Adiponectin upregulates hepatocyte CMKLR1 which is reduced in human fatty liver
Highlights
► Chemokine-like receptor 1 (CMKLR1) is the receptor for the adipokine chemerin. ► CMKLR1 is expressed by hepatocytes. ► CMKLR1 is reduced in fatty liver and in rodent fibrotic liver. ► Adiponectin induces CMKLR1 in hepatocytes. ► CMKLR1 is reduced in liver of adiponectin-deficient mice.
Introduction
Non-alcoholic fatty liver disease is becoming the most common cause of chronic liver diseases in westernized countries (Clark, 2006). Obesity is frequently accompanied by NAFLD and altered metabolic function and increased production of inflammatory adipokines are suggested to contribute to hepatic steatosis and progressive liver disease (Clark, 2006, Schaffler et al., 2005). IL-6 is preferentially released from visceral fat and upregulates suppressor of cytokine signaling 3 in the liver which causes hepatic insulin resistance (Fontana et al., 2007, Sabio et al., 2008, Wiest et al., 2011). Leptin is mainly produced by subcutaneous adipocytes, and obesity is characterized by elevated systemic levels. Leptin prevents lipid accumulation in the liver, and animal studies proved that leptin directly promotes fibrogenesis (Biddinger et al., 2006, Wang et al., 2009a). Systemic adiponectin which lowers hepatocyte lipid storage and protects from progressive liver damage is reduced in obesity and patients with fatty liver disease independent of body mass index (Polyzos et al., 2010, Schaffler et al., 2005, Wang et al., 2009b). Antiinflammatory effects of adiponectin include inhibition of TNF activity, a cytokine involved in the pathophysiology of metabolic liver injury (Tilg, 2010).
Chemokine-like receptor 1 (CMKLR1) is expressed by immune cells and is downregulated by TNF and LPS in macrophages (Zabel et al., 2006). TGFβ which deactivates macrophages induces CMKLR1 (Zabel et al., 2006). Mice with deficiency of CMKLR1 exhibit increased recruitment of inflammatory cells in the lung upon LPS challenge indicating antiinflammatory effects of CMKLR1 (Luangsay et al., 2009).
Chemerin is one of the ligands of CMKLR1 and circulating levels are increased in obesity (Meder et al., 2003, Weigert et al., 2010). Serum chemerin positively correlates with BMI, Homeostasis Model Assessment of Insulin Resistance, triglycerides, systolic blood pressure, leptin, resistin, C-reactive protein, TNF and IL-6 and negatively with HDL cholesterol suggesting a function of chemerin in metabolic disturbances associated with obesity (Bozaoglu et al., 2007, Ernst et al., 2010, Parlee et al., 2010, Stejskal et al., 2008, Weigert et al., 2010). In morbidly obese patients with non-alcoholic steatohepatitis (NASH) chemerin is further elevated (Sell et al., 2010). Chemerin is an attractant for immune cells and pro- and antiinflammatory effects of chemerin have been described (Ernst and Sinal, 2010). More recently it has been shown that chemerin impairs insulin signaling in adipocytes and skeletal muscle cells (Becker et al., 2010, Kralisch et al., 2009, Sell et al., 2009).
CMKLR1 also serves as a receptor for resolvin E1 which is derived from omega 3 polyunsaturated fatty acids at inflammatory sites (Arita et al., 2007). Resolvin E1 mediates insulin-sensitizing and antisteatotic effects in rodent models of obesity (Arita et al., 2007, Gonzalez-Periz et al., 2009).
CMKLR1 is expressed in the liver (Ernst et al., 2010, Parlee et al., 2010) and so far it has been shown that mRNA levels are not altered in the liver of ob/ob mice compared to wild type animals. It was hypothesized that CMKLR1 is also expressed by hepatocytes and that its level may be altered in NAFLD. Therefore, we intended to more closely analyse the expression of CMKLR1 in human liver cell populations, human and rodent fatty liver and rodent NASH.
Section snippets
Material
Dulbecco’s modified eagle medium (DMEM) was from PAA (Karlsruhe, Germany), RNeasy Mini Kit was from Qiagen (Hilden, Germany) and oligonucleotides were synthesized by Metabion (Planegg-Martinsried, Germany). LightCycler FastStart DNA Master SYBR Green I was purchased from Roche (Mannheim, Germany). The ACAT inhibitor Sandoz 58-035, LPS (Escherichia coli serotype 055:B5), palmitic acid and oleic acid were ordered from Sigma (Deisenhofen, Germany). Acetylated LDL was from Invitrogen GmbH
CMKLR1 expression in primary human liver cells
CMKLR1 mRNA was detected in primary human hepatocytes (PHH), hepatic stellate cells (HSC), endothelial cells, Kupffer cells (KC) and bile duct cells (Fig. 1A). CMKLR1 protein was analyzed in PHH, KC and HSC and was expressed by all of these cells. Although identical concentrations of the different protein lysates were used GAPDH protein was higher abundant in HSC, KC and macrophages compared to PHH (Fig. 1B). To confirm similar amounts of protein in the different lysates Coomassie stained
Discussion
In the current study we show that CMKLR1 protein is expressed by hepatocytes, hepatic stellate cells, bile duct epithelial cells and Kupffer cells. All of these cells express CMKLR1 mRNA but it has to be considered that primary cells are contaminated with other liver cells to some degree and this may give false positive results. To solve this problem in situ hybridisation to detect cellular mRNAs has to be performed. CMKLR1 protein is highly expressed in hepatoma cell lines suggesting that
Disclosure
The authors declare that they have no conflicts of interest related to this work.
Acknowledgments
The technical assistance of Yvonne Hader is greatly appreciated. The study was supported by a Grant from the Deutsche Forschungsgemeinschaft (BU 1141/3-3, BU 1141/7-1) to C. Buechler and a grant from the Regensburger Forschungsförderung (ReForm C) to C. Buechler, C. Hellerbrand and T.S. Weiss.
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