 57(8):858–862CrossRefGoogle Scholar
  39. 39.
    Nakagawa K, Eitsuka T, Inokuchi H, Miyazawa T (2004) DNA chip analysis of comprehensive food function: inhibition of angiogenesis and telomerase activity with unsaturated vitamin E, tocotrienol. BioFactors 21(1–4):5–10CrossRefGoogle Scholar
  40. 40.
    Eitsuka T, Nakagawa K, Miyazawa T (2006) Down-regulation of telomerase activity in DLD-1 human colorectal adenocarcinoma cells by tocotrienol. Biochem Biophys Res Commun 348(1):170–175CrossRefGoogle Scholar
  41. 41.
    Gizard F, Heywood EB, Findeisen HM, Zhao Y, Jones KL, Cudejko C, Post GR, Staels B, Bruemmer D (2011) Telomerase activation in atherosclerosis and induction of telomerase reverse transcriptase expression by inflammatory stimuli in macrophages. Arterioscler Thromb Vasc Biol 31(2):245–252CrossRefGoogle Scholar
  42. 42.
    von Zglinicki T (2002) Oxidative stress shortens telomeres. Trends Biochem Sci 27(7):339–344CrossRefGoogle Scholar
  43. 43.
    Parrado J, Miramontes E, Jover M, Marquez JC, Angeles Mejias M, Collantes De Teran L, Absi E, Bautista J (2003) Prevention of brain protein and lipid oxidation elicited by a water-soluble oryzanol enzymatic extract derived from rice bran. Eur J Nutr 42(6):307–314CrossRefGoogle Scholar
  44. 44.
    Santa-Maria C, Revilla E, Miramontes E, Bautista J, Garcia-Martinez A, Romero E, Carballo M, Parrado J (2010) Protection against free radicals (UVB irradiation) of a water-soluble enzymatic extract from rice bran. Study using human keratinocyte monolayer and reconstructed human epidermis. Food Chem Toxicol 48(1):83–88CrossRefGoogle Scholar
  45. 45.
    Brown AL, Zhu X, Rong S, Shewale S, Seo J, Boudyguina E, Gebre AK, Alexander-Miller MA, Parks JS (2012) Omega-3 fatty acids ameliorate atherosclerosis by favorably altering monocyte subsets and limiting monocyte recruitment to aortic lesions. Arterioscler Thromb Vasc Biol 32(9):2122–2130CrossRefGoogle Scholar
  46. 46.
    Dutta P, Courties G, Wei Y, Leuschner F, Gorbatov R, Robbins CS, Iwamoto Y, Thompson B, Carlson AL, Heidt T et al (2012) Myocardial infarction accelerates atherosclerosis. Nature 487(7407):325–329CrossRefGoogle Scholar
  47. 47.
    Henderson AJ, Ollila CA, Kumar A, Borresen EC, Raina K, Agarwal R, Ryan EP (2012) Chemopreventive properties of dietary rice bran: current status and future prospects. Adv Nutr 3(5):643–653CrossRefGoogle Scholar
  48. 48.
    de Nigris F, Franconi F, Maida I, Palumbo G, Anania V, Napoli C (2000) Modulation by alpha- and gamma-tocopherol and oxidized low-density lipoprotein of apoptotic signaling in human coronary smooth muscle cells. Biochem Pharmacol 59(11):1477–1487CrossRefGoogle Scholar
  49. 49.
    Khanduja KL, Avti PK, Kumar S, Mittal N, Sohi KK, Pathak CM (2006) Anti-apoptotic activity of caffeic acid, ellagic acid and ferulic acid in normal human peripheral blood mononuclear cells: a Bcl-2 independent mechanism. Biochim Biophys Acta 2:283–289CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  1. 1.Department of Pharmacology, School of PharmacyUniversity of SevilleSevilleSpain
  2. 2.Klinik für Innere Medizin III – Kardiologie, Angiologie und Internistische IntensivmedizinUniversitätsklinikum des SaarlandesHomburg/SaarGermany

Personalised recommendations