Expression of Leptin and Leptin Receptor During the Development of Liver Fibrosis and Cirrhosis

Abstract

Leptin is involved in the regulation of food intake and is mainly secreted by adipocytes. Major secretagogues are cytokines such as TNF-α or IL-1. Leptin in turn upregulates inflammatory immune responses. Elevated leptin serum levels have been detected in patients with liver cirrhosis, a disease frequently associated with elevated levels of circulating cytokines as well as hypermetabolism and altered body weight. Recently, leptin has been detected in activated hepatic stellate cells in vitro and an involvement of leptin in liver fibrogenisis has been suggested. The current study was designed to further clarify the role of leptin in liver disease by characterizing leptin and leptin receptor expression in the development and onset of experimental liver fibrosis.

Liver fibrosis and cirrhosis was induced in rats by use of phenobarbitone and increasing doses of CCl₄. Leptin and leptin receptor mRNA expression was determined by semiquantitative RT-PCR, protein expression by Western blot analysis and localization of leptin and its receptor by immunohistochemistry.

Normal liver tissue does not express leptin, but leptin receptor mRNA. Increasing levels of leptin mRNA were detected in fibrotic and cirrhotic livers correlated to the degree of fibrosis. Leptin receptor mRNA expression was not significantly altered in damaged livers. Increasing levels of leptin were detected in fibrotic and cirrhotic livers, whereas protein expression of the receptor remained unchanged. Throughout different stages of liver fibrosis, leptin immunreactivity was localized in activated hepatic stellate cells only, whereas immunoreactivity for the receptor was mainly seen on hepatocytes. In conclusion, leptin is expressed at increasing levels in activated hepatic stellate cells in vivo, which may therefore be a source of increased leptin tissue and serum levels contributing to the pathophysiology and morphological changes of chronic liver disease.

Key words

Leptin - liver fibrosis - stellate cells - inflammation

References

• 1 Alcolado R, Arthur M JP, Iredale J P. Pathogenesis of liver fibrosis.  Clin Sci. 1997;  92 103-112
• 2 Bado A, Levasseur S, Attoub S, Kermorgant S, Laigneau J P, Bortoluzzi M N, Moizo L. et al . The stomach is a source of leptin.  Nature. 1998;  394 790-793
• 3 Batts K P, Ludwig J. Chronic hepatitis. An update on terminology and reporting.  Am J Surg Pathol. 1995;  19 1409-1417
• 4 Bouloumie A, Marumo T, Lafontan M, Busse R. Leptin induces oxidative stress in human endothelial cells.  FASEB J. 1999;  13 1231-1238
• 5 Caro J F, Sinha M K, Kolaczynski J W, Zhang P L, Considine R V. Leptin: the tale of an obesity gene.  Diabetes. 1996;  45 1455-1462
• 6 Cohen B, Novick D, Rubinstein M. Modulation of insulin activities by leptin.  Science. 1996;  274 1185-1188
• 7 Cohen P, Zhao C, Cai X, Montez J M, Rohani S C, Feinstein P, Mombaerts P, Friedman J M. Selective deletion of leptin receptor in neurons leads to obesity.  J Clin Invest. 2001;  108 1113-1121
• 8 Faggioni R, Fantuzzi G, Gabay C, Moser A, Dinarello C A, Feingold K R, Grunfeld C. Leptin deficiency enhances sensitivity to endotoxin-induced lethality.  Am J Physiol. 1999;  276 R136-142
• 9 Faggioni R, Jones-Carson J, Reed D A, Dinarello C A, Feingold K R, Grunfeld C, Fantuzzi G. Leptin-deficient (ob/ob) mice are protected from T cell-mediated hepatotoxicity: role of tumor necrosis factor alpha and IL-18.  Proc Natl Acad Sci USA. 2000;  97 2367-2372
• 10 Frederich R C, Lollmann B, Hamann A, Napolitano-Rosen A, Kahn B B, Lowen B B, Flier J S. Expression of ob mRNA and its encoded protein in rodents. Impact of nutrition and obesity.  J Clin Invest. 1995;  96 1658-1663
• 11 Friedmann S L. The cellular basis of hepatic fibrosis: mechanisms and treatment strategies.  N Engl J Med. 1993;  328 1828-1835
• 12 Glasow A, Bornstein S R, Chrousos G P, Brown J W, Scherbaum W A. Detection of Ob-receptor in human adrenal neoplasms and effect of leptin on adrenal cell proliferation.  Horm Metab Res. 1999;  31 247-251
• 13 Grunfeld C, Zhao C, Fuller L, Pollock A, Moser A, Friedman J W, Feingold K R. Endotoxin and cytokines induce expression of leptin, the OB product, in hamsters. A role for leptin in the anorexia of infection.  J Clin Invest. 1996;  97 2152-2157
• 14 Henriksen J H, Holst J J, Moller S, Brinch K, Bendtsen F. Increased circulating leptin in alcoholic cirrhosis: relation to release and disposal.  Hepatology. 1999;  29 1818-1824
• 15 Ikejima K, Honda H, Yoshikawa M, Hirose M, Kitamura T, Takei Y, Sato N. Leptin augments inflammatory and profibrogenic responses in the murine liver induced by hepatotoxic chemicals.  Hepatology. 2001;  34 288-297
• 16 Ikejima K, Takei Y, Honda H, Hirose M, Yoshikawa M, Zhang Y J, Lang T, Fukuda T, Yamashina S, Kitamura T, Sato N. Leptin receptor-mediated signaling regulates hepatic fibrogenesis and remodeling of extracellular matrix in the rat.  Gastroenterology. 2002;  122 1399-1410
• 17 Kaplan L M. Leptin, obesity, and liver disease.  Gastroenterology. 1998;  115 997-1001
• 18 Kennedy A, Gettys T W, Watson P, Wallace P, Ganaway E, Pan Q, Garvey W T. The metabolic significance of leptin in humans: gender-based differences in relationship of adiposity, insulin sensivity, and energy expenditure.  J Clin Endocrinol Metab. 1997;  82 1293-1300
• 19 Leclercq I A, Farrell G C, Schriemer R, Robertson G R. Leptin is essential for the hepatic fibrogenic response to chronic liver injury.  J Hepatol. 2002;  37 206-213
• 20 Lee G H, Proenca R, Montez J M, Carroll K M, Darishzadeh J G, Lee J I, Friedmann J M. Abnormal splicing of the leptin receptor in diabetic mice.  Nature. 1996;  379 632-635
• 21 Loffreda S, Yang S Q, Lin H Z, Karp C L, Brengman M L, Wang D J, Klein A S, Bulkley G B, Bao C, Noble P W, Lane M D, Diehl A M. Leptin regulates proinflammatory responses.  FASEB J. 1998;  12 57-65
• 22 Lopez-Soriano J, Carbo N, Tessitore L, Lopez-Soriano F J, Argiles J M. Leptin and tumor growth in rats.  Int J Cancer. 1999;  81 726-729
• 23 Lord G M, Matarese G, Howard J K, Baker R J, Bloom S R, Lechler R I. Leptin modulates the T-cell immune response and reverses starvation-induced immunosuppression.  Nature. 1998;  394 897-901
• 24 Madej T, Boguski M S, Bryant S H. Threading analysis suggests that the obese gene product may be a helical cytokine.  FEBS Lett. 1995;  373 13-18
• 25 Masuzaki H, Ogawa Y, Hosoda K, Miyawaki T, Hanaoka I, Hiroka J, Yasuno Nishimura A H, Yoshimasa Y, Nishi S, Nakao K. Glucocorticoid regulation of leptin synthesis and secretion in humans: elevated plasma leptin levels in Cushing's syndrome.  J Clin Endocrinol Metab. 1997;  82 2542-2547
• 26 McCullough A J, Mullen K D, Kalkan S C. Measurement of total body and extracellular water in patients with and without ascites.  Hepatology. 1991;  14 1102-1111
• 27 McCullough A J, Bugianesi E, Marchesini G, Kalhan S C. Gender-dependent alterations in serum leptin in alcoholic cirrhosis.  Gastroenterology. 1998;  115 947-953
• 28 Mercer J G, Moar K M, Rayner D V, Trayhurn P, Hoggard N. Regulation of leptin receptor and NPY gene expression in hypothalamus of leptin-treated obese (ob/ob) and cold-exposed lean mice.  FEBS Lett. 1997;  402 185-188
• 29 Meyer-Alber A, Hartmann H, Stümpel F, Creutzfeldt W. Mechanism of insulin resistance in CCl₄-induced cirrhosis of rats.  Gastroenterology. 1992;  102 223-229
• 30 Mikhail A A, Beck E X, Shafer A, Barut B, Gbur J S, Zupancic T J, Schweitzer A C, Cioffi J A, Lacaud G, Ouyang B, Keller G, Snodgrass H R. Leptin stimulates fetal and adult erythroid and myeloid development.  Blood. 1997;  89 1507-1512
• 31 Muller M J, Lautz H U, Plogmann B, Berger M, Korber J, Schmidt F W. Energy expenditure and substrate oxidation in patients with cirrhosis: the impact of cause, clinical stage and nutritional state.  Hepatology. 1992;  15 782-797
• 32 Nemecz M, Preininger K, Englisch R, Furnsinn C, Schneider B, Waldhausl W, Roden M. Acute effect of leptin on hepatic glycogenolysis and gluconeogenesis in perfused rat liver.  Hepatology. 1999;  29 166-172
• 33 Potter J J, Womack L, Mezey E, Anania F A. Transdifferentiation of rat hepatic stellate cells results in leptin expression.  Biochem Biophys Res Com. 1998;  244 178-182
• 34 Santos-Alvarez J, Goberna R, Sanchez-Margalet V. Human leptin stimulates proliferation and activation of human circulating monocytes.  Cell Immunol. 1999;  194 6-11
• 35 Saxena N K, Ikeda K, Rockey D C, Friedman S L, Anania F A. Leptin in hepatic fibrosis: evidence for increased collagen production in stellate cells and lean littermates of ob/ob mice.  Hepatology. 2002;  35 762-771
• 36 Takahashi N, Waelput W, Guisez Y. Leptin is an endogenous protective protein against the toxicity exerted by tumor necrosis factor.  J Exp Med. 1999;  189 207-212
• 37 Tang M, Potter J J, Mezey E. Leptin enhances the effect of transforming growth factor beta in increasing type I collagen formation.  Biochem Biophys Res Commun. 2002;  297 906-911
• 38 Tartaglia L A, Dembski M, Weng X, Deng N, Culpepper J, Devos R, Richards G J. et al . Identification and expression cloning of a leptin receptor, OB-R.  Cell. 1995;  83 1263-1271
• 39 Tartaglia L A. The leptin receptor.  J Biol Chem. 1997;  272 6093-6096
• 40 Tsuchiya T, Shimizu H, Horie T, Mori M. Expression of leptin receptor in lung: leptin as a growth factor.  Eur J Pharmacol. 1999;  365 273-279
• 41 Vaisse C, Halaas J L, Horvath C M, Darnell J E, Stoffel M, Friedmann J M. Leptin activation of stat 3 in the hypothalamus of wild-type and ob/ob mice but not db/db mice.  Nat Gene. 1996;  14 95-97
• 42 Wolf G, Hamann A, Han D C, Helmchen U, Thaiss F, Ziyadeh F N, Stahl R A. Leptin stimulates proliferation and TGF-beta expression in renal glomerular endothelial cells: potential role in glomerulosclerosis.  Kidney Int. 1999;  56 860-872
• 43 Zhang Y, Proenca R, Maffei M, Barone M, Leopold L, Friedman J M. Positional cloning of the mouse obese gene and its human homologue.  Nature. 1994;  372 425-432
• 44 Zhao A Z, Shinohara M M, Huang D, Shimizu M, Eldar-Finkelman H, Krebs E G, Beavo J A, Bornfeldt K E. Leptin induces insulin-like signaling that antagonizes cAMP elevation by glucagon in hepatocytes.  J Biol Chem. 2000;  275 11348-11354