Background: Non-alcoholic fatty liver disease is considered a hepatic manifestation of the metabolic syndrome. It is associated with endothelial dysfunction as an early event of generalized atherosclerosis. However, it is unclear whether steatotic hepatocytes influence endothelial function directly. Objective: Explore the influence of hepatocyte steatosis on the function of endothelial cells. Methods: Oleic and palmitic acid (2:1 mixture, final concentration: 1 mM for 24 hours) was used to induce a normal adult hepatocyte strain (L-02) for transformation into steatosis cells. This was followed by oil red O staining and transmission electron microscopy (TEM) for verification. The culture solution of steatotic L-02 cells was filtered and collected, and added into the culture substrate of human umbilical vein endothelial cells (HUVECs). The expression of vascular cellular adhesion molecule -1 (VCAM-1), intercellular adhesion molecule-1 (ICAM-1), and E-selectin in HUVECs was detected by real-time polymerase chain reaction and Western blot assays. The apoptosis and proliferation of HUVECs was determined using flow cytometry. The experimental results were compared with the controls. Results: Oil red O staining and microscopic observation showed that the cytoplasm of induced L-02 cells contained a large amount of red lipid droplets. TEM results showed that the cytoplasm had lipid accumulation, swelling mitochondria, fewer cristae, and reduced number of rough endoplasmic reticula accompanied with degranulation. However, these changes were not observed in normal L-02 cells. As to the group of HUVECs treated by the filtrate of steatosis L-02 cells, the mRNA and protein expression of VCAM-1, ICAM-1, and E-selectin was higher than that in the control group. The difference was statistically significant (p <0.01). No significant difference was found when HUVECs apoptosis and proliferation were assessed by flow cytometry. Conclusion: Secretion from steatotic hepatocytes could boost the expression of VCAM-1, ICAM-1, and E-selectin in endothelial cells, indicating that hepatocyte steatosis could induce endothelial cell dysfunction. The proliferation and apoptosis of endothelial cells did not change, suggesting that hepatocyte steatosis had no influence on the viability of endothelial cells under this condition.
1. Angulo P. GI epidemiology: nonalcoholic fatty liver disease. Aliment Pharmacol Ther. 2007; 25: 883-9.
2. Adams LA, Angulo P. Recent concepts in nonalcoholic fatty liver disease. Diabet Med. 2005; 22: 1129-33.
3. Fan JG, Zhu J, Li XJ, Chen L, Li L, Dai F, et al. Prevalence of and risk factors for fatty liver in a general population of Shanghai, China. J Hepatol. 2005; 43: 508-14.
4. Sung KC, Ryan MC, Wilson AM. The severity of nonalcoholic fatty liver disease is associated with increased cardiovascular risk in a large cohort of nonobese Asian subjects. Atherosclerosis. 2009; 203: 581-6.
5. Targher G, Bertolini L, Poli F, Rodella S, Scala L, Tessari R, et al. Nonalcoholic fatty liver disease and risk of future cardiovascular events among type 2 diabetic patients. Diabetes. 2005; 54: 3541-6.
6. Ekstedt M, Franzen LE, Mathiesen UL, Thorelius L, Holmqvist M, Bodemar G, et al. Long-term follow-up of patients with NAFLD and elevated liver enzymes. Hepatology. 2006; 44: 865-73.
7. Targher G, Bertolini L, Scala L, Zenari L, Lippi G, Franchini M, et al. Plasma PAI-1 levels are increased in patients with nonalcoholic steatohepatitis. Diabetes Care. 2007; 30: e31-2.
8. Picardi A, Vespasiani-Gentilucci U. Association between non-alcoholic fatty liver disease and cardiovascular disease: a first message should pass. Am J Gastroenterol. 2008; 103: 3036-8.
9. Targher G, Arcaro G. Non-alcoholic fatty liver disease and increased risk of cardiovascular disease. Atherosclerosis. 2007; 191: 235-40.
10. Okui H, Hamasaki S, Ishida S, Kataoka T, Orihara K, Fukudome T, et al. Adiponectin is a better predictor of endothelial function of the coronary artery than HOMA-R, body mass index, immunoreactive insulin, or triglycerides. Int J Cardiol. 2007; 126: 53-61.
11. Jang Y, Lincoff AM, Plow EF, Topol EJ. Cell adhesion molecules in coronary artery disease. J Am Coll Cardiol. 1994; 24: 1591-601.
12. Schieffer B, Schieffer E, Hilfiker-Kleiner D, Hilfiker A, Kovanen PT, Kaartinen M, et al. Expression of angiotensin II and interleukin 6 in human coronary atherosclerotic plaques: potential implications for inflammation and plaque instability. Circulation. 2000; 101: 1372-8.
13. Mazzone GL, Rigato I, Ostrow JD, Bossi F, Bortoluzzi A, Sukowati CH, et al. Bilirubin inhibits the TNFalpharelated induction of three endothelial adhesion molecules. Biochem Biophys Res Commun. 2009; 386: 338-44.
14. Jin X, Yang YD, Chen K, Lv ZY, Zheng L, Liu YP, et al. HDMCP uncouples yeast mitochondrial respiration and alleviates steatosis in L02 and hepG2 cells by decreasing ATP and H2O2 levels: a novel mechanism for NAFLD. J Hepatol. 2009; 50: 1019-28.
15. Kralisch S, Sommer G, Stangl V, Köhler U, Kratzsch J, Stepan H, et al. Secretory products from human adipocytes impair endothelial function via nuclear factor kappaB. Atherosclerosis. 2008; 196: 523-31.
16. Havel PJ. Role of adipose tissue in body-weight regulation: mechanisms regulating leptin production and energy balance. Proc Nutr Soc. 2000; 59: 359-71.
17. Yamawaki H, Hara N, Okada M, Hara Y.Visfatin causes endothelium-dependent relaxation in isolated blood vessels. Biochem Biophys Res Commun. 2009; 383: 503-8.
18. Woo SH, Park MJ, An S, Lee HC, Jin HO, Lee SJ, et al. Diarsenic and tetraarsenic oxide inhibit cell cycle progression and bFGF- and VEGF-induced proliferation of human endothelial cells. J Cell Biochem. 2005; 95: 120-30.
19. Hwang SJ, Ballantyne CM, Sharrett AR, Smith LC, Davis CE, Jr Gotto AM, et al. Circulating adhesion molecules VCAM-1, ICAM-1 and E-selectin in carotid atherosclerosis and incident coronary heart disease patients: the Atherosclerosis Risk in Communities Study (ARIC). Circulation. 1997; 96: 4219-25.
20. Davies MJ, Gordon JL, Gearing AJ, Pigott R, Woolf N, Katz D, et al. The expression of the adhesion molecules ICAM-1, VCAM-1, PECAM, and E-selectin in human atherosclerosis. J Pathol. 1993; 171: 223-9.
21. O’Brien KD, McDonald TO, Chait A, Allen MD, Alpers CE. Neovascular experssion of E-selectin, intercellular adhesion molecule-1 in human atheroscleorsis and their relation to intimal leukocyte content. Circulation. 1996; 93: 672-82.
22. Blankenberg S, Rupprecht HJ, Bickel C, Peetz D, Hafner G, Tiret L, et al. Circulating cell adhesion molecules and death in patients with coronary artery disease. Circulation. 2001; 104: 1336-42.
23. Ivan L, Antohe F. Hyperlipidemia induces endothelialderived foam cells in culture. J Recept Signal Transduct Res. 2010; 30: 106-14.
24. Ross R. The pathogenesis of atherosclerosis: a perspective for the 1990s. Nature. 1993; 362: 801-9.
25. Fan JG. Impact of non-alcoholic fatty liver disease on accelerated metabolic complications. J Dig Dis. 2008; 9: 63-7.
26. Jarrar MH, Baranova A, Collantes R, Ranard B, Stepanova M, Bennett C, et al. Adipokines and cytokines in non-alcoholic fatty liver disease. Aliment Pharmacol Ther. 2008; 27: 412-21.
27. Baudin B, Bruneel A, Bosselut N, Vaubourdolle M. A protocol for isolation and culture of human umbilical vein endothelial cells. Nat Protoc. 2007; 2: 481-5.