372 Gut and Liver, Vol. 5, No. 3, September 2011 Fig. 3. Analysis of cyclooxygenase (COX)-2 expression in the 2-acetylaminofluorene (2-AAF)/70% partial hepatectomy (PHx) model and in sorted hepatic oval cells (HOCs). (A) RT-PCR analysis of COX-2 in normal liver (NL), 2AAF/PHx liver (day 9), PHx liver (after 12 hours), and Thy1+-sorted HOCs. (B) Western blot of a whole-liver homogenate demonstrating the accumulation of c-Met and COX-2. β-actin is used as a loading control. (C) Immunofluorescence staining (Texas Red) to examine the expression of COX-2, albumin, alpha-fetoprotein (AFP), fibronectin (FN), and CD45 in Thy1+ HOCs sorted from the livers of 2AAF/PHx-treated rats on day 9 (original magnification, ×600) Fig. 4. Detection of prostaglandin (PG) receptors in hepatic oval cells (HOCs). RT-PCR amplification of the PG receptor family (EP1-4) in normal liver (NL), the liver regeneration model (PHx), and Thy1+- sorted HOCs. GAPDH is used as an internal standard. PG signaling during liver regeneration. The relationship between COX-2 and HOC proliferation-related signals involved two ECM proteins (CTGF and FN) and AKT activity. The COX-2 inhibitor NS398 decreased HOC proliferation and significantly suppressed CTGF and FN expression and AKT activation, but not ERK 1/2 activation. Moreover, the observation that NS398 did not affect HOC proliferation on FN-coated dishes indicates that the FN- induced proliferation of HOCs occurs through COX-2-induced PG signaling. The observation that HOCs treated with NS398 were shifted to an HGF-containing medium for HOC regulation Fig. 5. Western blot analysis of NS398-treated hepatic oval cells (HOCs). HOCs cultured with 100 μM NS398 for 24 hours are exam- ined. Control cells are treated with DMSO without NS398. NS398 suppress cyclooxygenase (COX)-2, connective tissue growth factor (CTGF), fibronectin (FN), and AKT expression, but not ERK expres- sion. β-Actin is used as a loading control. is entirely consistent with a role for HGF in HOC proliferation. In the liver regeneration model, HOCs participate in liver regeneration by activating a complex mixture of compounds, including cytokines and growth factors.2-6 Several growth fac- tors (e.g., HGF and epidermal growth factor) and intracellular signaling pathways (e.g., ERK and Akt) are important regulators of HOC-mediated liver regeneration.41-43 Furthermore, the upreg-