Many receptor tyrosine kinases are expressed in activated HS
Many receptor tyrosine kinases are expressed in activated HSCs and are associated with the development of liver fibrosis. These include platelet-derived growth factor receptor (PDGFR) , vascular endothelial growth factor receptor (VEGFR) , and epidermal growth factor receptor (EGFR) . Only recently, EGFR has been studied in models of liver injury including the diethylnitrosamine (DEN) and the carbon tetrachloride (CCl4) model . EGFR is composed of a ligand-binding extracellular domain and a tyrosine kinase intracellular domain , . EGFR ligands include epidermal growth factor (EGF), transforming growth factor-α (TGF-α), amphiregulin (AR), epiregulin (EREG), β-cellulin (BTC), heparin-binding EGF (HB-EGF), and epigen (EPGN) . Upon binding to a ligand, EGFR forms homo- or heterodimers with other EGFR family members. Following activation of the intrinsic kinase domain, several proteins containing Src-homology 2 domains bind and activate downstream signaling cascades , , . The main activated downstream signaling pathways are extracellular signal-regulated kinase (ERK) and the phosphatidylinositol-3-kinase (PI3K)-Akt pathway , , .
EGFR has been well demonstrated as a therapeutic target for cancer treatment and several EGFR inhibitors have been used in clinical as anti-cancer agents. Recently, erlotinib, a small-molecule EGFR inhibitor, has also been shown to reduce the number of activated HSCs by depressing EGFR Necrosulfonamide , and is also being evaluated in an ongoing clinical trial (NCT02273362) for fibrogenesis inhibition and HCC prevention. Although EGFR inhibition has been reported to reduce HSCs, the underlying mechanisms are not known. In addition, the role of EGFR signaling has not been elucidated in a more clinically applicable diet-induced model of NAFLD. In this study, we utilized the high fat diet-induced model of liver injury and show that inhibition of EGFR prevents fat accumulation and liver fibrosis. We found the mechanism to involve modulation of oxidative stress. Furthermore, we show that EGFR regulates HSC activation and deposition of matrix proteins through inducing oxidative stress.
Materials and methods
Discussion HSC activation is well established to play a role in the development and progression of liver fibrosis. These peculiar cells are defined as fat-storing pericytes that constitute 5–8% of all liver cells . HSCs are quiescent but are activated through inflammatory stimuli and through paracrine interactions with Kupffer cells to become highly proliferative. Associated with this proliferative phase is the ability to synthesize and deposit matrix proteins. In experimental models of liver injury, this activated phase is denoted by induction of HSCs α-SMA , . Our studies show that HFD-feeding induces α-SMA in the liver tissues. We further show co-induction of collagen and fibrogenic TGF-β. Both of these readouts of HSCs are prevented when mice are treated with EGFR inhibitors. In addition, ox-LDL exposure of HSCs in culture improves growth and mimics activation as illustrated through α-SMA induction and matrix deposition. The precise mechanism by which EGFR activation facilitates HSC transdifferentiation is not known. However, we are able to speculate that the mechanism may involve ROS production. ROS may be derived from HSCs themselves or from the environment. Our studies show that ROS is induced when HSCs are exposed to ox-LDL suggesting an autocrine source. ROS may also be produced by Kupffer cells  and damaged hepatocytes . In both cases, however, it is expected that a pro-inflammatory and imbalanced redox environment is created. Agents which display anti-oxidative effects protect the liver from injury induced by hepatotoxins and prevent fibrogenesis , . In the present study, we found that oxidative stress is remarkably increased in the livers of HFD-fed mice in association with liver fibrosis and injury. In addition, we show that ROS scavenging by NAC prevent HSC activation and fibrosis.