(580.3) A Mechanistic Role for RECK in the Regulation of Hepatocellular Inflammation

Poster Board Number: E308

Ryan Dashek (University of Missouri, University of Missouri), Connor Diaz (University of Missouri), Bysani Chandrasekar (University of Missouri, University of Missouri, University of Missouri, University of Missouri), R Scott Rector (University of Missouri, University of Missouri, University of Missouri, University of Missouri)

RECK (Reversion Inducing Cysteine Rich Protein with Kazal Motifs), a membrane-anchored glycoprotein, modifies the extracellular matrix involved in various cancers, including hepatocellular carcinoma (HCC). Its role in regulating inflammatory and fibrogenic processes has also been postulated. Both inflammation and fibrosis contribute to progression of nonalcoholic steatohepatitis (NASH) to HCC. Here, we tested the hypothesis that inducing or sustaining RECK expression will inhibit proinflammatory amphiregulin and epidermal growth factor receptor (EGFR) signaling, progression of NASH and the development of HCC using an in vitro cell culture model employing isolated adult mouse primary hepatocytes. Ectopic RECK overexpression (gain-of-function) was achieved by the adenoviral (Adv) transduction of murine RECK cDNA. Adv.GFP served as a control. RECK expression was silenced (loss-of-function) by transducing RECK-specific siRNA using lipofectamine. Nonspecific siRNA served as a control. Hepatocytes were exposed to TNFα in the presence or absence of inhibitors against ADAM 10 (A Disintegrin and Metalloproteinase Domain-Containing Protein 10) and/or ADAM17 – two well-established pro-inflammatory sheddases.  Results show that RECK overexpression inhibited TNFα-induced ADAM10/17 activity, and amphiregulin (an EGFR ligand) and EGFR expression. In the presence of ADAM10/17 inhibitors, no further reduction was observed in amphiregulin and EGFR activity, suggesting a maximal inhibitory effect of RECK on EGFR activity. In contrast, silencing RECK significantly increased amphiregulin secretion, and this effect was reversed by inhibition of ADAM10/17, indicating RECK signals via regulation of these sheddases. Thus, RECK not only regulates sheddase activity of ADAM10 and 17, but also downstream amphiregulin and EGFR signaling under pro-inflammatory stimuli.  Because increased and sustained EGFR activity contributes to progression of NASH to HCC in preclinical models, our results indicate that inducing RECK has the potential to inhibit hepatocellular inflammation in the setting of NASH and HCC.

This work was supported by VA-Merit Grant I01BX003271-01 (R.S.R.) and the Truman VA Medical Research Foundation. RD was also funded by NIH T32 OD011126.