![[Feedback]](/icons/banner/feedbackACT.gif){kind=icon}
![[For Subscribers]](/icons/banner/subscriberACT.gif){kind=icon}
![[Archive]](/icons/banner/archiveACT.gif){kind=icon}
![[Search]](/icons/banner/searchACT.gif){kind=icon}
![[Contents]](/icons/banner/tocACT.gif){kind=icon}
|
|
|
|
|
||
* Servicio de Bioquímica-Investigación, Hospital Ramón y Cajal, Madrid, Spain; Instituto de Fermentaciones Industriales, CSIC, Madrid, Spain; and ** Departamento de Bioquímica y Biología Molecular, Universidad de Alcalá, Madrid, Spain
4 To whom correspondence should be addressed. E-mail: miguel.a.lasuncion{at}hrc.es.
 |
ABSTRACT |
|---|
 TOP ABSTRACT MATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
KEY WORDS: • LDL receptor • red grape juice polyphenols • cholesterol trafficking • LDL cholesterol
Polyphenols, a ubiquitous group of secondary plant metabolites, represent a large group of natural antioxidants abundant in fruits, vegetables, and beverages such as grape juice, wine, and tea. Several studies have linked wine consumption to a reduction in cardiovascular diseases (1–3), attributing some of the cardioprotective effects to the polyphenols present in red wine and grape products (4). Oxidation of LDL was proposed to play a crucial role in atherogenesis (5), and polyphenols were shown to exhibit strong antioxidant properties that could protect LDL from oxidation (6–9).
In addition to their antioxidant activity, polyphenols also possess many different biological properties that may contribute to their cardioprotective effects, including the ability to inhibit platelet activity and thrombosis (10–12) and the potential to reduce plasma lipids. In vivo studies showed that administration of green tea (13), dealcoholized red wine (8), grape seed procyanidins (14), naringin (15), or polymethoxylated flavones (16) lowered plasma cholesterol in laboratory animals with diet-induced hypercholesterolemic. Recently, reduction of cholesterol LDL plasma concentration was reported in women consuming a lyophilized grape powder (17) or red wine (18). The mechanisms underling this hypolipidemic effect may involve the upregulation of LDL receptor expression (19,20), inhibition of hepatic lipid synthesis (21) and lipoprotein secretion (22,23), or increased cholesterol elimination via bile acids (14).
Intracellular cholesterol homeostasis is under the control of the sterol regulatory element-binding protein (SREBP)5 transcription factor (24,25). Once LDL is internalized by the LDL receptor, endosomes fuse with lysosomes and LDL cholesteryl esters are hydrolyzed. Then, free cholesterol is released into the cytoplasm and reaches the different intracellular compartments. Free cholesterol in the endoplasmic reticulum (ER) can be converted to cholesteryl ester by the enzyme acyl-CoA cholesterol acyltransferase (ACAT), transported to the plasma membrane, or used for other metabolic purposes. Free cholesterol in the ER has a negative feedback effect on cholesterol biosynthesis and expression of the LDL receptor via SREBP (24,25). To elucidate the mechanism that accounts for the hypocholesterolemic effect of red grape juice consumption, the present study addresses the effects of red grape juice on intracellular cholesterol homeostasis in human cell lines in vitro.
|
MATERIALS AND METHODS |
|---|
|
TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
Cell culture. Human hepatocarcinoma HepG2 cells (ATCC HB 8065) obtained from the American Type Culture Collection were maintained in DMEM supplemented with 10% FBS and antibiotics, at 37°C in a humidified atmosphere containing 5% CO2. For experiments, cells were incubated in DMEM containing 10% LPDS, supplemented or not with human LDL and/or RGJ. On other occasions, pure antioxidants, lovastatin, and/or compound S-58035 were also added to the incubation medium. For controls, cells were supplemented with glucose and fructose, each at a concentration of 2.1 g/L of medium.
Analysis of polyphenols in red grape juice. Analysis of flavonoids, phenolic acids, and flavan-3-ols was performed following a previously described method (26). Briefly, 5 mL of RGJ (68° Brix) was diluted with 15 mL of water, separated into 2 aliquots and extracted 3 times each with diethyl ether and ethyl acetate. The organic fractions were mixed and dried with anhydrous Na2SO4 for 30 min, concentrated to dryness and dissolved in 1 mL of methanol/water (50:50). After being filtered through a 0.45-µm membrane, a 100 µL of sample was injected into a reverse-phase Nova-Pak C18 column (300 mm x 3.9 mm, 4µm), using a Waters liquid chromatography system equipped with a 600-MS controller, a 717Plus autosampler, and a 996 photodiode-array detector (26). Detection was performed by scanning from 220 to 380 nm. For anthocyanins, 100 µL of diluted RGJ, previously filtered through a 0.45-µm membrane, was injected into a reverse-phase Nova-Pak C18 column (150 mm x 3.9 mm, 4 µm), and separated by a multistep linear gradient method, as described (27). Detection was performed by scanning from 260 to 600 nm.
Uptake of RGJ polyphenols by HepG2 cells. HepG2 cells were cultured in 175-cm2 flasks in DMEM containing 10% FBS to reach 80% confluence; then the medium was changed to DMEM containing 10% LPDS, LDL (0.31 mmol cholesterol/L), and RGJ (5 mL/L). At 0, 3, 9, and 20 h of incubation, cells were harvested, washed with PBS, and treated on ice with 300 µL of lysis buffer (Cell Signaling) for 20 min. The cell protein content was determined [bicinchoninic acid (BCA) protein assay kit, Pierce]. Lysates were incubated with 200 µL of 0.1 mol/L ascorbic acid, 100 µL of 0.78 mol/L sodium acetate buffer (pH 4.8), and 100 µL ß-glucuronidase (131100 kU/L ß-glucuronidase and 1070 kU/L sulfatase) from Helix pomatia at 37°C for 8 h. The mixture was extracted twice with 1 mL of ethyl acetate, and pooled supernatants were concentrated to dryness with a SPD121P Speed-Vac system (Thermo Electron). The sample was reconstituted in 300 µL of methanol:water (50:50, v:v), filtered through a 0.45-µm membrane, and 200 µL were subjected to HPLC analysis as described (26).
LDL receptor activity and cell-surface expression. Human LDL was isolated by ultracentrifugation and labeled with the fluorescent probe DiI as described elsewhere (28). HepG2 cells (4 x 105 cells/well), treated with the test compounds for 20 h in 10% LPDS, were washed twice with PBS and then incubated in duplicate in 10% LPDS medium containing DiI-LDL (0.10 mmol cholesterol/L) for 2 h at 37°C to allow DiI-LDL uptake. Nonspecific DiI-LDL uptake was assessed in a third well containing a 50-fold excess of unlabeled LDL. Cells were washed and analyzed by flow cytometry (FACScalibur, BD Biosciences).
To analyze cell-surface expression of the LDL receptor protein, cells were incubated with anti-LDL receptor monoclonal antibody IgG C7 (15 mg/L) and then FITC-labeled goat anti-mouse IgG, and analyzed by flow cytometry as described previously (29).
Analysis of cholesterol biosynthesis and lipid determinations. To examine cholesterol biosynthesis, HepG2 cells (6 x 106 cells) were incubated in 10% LPDS medium containing [14C]-acetate (1.5 mCi/L) for 20 h, lysed with 10% KOH, and total cellular de novo cholesterol was analyzed by reverse-phase HPLC as described previously (30). Cell total cholesterol and free cholesterol were measured using an enzymatic method followed by HPLC analysis using cholesterol oxidase and cholesterol esterase as described previously (31). For free cholesterol, cholesterol esterase was not included in the reaction mixture; esterified cholesterol was calculated by subtracting free cholesterol from total cholesterol. Triglycerides were measured using a colorimetric enzymatic kit (A. Menarini Diagnostics).
Quantitative real-time RT-PCR assay. Total RNA was extracted with Trizol reagent according to the manufacturer's instructions. Total RNA (2 µg) was reverse-transcribed using M-MLV RT enzyme (Promega). Real-time PCR amplification was performed on a LightCycler and data analysis software version 3.5 (Roche Diagnostics GmbH), using FastStart DNA Master SYBR Green I (Roche Diagnostics GmbH). The primers used are listed in supplemental Table S1. Amplification cycles used were 95°C for 10 s, 60°C for 20 s, and 72°C for 20 s, except for ABCG1 and ABCG5, for which the annealing temperature was 62°C. Melting curves were evaluated for each gene, and PCR reaction products were separated on a 2% agarose gel and stained with ethidium bromide to confirm the presence of a single product. All analyses were performed in duplicate, and relative mRNA levels were determined using glyceraldehyde-3-phosphate dehydrogenase as a reference. The relative expression ratio (R) for each target gene was calculated taken into account the reaction efficiency (E) and the crossing point (CP) of the unknown sample vs. the control (Roche Diagnostics, LC relative quantification software, 2001). At least 4 independent experiments were performed in duplicate for each determination.
ACAT inhibition. ACAT inhibition with S-58035 was used as an indirect measure of free cholesterol reaching the ER. Cells (4 x 105 per well) were incubated in 10% LPDS-containing DMEM supplemented with LDL (0.31 mmol cholesterol/L), in the presence or absence of lovastatin (2.5 µmol/L), S-58035 (5 mg/L), and RGJ (5 mL/L) for 20 h. Cells were washed twice with PBS, and LDL receptor activity was assessed using the DiI-LDL uptake assay described above.
Western blot analysis of SREBP-1. After incubation, cells were washed with PBS and then treated with lysis buffer. Cell protein content was determined (BCA protein assay kit) and loaded equally (50 µg protein/lane) onto 10% polyacrylamide gels, electrophoresed, and transferred onto a nitrocellulose membrane. Membranes were blocked with 1% defatted milk powder in Tris-buffered saline (TBS; 25 mmol/L Tris pH 7.4, 136 mmol/L NaCl, 2.6 mmol/L KCl) containing 0.1% Tween 20, incubated overnight at 4°C with anti-mouse SREBP-1 (Santa Cruz Biotechnology) and then with horseradish peroxidase-conjugated secondary antibody, and developed with the enhanced chemiluminescence detection system (Amersham Biosciences). Nuclear extracts were obtained as described previously (32) and Western blot was performed as described above, with 100 µg protein loaded per lane. To inhibit rapid degradation of nuclear fragments of SREBPs, cells were treated with N-acetyl-leucinal-leucinal-norleucinal at a final concentration of 25 mg/L 2 h before protein extraction and throughout the extraction procedure.
Fluorescence microscopy. HepG2 cells were cultured on 12-mm glass coverslips in 24-well dishes for 48 h in medium containing 10% FBS. The medium was then replaced with medium containing 10% LPDS and incubated in the presence of DiI-LDL (0.155 mmol cholesterol/L), RGJ (5 mL/L), or tamoxifen (5 µmol/L) for 20 h. For staining of free cholesterol, cells were washed and fixed in 4% paraformaldehyde for 15 min, washed 3 times with PBS and then stained with filipin (50 mg/L in PBS) for 45 min. Immunostaining was performed as previously described (33), using anti-CD63 (1:100) and Alexa-conjugated anti-mouse IgG (1:400) antibodies. DNA was stained with DAPI. Cells were mounted for microscopy and examined on an Olympus BX51 reflected fluorescence microscope. Merged images were prepared using Adobe Photoshop 6.0.
Statistical analysis. All values are presented as means ± SEM. Means were compared using Student's t test or ANOVA followed by post hoc multiple comparisons. Comparisons of cholesterol biosynthesis data were made by 1-way repeated measures ANOVA and pairwise comparisons by the Student-Newman-Keuls method. The interactions between RGJ and LDL on LDL receptor activity were analyzed by two-way ANOVA. When the interaction was significant, the subsequent analysis of the effect of RGJ was performed by 1-way ANOVA separately for each group (presence of LDL in the medium). Comparisons of gene expression data were made either by the confidence interval method (for each experimental condition vs. the control) or by 1-way ANOVA and Tukey's HSD post hoc test (for comparisons between the experimental conditions); in the latter case, data from the control condition were excluded from the analyses. Differences with P-values < 0.05 were considered significant. Analyses were performed with Statgraphics Plus v5.0 (Statistical Graphics).
|
RESULTS |
|---|
|
TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
|
|
|
|
To confirm that the effects of RGJ were not mediated by its antioxidant activity, other antioxidants were also evaluated. BHT (1–10 µmol/L) and Trolox (0.1–10 µmol/L) did not stimulate LDL receptor activity in cells exposed to LDL (data not shown), confirming that the effect of RGJ polyphenols cannot be attributed to antioxidant activity.
Effects of RGJ on cellular lipid mass and cholesterol biosynthesis. To explore whether changes in LDL receptor activity and cell-surface expression are correlated with cell lipid content, both cell cholesterol and triglycerides were quantified. Incubation of HepG2 cells in the presence of LDL significantly increased the cell total cholesterol content, whereas the effect was proportionally much higher for esterified cholesterol (403% of the control) than for free cholesterol (125% of the control) (Table 3). RGJ did not have any detectable effect on cell total cholesterol content in cells incubated in medium lacking LDL. However, in the presence of LDL, RGJ significantly increased total cholesterol. This was confirmed by oil red O staining for neutral lipids (data not shown). Analysis of free and esterified cholesterol revealed significant changes only in esterified cholesterol (149% of the control). Although incubation with LDL did not cause appreciable changes in the triglyceride content of the cells, RGJ caused a significant reduction either in the presence (78.4% of the control) or absence of LDL (81.7% of the control) (Table 3).
|
Effect of RGJ on gene expression. We next used quantitative RT-PCR to assess relative changes in the expression of selected genes related to lipid metabolism (Table 4). In cells incubated in the absence of LDL, RGJ significantly reduced the expression of ABCA1, ABCG8, and apoB, whereas the other genes were unaffected. LDL repressed the expression of the LDL receptor, hydroxymethylglutaryl-CoA (HMG-CoA) reductase, and NPC1L1, which are all involved in cholesterol accretion. LDL did not affect the expression of the other genes examined. The simultaneous addition of RGJ abrogated the effect of LDL on the expression of the LDL receptor, partially counteracted the effect on HMG-CoA reductase, and markedly reduced the expression of CYP7A1 and apoB (Table 4).
|
|
|
|
|
|
DISCUSSION |
|---|
|
TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
The LDL receptor and most of the enzymes involved in the cholesterol biosynthesis pathway are regulated at the transcriptional level by a family of membrane-bound transcription factors, the SREBPs (24). In response to a decrease in free cholesterol availability in the ER, SREBP is processed proteolytically, and the active fragment migrates to the nucleus where it interacts with the sterol response elements present in the promotor of those genes and activates their transcription (24). Consistent with this, we observed that incubation of cells with LDL was accompanied by a reduction in SREBP-1 processing. Interestingly, simultaneous treatment with RGJ prevented this effect of LDL. Very recently, Kuhn et al. (19) proposed that the upregulation of LDL receptor expression induced by green tea polyphenols is mediated by inhibition of ubiquitin/proteosome-mediated degradation of active SREBP-2. It remains to be seen whether this mechanism also applies to RGJ polyphenols. Nevertheless, the increased levels of active SREBP are likely to be responsible for the effect of RGJ on the expression of the LDL receptor and HMG-CoA reductase.
In contrast to the LDL receptor and enzymes involved in cholesterol biosynthesis, the expression of genes involved in cholesterol efflux is induced in response to cholesterol loading (38,39). In cells incubated in the presence of LDL, we found that RGJ caused repression of ABCA1 and ABCG5, which are involved in cholesterol efflux, CYP7A1, which directs cholesterol to the synthesis of bile acids, and apoB, which is required for lipoprotein synthesis and secretion. This suggests that RGJ interferes with the intracellular availability of free cholesterol for regulatory purposes. To study how LDL-derived free cholesterol reached the ER, cells were treated with S-58035, a specific inhibitor of ACAT. As expected, LDL inhibited LDL receptor activity, and S-58035 further decreased this activity, a finding that is in agreement with the results of previous studies (40,41). In contrast, S-58035 did not have any effect on RGJ-treated cells, which displayed increased LDL receptor activity. Taken together, these results suggest that RGJ compromises the availability of LDL-derived free cholesterol in the ER, thus explaining the counteracting effect of RGJ in the response to LDL.
Next, we addressed whether RGJ affects the endocytic pathway. After being endocytosed, LDL enters distinct endosomal compartments: first, it enters the early endosome compartments and then the late endosome/lysosome compartments, where LDL cholesteryl ester is hydrolyzed by acid lipase (42,43). It was reported that disruption of intracellular cholesterol trafficking induces upregulation of LDL receptor expression in human hepatocytes (29,44,45). Treatment with tamoxifen, for instance, was shown to exert a strong stimulatory effect on LDL receptor expression in cells exposed to LDL, an effect that was attributed to the retention of LDL-derived free cholesterol in the endosomal/lysosomal compartment (29). In the present study, the filipin staining pattern in cells treated with RGJ was weak and diffuse, quite different from that of tamoxifen-treated cells, which showed intensely stained enlarged vesicles (Fig. 5, F). Because DiI-LDL uptake was similar in the presence of either tamoxifen or RGJ, we conclude that the effect of RGJ was not mediated by the retention of free cholesterol in endosome/lysosome vesicles.
Finally, we analyzed the nature of the LDL-accumulating compartment by examining the expression of CD63, a protein localized to late endosomes (34). Most vesicles containing DiI-LDL also showed CD63 labeling in control and tamoxifen-treated cells, whereas this colocalization was not observed in cells treated with RGJ (Fig. 6, F). These results indicate that RGJ interferes with LDL trafficking through the endocytic pathway, probably by preventing the fusion of lysosomes with endosomes. Based on current understanding of this pathway (43,46), this effect would delay lipolytic processing of LDL and reduce the efflux of lipoprotein cholesterol to the ER, thus explaining the counteracting effect of RGJ on gene expression in cells exposed to LDL.
Epidemiologic and clinical studies suggest that polyphenols are good candidates to explain the protective effects of vegetables, fruits, and red wine against certain cancers and cardiovascular diseases (3,4,12,47). In addition to their antioxidant (6,7), vasorelaxant (48), and anti-inflammatory (17) properties, the effects on cholesterol homeostasis shown here may contribute to the cardioprotective effects of Vitis vinifera polyphenols. Although we were not able to attribute the effects of RGJ to a single polyphenol or family of polyphenols present in RGJ, the variety of polyphenols present in the RGJ concentrate, including flavonols, anthocyanins, flavan-3-ols, procyanidins and phenolic acids, and the limited availability of pure compounds, make the search for the active compound(s) a complex task. The finding that other potent antioxidants do not mimic the effect of RGJ indicates that antioxidant protection is not responsible for the effects of RGJ on LDL receptor activity. Our observation that RGJ interferes with the endocytic pathway offers an alternative mechanism to account for the activity of grape polyphenols.
|
ACKNOWLEDGMENTS |
|---|
|
FOOTNOTES |
|---|
2 Supplemental Table 1 and supplemental references are available with the online posting of this paper at www.nutrition.org.
3 Present address: Department of Pharmacology, Boyer Center for Molecular Medicine, Yale University, School of Medicine, New Haven, CT 06536-0812.
5 Abbreviations used: ACAT, acyl-CoA cholesterol acyltransferase; apo B, apolipoprotein B; BCA, bicinchoninic acid; DiI,1,1'-dioctadecyl-3,3,3,3'-tetramethylindocarbocyanineperchlorate; ER, endoplasmic reticulum; FBS, fetal bovine serum; HMG-CoA, hydroxymethylglutaryl-CoA; LPDS, lipoprotein-deficient serum; RGJ, concentrated red grape juice; SREBP, sterol regulatory element-binding protein.
Manuscript received 21 December 2005. Initial review completed 8 February 2006. Revision accepted 9 April 2006.
|
LITERATURE CITED |
|---|
|
TOPABSTRACTMATERIALS AND METHODSRESULTSDISCUSSIONLITERATURE CITED |
|---|
1. Renaud S, de Lorgeril M. Wine, alcohol, platelets, and the French paradox for coronary artery heart disease. Lancet. 1992;339:1523–6.[Medline]
2. Rimm E, Giovannucci E, Willet W, Coldits G. Acheriori A, Rosner B, Stampfer M. Prospective study of alcohol consumption and risk of coronary disease in men. Lancet. 1991;338:464–8.[Medline]
3. Hertog MG, Feskens EJ, Hollman PC, Katan MB, Kromhout D. Dietary antioxidant flavonoids and risk of coronary heart disease: the Zutphen Elderly Study. Lancet. 1993;342:1007–11.[Medline]
4. Folts JD. Potential health benefits from the flavonoids in grape products on vascular disease. Adv Exp Med Biol. 2002;505:95–111.[Medline]
5. Witztum JL, Steinberg D. Role of oxidized low density lipoprotein in atherogenesis. J Clin Invest. 1991;88:1785–92.[Medline]
6. De Whalley CV, Rankin SM, Hoult RS, Jessup W, Leake DS. Flavonoids inhibit the oxidative modification of low density lipoproteins by macrophages. Biochem Pharmacol. 1990;39:1743–50.[Medline]
7. Frankel EN, Kanner J, German JB, Parks E, Kinsella JE. Inhibition of oxidation of human low-density lipoprotein by phenolic substances in red wine. Lancet. 1993;341:454–7.[Medline]
8. Vinson JA, Teufel K, Wu N. Red wine, dealcoholized red wine, and especially grape juice, inhibit atherosclerosis in hamster model. Atherosclerosis. 2001;156:67–72.[Medline]
9. O'Byrne DJ, Devaraj S, Grundy SM, Jialal I. Comparison of antioxidant effects of Concord grape juice flavonoides and 
-tocopherol on markers of oxidative stress in healthy adults. Am J Clin Nutr. 2002;76:1367–74.
10. Demrow H, Slane PR, Folts JD. Administration of wine and grape juice inhibits in vivo platelet activity and thrombosis in stenosed canine coronary arteries. Circulation. 1995;91:1182–88.
11. Osman HE, Maalej N, Shanmuganayagam D, Folts JD. Grape juice but not orange or grapefruit juice inhibits platelet activity in dogs and monkey. J Nutr. 1998;128:2307–12.
12. Freedman JE, Parker, III C, Li L, Perlman JA, Frei B, Ivanov V, Deak LR, Iafrati MD, Folts JD. Select flavonoids and whole juice from purple grapes inhibit platelet function and enhance nitric oxide release. Circulation. 2001;103:2792–98.
13. Yang TT, Koo MWL. Chinese green tea lowers cholesterol level through an increase in fecal lipid excretion. Life Sci. 2000;66:411–23.[Medline]
14. Del Bas JM, Fernández-Larrea J, Blay M, Ardevol A, Salvadó MJ, Arola L, Blade C. Grape seed procyanidins improve atherosclerotic risk index and induce liver CYP7A1 and SHP expression in healthy rats. FASEB J. 2005;19:479–81.
15. Kim HJ, Oh GT, Park YB, Lee MK, Seo HJ, Choi MS. Naringin alters the cholesterol biosynthesis and antioxidant enzyme activities in LDL receptor-knockout mice under cholesterol fed condition. Life Sci. 2004;74:1621–34.[Medline]
16. Kurowska EM, Manthey JA. Hypolipidemic effects and absorption of citrus polymethoxylated flavones in hamsters with diet-induced hypercholesterolemia. J Agric Food Chem. 2004;52:2879–86.[Medline]
17. Zern TL, Wood RJ, Greene C, West KL, Liu Y, Aggarwal D, Shachter NS, Fernández ML. Grape polyphenols exert a cardioprotective effect in pre- and postmenopausal women by lowering plasma lipids and reducing oxidative stress. J Nutr. 2005;135:1911–7.
18. Naissides M, Mamo JCL, James AP, Pal S. The effect of chronic consumption of red wine on cardiovascular disease risk factors in postmenopausal women. Atherosclerosis. 2006;185:438–45.[Medline]
19. Kuhn DJ, Burns AC, Kazi A, Dou QP. Direct inhibition of the ubiquitin-proteosome pathway by ester bond-containing green tea polyphenols is associated with increased expression of sterol regulatory element-binding protein 2 and LDL receptor. Biochim Biophys Acta. 2004;1682:1–10.[Medline]
20. Pal S, Ho N, Santos C, Dubois P, Mamo J, Croft K, Allister E. Red wine polyphenolics increase LDL receptor expression and activity and suppress the secretion of apoB100 from human HepG2 cells. J Nutr. 2003;133:700–6.
21. Theriault A, Wang Q, Van Iderstine SC, Chen B, Franke AA, Adeli K. Modulation of hepatic lipoprotein synthesis and secretion by taxifolin, a plant flavonoid. J Lipid Res. 2000;41:1969–79.
22. Casaschi A, Wang Q, Dang K, Richards A, Theriault AG. Intestinal apolipoprotein B secretion is inhibited by the flavonoid quercetin: potential role of microsomal triglyceride transfer protein and diacylglycerol acyltransferase. Lipids. 2002;37:647–52.[Medline]
23. Borradaile NM, de Dreu LE, Barrett PHR, Behrsin CD, Huff MW. Hepatocyte apoB-containing lipoprotein secretion is decreased by grapefruit flavonoid, naringenin, via inhibition of MTP-mediated microsomal triglyceride accumulation. Biochemistry. 2003;42:1283–91.[Medline]
24. Brown MS, Goldstein JL. The SREBP pathway: regulation of cholesterol metabolism by proteolysis of a membrane-bound transcription factor. Cell. 1997;89:331–40.[Medline]
25. Horton JD, Goldstein JL, Brown MS. SREBPs: activators of the complete program of cholesterol and fatty acids synthesis in the liver. J Clin Invest. 2002;109:1125–31.[Medline]
26. Monagas M, Gómez-Cordovés C, Bartolomé B. Evolution of polyphenols in red wines from Vitis vinifera L. during aging in the bottle. II. Non-anthocyanin phenolic compounds. Eur Food Res Technol. 2005;220:331–40.
27. Monagas M, Gómez-Cordovés C, Bartolomé B. Evolution of polyphenols in red wines from Vitis vinifera L. during aging in the bottle. I. Anthocyanins and pyranoanthocyanins. Eur Food Res Technol. 2005;220:607–14.
28. Calvo D, Gómez-Coronado D, Suárez Y, Lasunción MA, Vega MA. Human CD63 is a high affinity receptor for the native lipoproteins HDL, LDL, and VLDL. J Lipid Res. 1998;39:777–88.
29. Suárez Y, Fernández C, Gómez-Coronado D, Ferruelo AJ, Dávalos A, Martínez-Botas J, Lasunción MA. Synergistic upregulation of low-density lipoprotein receptor activity by tamoxifen and lovastatin. Cardiovasc Res. 2004;64:346–55.[Medline]
30. Fernández C, Suárez Y, Ferruelo AJ, Gómez-Coronado D, Lasunción MA. Inhibition of cholesterol biosynthesis by 
24-reductase in mammalian cells. Biochem J. 2002;366:109–19.[Medline]
31. Contreras JA, Castro M, Bocos C, Herrera E, Lasunción MA. Combination of an enzymatic method and HPLC for the quantitation of cholesterol in cultured cells. J Lipid Res. 1992;33:931–6.[Abstract]
32. Yin L, Zhang Y, Hillgartner FB. Sterol regulatory element-binding protein-1 interacts with the nuclear thyroid hormone receptor to enhance acetyl-CoA carboxylase-alpha transcription in hepatocytes. J Biol Chem. 2002;277:19554–65.
33. Suárez Y, González L, Cuadrado A, Berciano M, Lafarga M, Muñoz A. Kahalalide F, a new marine-derived compound, induces oncosis in human prostate and breast cancer cells. Mol Cancer Ther. 2003;2:863–72.
34. Escola JM, Kleijmeer MJ, Stoorvogel W, Griffith JM, Yoshie O, Geuze HJ. Selective enrichment of tetraspan proteins on the internal vesicles of multivesicular endosomes and on exosomes secreted by human B-lymphocytes. J Biol Chem. 1998;273:20121–27.
35. Brown MS, Goldstein JL. Lowering plasma cholesterol by raising LDL receptor. Atheroscler Suppl. 2004;5:57–9.[Medline]
36. Bursill C, Roach PD, Bottema CD, Pal S. Green tea upregulates the low-density lipoprotein receptor through the sterol-regulated element binding protein in HepG2 liver cells. J Agric Food Chem. 2001;49:5639–45.[Medline]
37. Bursill CA, Roach PD. Modulation of cholesterol metabolism by green tea polyphenol (-)-epigallocatechin gallate in cultured human liver (HepG2) cells. J Agric Food Chem. 2006;54:1621–6.[Medline]
38. Liao H, Langmann T, Schmitz G, Zhu Y. Native LDL upregulation of ATP-binding cassette transporter-1 in human vascular endothelial cells. Arterioscler Thromb Vasc Biol. 2002;22:127–32.
39. Hoekstra M, Out R, Kruijt JK, Van Eck M, Van Berkel TJC. Diet induced regulation of genes involved in cholesterol metabolism in rat liver parenchymal and Kupffer cells. J Hepatol. 2005;42:400–7.[Medline]
40. Tabas I, Weiland DA, Tall AR. Inhibition of acyl coenzyme A:cholesterol acyl transferase in J774 macrophages enhances down-regulation of the low density lipoprotein receptor and 3-hydroxy-3-methylglutaryl-coenzyme A reductase and prevents low density lipoprotein-induced cholesterol accumulation. J Biol Chem. 1986;261:3147–55.
41. Havekes LM, De Wit ECM, Princen HMG. Cellular free cholesterol in HepG2 cells is only partially available for down-regulation of low-density-lipoprotein receptor activity. Biochem J. 1987;247:739–46.[Medline]
42. Brown MS, Goldstein JL. A receptor-mediated pathway for cholesterol homeostasis. Science. 1986;232:34–47.
43. Sugii S, Reid PC, Ohgami N, Du H, Chang TY. Distinct endosomal compartment in early trafficking of low density lipoprotein-derived cholesterol. J Biol Chem. 2003;278:27180–89.
44. Issandou M, Guillard R, Boullay A-B, Linhart V, López-Pérez E. Up-regulation of low-density lipoproteins receptor in human hepatocytes induced by sequestration of free cholesterol in the endosomal/lysosomal compartments. Biochem Pharmacol. 2004;67:2281–89.[Medline]
45. Zhang J, Dudley-Rucker N, Crowley JR, López-Pérez E, Issandou M, Schaffer JE, Ory DS. The steroidal analog GW707 activates SREBP pathway through disruption of intracellular cholesterol trafficking. J Lipid Res. 2004;45:223–31.
46. Schroeder F, Gallegos AM, Atshaves BP, Storey SM, McIntosh AL, Petrescu AD, Huang H, Starodub O, Chao H, et al. Recent advances in membrane microdomains: rafts, caveolae, and intracellular cholesterol trafficking. Exp Biol Med (Maywood). 2001;226:873–90.
47. Arts IC, Hollman PC. Polyphenols and disease risk in epidemiologic studies. Am J Clin Nutr. 2005;81:317S–25.
48. Caimi G, Carollo C, Lo Presti R. Wine and endothelial function. Drugs Exp Clin Res. 2003;29:235–42.[Medline]
This article has been cited by other articles:
|
|
 |
|
  M. M. Dohadwala and J. A. Vita Grapes and Cardiovascular Disease J. Nutr., September 1, 2009; 139(9): 1788S - 1793S. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. Castilla, A. Davalos, J. L. Teruel, F. Cerrato, M. Fernandez-Lucas, J. L. Merino, C. C. Sanchez-Martin, J. Ortuno, and M. A Lasuncion Comparative effects of dietary supplementation with red grape juice and vitamin E on production of superoxide by circulating neutrophil NADPH oxidase in hemodialysis patients Am. J. Clinical Nutrition, April 1, 2008; 87(4): 1053 - 1061. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. Soundararajan, A. D. Wishart, H. P. V. Rupasinghe, M. Arcellana-Panlilio, C. M. Nelson, M. Mayne, and G. S. Robertson Quercetin 3-Glucoside Protects Neuroblastoma (SH-SY5Y) Cells in Vitro against Oxidative Damage by Inducing Sterol Regulatory Element-binding Protein-2-mediated Cholesterol Biosynthesis J. Biol. Chem., January 25, 2008; 283(4): 2231 - 2245. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. W. Finley, J. B. Burrell, and P. G. Reeves Pinto Bean Consumption Changes SCFA Profiles in Fecal Fermentations, Bacterial Populations of the Lower Bowel, and Lipid Profiles in Blood of Humans J. Nutr., November 1, 2007; 137(11): 2391 - 2398. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
