Osteopontin is an important mediator of alcoholic liver disease via hepatic stellate cell activation

Figure 1 Over-expression of Osteopontin expression increased in human alcoholic liver disease, in vivo alcoholic steatosis mouse model and in vitro in stellate LX2 cells with a single acute dose of alcohol. A: Over-expression of Osteopontin (OPN)-C isoform mRNA (per ìg total RNA) increased progressively in livers from patients with alcoholic steatosis (AS), alcoholic hepatitis (AH) and alcoholic cirrhosis (AC) compared to non-diseased (ND) donor livers; B: Circulating OPN protein significantly increased (> 4-fold) in serum from patients with alcoholic cirrhosis compared to non-diseased alcoholics; C: In vivo: Hepatic OPN mRNA expression significantly increased from saline control with increasing doses of 2, 4 and 6 g/kg alcohol in wild type mice. No change was observed for â-actin expression used as control; D: Full length (about 53 kD) and cleaved (about 47 kD) OPN protein expression was visible with all alcohol doses (WB panel) and significantly increased with 6 g/kg alcohol on quantitation, normalized to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (bar graph); E: In vitro: Isoforms OPN-A (15-fold) and OPN-C (about 286-fold) mRNAs (transcripts per ìg total RNA) significantly increased with a single acute dose of alcohol (10 mmol/L per 4 h) compared to untreated control cells; F: Total secreted OPN protein (enzyme-linked immunosorbent assay) in culture media from alcohol treated LX2 cells also increased with respect to control cells but did not reach significance; G: Intracellular OPN (red) was associated with alcohol-induced LX2 activation (green) observed as recessed processes in alcohol treated LX2 compared to control cells (immunofluorescence). Overlay shows increased expression of OPN in LX2 cells exposed to alcohol. ^aP < 0.05, ^bP < 0.01 vs ND; ^dP < 0.01 vs saline control; ^eP < 0.01 vs control.