Abstract
Non-alcoholic fatty liver disease (NAFLD) is characterized by steatosis associated with liver inflammation. As NAFLD progresses, triglycerides increase within hepatocytes, causing typical vacuoles that resemble adipocytes. However, whether these morphological changes in hepatocytes indicate potential functional changes is unclear. C57BL/6J mice were fed a high-fat diet (HFD) containing 42 % fat. Markers for adipocytes in the liver were measured using real-time PCR, Western blot, and double immunofluorescent labeling. Cytokines in cell culture supernatants were quantified with ELISA. To determine the macrophage phenotype, hepatic classical M1 markers and alternative M2 markers were analyzed. After a 24-week feeding period, adipocyte markers aP2 and PPARγ increased at both the mRNA and protein level in the liver of HFD-fed mice. FITC-labeled aP2 and rhodamine-labeled albumin were both stained in the cytoplasm of steatotic hepatocytes as observed under confocal laser scanning microscopy. Cell membrane-bound E-cadherin and albumin expression were reduced in steatotic hepatocytes compared to controls. However, hepatic adiponectin and adiponectin receptor-2 expression decreased with upregulation of hepatic CD36, suggesting impaired adiponectin activity in livers of HFD-fed mice. Moreover, steatotic primary hepatocytes not only released pro-inflammatory cytokines such as TNFα, MCP-1, IL-6, and IL-18, but also could activate macrophages when co-cultured in vitro. In vivo, hepatic expression of M1 genes such as iNOS and TNFα was markedly increased in HFD-fed mice. In contrast, hepatic expression of M2 genes such as Arg1 and CD206 was significantly reduced. Specifically, the ratio of TNFα to CD206 in HFD-fed mice was notably upregulated. Overexpression of adipocyte-specific genes in hepatocytes and their secretory function and epithelial phenotype impairment in NAFLD cause functional changes in steatotic hepatocytes aside from morphological changes. This suggests that adipogenic changes in hepatocytes are involved in pathogenesis of NAFLD.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
D.G. Tiniakos, M.B. Vos, E.M. Brunt, Nonalcoholic fatty liver disease: pathology and pathogenesis. Annu. Rev. Pathol. 5, 145–171 (2010)
E. Fabbrini, S. Sullivan, S. Klein, Obesity and nonalcoholic fatty liver disease: biochemical, metabolic, and clinical implications. Hepatology 51, 679–689 (2010)
H. Tilg, A.R. Moschen, Evolution of inflammation in nonalcoholic fatty liver disease: the multiple parallel hits hypothesis. Hepatology 52, 1836–1846 (2010)
S. Yu, K. Matsusue, P. Kashireddy, W.Q. Cao, V. Yeldandi, A.V. Yeldandi et al., Adipocyte-specific gene expression and adipogenic steatosis in the mouse liver due to peroxisome proliferator-activated receptor gamma1 (PPARgamma1) overexpression. J. Biol. Chem. 278, 498–505 (2003)
A. Vidal-Puig, M. Jimenez-Linan, B.B. Lowell, A. Hamann, E. Hu, B. Spiegelman et al., Regulation of PPAR gamma gene expression by nutrition and obesity in rodents. J. Clin. Invest. 97, 2553–2561 (1996)
N. Ouchi, J.L. Parker, J.J. Lugus, K. Walsh, Adipokines in inflammation and metabolic disease. Nat. Rev. Immunol. 11, 85–97 (2011)
B.L. Kreamer, J.L. Staecker, N. Sawada, G.L. Sattler, M.T. Hsia, H.C. Pitot, Use of a low-speed, iso-density percoll centrifugation method to increase the viability of isolated rat hepatocyte preparations. In Vitro Cell Dev. Biol. 22, 201–211 (1986)
S.E. Schadinger, N.L. Bucher, B.M. Schreiber, S.R. Farmer, PPARgamma2 regulates lipogenesis and lipid accumulation in steatotic hepatocytes. Am. J. Physiol. Endocrinol. Metab. 288, 1195–1205 (2005)
D.E. Kleiner, E.M. Brunt, M. Van Natta, C. Behling, M.J. Contos, O.W. Cummings et al., Design and validation of a histological scoring system for nonalcoholic fatty liver disease. Hepatology 41, 1313–1321 (2005)
P. Puri, R.A. Baillie, M.M. Wiest, F. Mirshahi, J. Choudhury, O. Cheung et al., A lipidomic analysis of nonalcoholic fatty liver disease. Hepatology 46, 1081–1090 (2007)
M. Bell, H. Wang, H. Chen, J.C. McLenithan, D.W. Gong, R.Z. Yang et al., Consequences of lipid droplet coat protein downregulation in liver cells: abnormal lipid droplet metabolism and induction of insulin resistance. Diabetes 57, 2037–2045 (2008)
K. Yamaguchi, L. Yang, S. McCall, J. Huang, X.X. Yu, S.K. Pandey et al., Inhibiting triglyceride synthesis improves hepatic steatosis but exacerbates liver damage and fibrosis in obese mice with nonalcoholic steatohepatitis. Hepatology 45, 1366–1374 (2007)
C.Z. Larter, M.M. Yeh, J. Williams, K.S. Bell-Anderson, G.C. Farrell, MCD-induced steatohepatitis is associated with hepatic adiponectin resistance and adipogenic transformation of hepatocytes. J. Hepatol. 49, 407–416 (2008)
J.C. Cohen, J.D. Horton, H.H. Hobbs, Human fatty liver disease: old questions and new insights. Science 332, 1519–1523 (2011)
J.M. Lehmann, L.B. Moore, T.A. Smith-Oliver, W.O. Wilkison, T.M. Wilkison, S.A. Kliewer, An antidiabetic thiazolidinedione is a high affinity ligand for peroxisome proliferator-activated receptor gamma (PPAR gamma). J. Biol. Chem. 270, 12953–12956 (1995)
W. Tang, D. Zeve, J. Seo, A.Y. Jo, J.M. Graff, Thiazolidinediones regulate adipose lineage dynamics. Cell Metab. 14, 116–122 (2011)
A.G. Cristancho, M.A. Lazar, Forming functional fat: a growing understanding of adipocyte differentiation. Nat. Rev. Mol. Cell Biol. 12, 722–734 (2011)
E. Moran-Salvador, M. Lopez-Parra, V. Garcia-Alonso, E. Titos, M. Martinez-Clemente, A. Gonzalez-Periz et al., Role for PPARgamma in obesity-induced hepatic steatosis as determined by hepatocyte- and macrophage-specific conditional knockouts. FASEB J. 25, 2538–2550 (2011)
M. Hemmingsen, S. Vedel, P. Skafte-Pedersen, D. Sabourin, P. Collas, H. Bruus et al., The role of paracrine and autocrine signaling in the early phase of adipogenic differentiation of adipose-derived stem cells. PLoS ONE 8, 63638 (2013)
H. Yoshida, Y. Kanamori, H. Asano, O. Hashimoto, M. Murakami, T. Kawada et al., Regulation of brown adipogenesis by the Tgf-beta family: involvement of Srebp1c in Tgf-beta- and Activin-induced inhibition of adipogenesis. Biochim. Biophys. Acta 1830, 5027–5035 (2013)
J. Wanninger, M. Neumeier, C. Hellerbrand, D. Schacherer, S. Bauer, T.S. Weiss et al., Lipid accumulation impairs adiponectin-mediated induction of activin A by increasing TGFbeta in primary human hepatocytes. Biochim. Biophys. Acta 1811, 626–633 (2011)
P. Charatcharoenwitthaya, K.D. Lindor, P. Angulo, The spontaneous course of liver enzymes and its correlation in nonalcoholic fatty liver disease. Dig. Dis. Sci. 57, 1925–1931 (2012)
S. Kaser, A. Moschen, A. Cayon, A. Kaser, J. Crespo, F. Pons-Romero et al., Adiponectin and its receptors in non-alcoholic steatohepatitis. Gut 54, 117–121 (2005)
J. Tam, G. Godlewski, B.J. Earley, L. Zhou, T. Jourdan, G. Szanda et al., Role of adiponectin in the metabolic effects of cannabinoid type 1 receptor blockade in mice with diet-induced obesity. Am. J. Physiol. Endocrinol. Metab. 306, E457–E468 (2014)
C. Finelli, G. Tarantino, What is the role of adiponectin in obesity related non-alcoholic fatty liver disease? World J. Gastroenterol. 19, 802–812 (2013)
J.M. Pappachan, F.A. Antonio, M. Edavalath, A. Mukherjee, Non-alcoholic fatty liver disease: a diabetologist’s perspective. Endocrine 45, 344–353 (2014)
G. Sabio, M. Das, A. Mora, Z. Zhang, J.Y. Jun, H.J. Ko et al., A stress signaling pathway in adipose tissue regulates hepatic insulin resistance. Science 322, 1539–1543 (2008)
L.C. Davies, S.J. Jenkins, J.E. Allen, P.R. Taylor, Tissue-resident macrophages. Nat. Immunol. 14, 986–995 (2013)
N. Lanthier, O. Molendi-Coste, Y. Horsmans, N. van Rooijen, P.D. Cani, I.A. Leclercq, Kupffer cell activation is a causal factor for hepatic insulin resistance. Am. J. Physiol. Gastrointest. Liver Physiol. 298, G107–G116 (2010)
R. Stienstra, F. Saudale, C. Duval, S. Keshtkar, J.E. Groener, N. van Rooijen et al., Kupffer cells promote hepatic steatosis via interleukin-1beta-dependent suppression of peroxisome proliferator-activated receptor alpha activity. Hepatology 51, 511–522 (2010)
W. Huang, A. Metlakunta, N. Dedousis, P. Zhang, I. Sipula, J.J. Dube et al., Depletion of liver Kupffer cells prevents the development of diet-induced hepatic steatosis and insulin resistance. Diabetes 59, 347–357 (2010)
A.H. Clementi, A.M. Gaudy, N. van Rooijen, R.H. Pierce, R.A. Mooney, Loss of Kupffer cells in diet-induced obesity is associated with increased hepatic steatosis, STAT3 signaling, and further decreases in insulin signaling. Biochim. Biophys. Acta 1792, 1062–1072 (2009)
N. Lanthier, O. Molendi-Coste, P.D. Cani, N. van Rooijen, Y. Horsmans, I.A. Leclercq, Kupffer cell depletion prevents but has no therapeutic effect on metabolic and inflammatory changes induced by a high-fat diet. FASEB J. 25, 4301–4311 (2011)
Acknowledgments
This work was supported by the National Natural Science Foundation of China (Grant No. 81370525).
Conflict of interest
The authors declare that they have no conflict of interest.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Pan, X., Wang, P., Luo, J. et al. Adipogenic changes of hepatocytes in a high-fat diet-induced fatty liver mice model and non-alcoholic fatty liver disease patients. Endocrine 48, 834–847 (2015). https://doi.org/10.1007/s12020-014-0384-x
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s12020-014-0384-x