RNA Interference Against Hepatic Epidermal Growth Factor Receptor Has Suppressive Effects on Liver Regeneration in Rats

Abstract

Liver regeneration after a two-thirds partial hepatectomy (PHx) is a complex process requiring interaction and cooperation of many growth factors and cytokines and cross talk between multiple pathways. Along with hepatocyte growth factor and its receptor MET (HGF-MET), the epidermal growth factor receptor (EGFR) signaling pathway is activated within 60 minutes after PHx. To investigate the role of EGFR in liver regeneration, we used two EGFR-specific short hairpin silencing RNAs to inhibit EGFR expression in regenerating normal rat liver. Suppression of EGFR mRNA and protein was evident in treated rats. There was also a demonstrable decrease but not complete elimination of bromo-deoxyuridine incorporation and mitoses at 24 hours after PHx. In addition, we observed up-regulation of MET and Src as well as activation of the ErbB-3-ErbB-2-PI3K-Akt pathway and down-regulation of STAT 3, cyclin D1, cyclin E1, p21, and C/EBP ␤. The decrease in the ratio of C/EBP ␣ to C/EBP ␤ known to occur after PHx was offset in shEGFR-treated rats. Despite suppression of hepatocyte proliferation lasting into day 3 after PHx, liver weight restoration occurred. Interestingly, hepatocytes in shEGFR-treated rats were considerably larger when compared with ScrRNA-treated controls. The data indicate that although the MET and EGFR pathways are similar, the contributions made by MET and EGFR are unique and are not compensated by each other or other cytokines. Partial hepatectomy (PHx), in which two thirds of the rat liver is surgically removed, has been extensively used to study the highly complex phenomenon of liver regeneration. Although hepatocytes in normal adult liver are quiescent and rarely divide, they do retain an astounding ability to reenter the cell cycle and regenerate on surgical insult or injury. PHx in rats/mice results in rapid induction of more than 100 genes that are not expressed in the normal resting liver.1 A rapid up-regulation of genes encoding transcriptional factors like AP1 breakdown of extracellular matrix by uPA and release of pre-existing stores of HGF is observed within 60 minutes of a PHx. 2The hepatocytes leave the quiescent G0 phase and enter the cell cycle. Methods to identify extrahepatic signals leading to liver regeneration have included mitogenic effects on hepatocyte cultures, stimulation of DNA synthesis in the liver of normal (unoperated) animals, and decrease in regeneration-related events in animals genetically or pharmacologically depleted of the agent under study. Of the various agents implicated in liver regeneration, HGF and ligands of the epidermal growth factor receptor (EGFR) are the only ones that stimulate DNA synthesis in hepatocyte cultures maintained in chemically defined media.3 They are also the only ones that stimulate DNA synthesis in the liver of normal mice and rats. 4 -6 HGF and EGF signaling pathways are activated within 60 minutes after a PHx, 7,8 as evidenced by tyrosine phosphorylation of MET and EGFR within 30 to 60 minutes after PHx. There is evidence of cross talk and c...

Branches of NF-κb signaling pathway regulate hepatocyte proliferation in rat liver regeneration

“…The activities of physiological processes reflected by the expression changes were analyzed then by systems biology methods, and both growth factor branches and its regulation of hepatocyte proliferation activity were elevated in rat liver regeneration (Figure 1). The above results were consistent with those of other authors (Gezginci-Oktayoglu et al, 2010;Paranjpe et al, 2010;Gui et al, 2011;Tsai and Wang, 2011;Wu et al, 2011), who demonstrated the correlation between the signal transduction activity of the growth factor branch of the NF-κB signaling pathway and hepatocyte proliferation activity. The above results helped deduce the following possible connection between the growth factor branch and hepatocyte proliferation activity.…”

A disintegrin and metalloproteinase 17 regulates TNF and TNFR1 levels in inflammation and liver regeneration in mice

“…We show that the induction of AR protein is abrogated in ADAM17 h-KO mice, implicating ADAM17 as a major regulator of AR in the liver. EGFR activation is key to hepatocyte cell cycle progression, and rodents in which liver EGFR is conditionally inactivated have impaired liver regeneration after PH (28,29). TGF-␣ and HB-EGF, which are other EGFR ligands and hepatocyte mitogens, are also cleaved by ADAM17 in cell culture (32).…”

Identification of an epigenetic signature of early mouse liver regeneration that is disrupted by Zn-HDAC inhibition

“…Many of these hyperacetylated genes are known to be induced during, to positively regulate, or to interact with factors that promote liver regeneration. For example, Ccnd1 (i.e., cyclin d1), Cdkn1a (i.e., p21), Gadd45g, Lcn2, and Myc are induced in early regenerating liver [4][5][6][7] ; Ccnd1, Egfr, and Prlr promote liver regeneration [8][9][10][11] ; and Tcf7l2 (also known as Tcf4) binds to b-catenin, whose signaling is activated during and promotes regeneration. 12 Deacetylated genes Similar examination of genes deacetylated in response to PH showed 37 of 392 associated with cell cycle-and 31 linked to cell death-related gene category terms ( Tables 1 and 2).…”