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. 2008 Jan 7;28(5):1489–1502. doi: 10.1128/MCB.01090-07

FIG. 5.

FIG. 5.

Molecular analysis of HCC in Iqgap2−/− mice. (A) Immunohistochemical detection of IQGAP1, β-catenin, and E-cadherin in livers from 4-month-old wild-type or Iqgap2−/− mice without HCC or 2-year-old Iqgap2−/− mice with HCC. Note the loss of E-cadherin from cell membranes, along with the enhanced cytoplasmic expression and translocation of both IQGAP1 and β-catenin, occurring only with HCC. Scale bars correspond to 50 μm. (B) Immunoblot analysis was completed using an anti-β-catenin monoclonal antibody and liver lysates (10 μg/lane) from mice (2 years old) with HCC or age-matched controls; the arrow indicates the 92-kDa β-catenin band, with clear evidence for truncated β-catenin mutant forms in the case of HCC. (C) Immunoblot (10 μg of lysate/lane) demonstrating the presence of active (dephosphorylated) β-catenin mutant forms in HCC liver tissue. Results for livers from two 4-month-old wild-type (WT) and Iqgap2−/− (−/−) mice are also shown. Cell lysate from the A431 cell line (Upstate Biotechnology/Millipore) was used as a positive control. (D) Immunoblot analysis (10 μg of lysate/lane) to assess E-cadherin, cyclin D1, and IQGAP1 expression in Iqgap2−/− liver lobes without HCC or affected by HCC; paired normal (N) and tumor (T) samples were collected from the same animals. (E) Immunoblot densitometric analysis of E-cadherin, cyclin D1, and IQGAP1 expression was conducted using liver lysates (10 μg/lane) from lobes affected by HCC (n = 4), those of 4- and 12-month-old Iqgap2−/− mice without HCC (n = 3), and those of 4-month-old wild-type mice (n = 2). Data are presented as mean ratios ± standard deviations of the expression of individual proteins relative to that of proteins in the wild-type samples. (F) IQGAP1 expression was assessed by immunoblotting of liver lysates (10 μg/lane) from wild-type (WT) and Iqgap2−/− (−/−) mice of various ages (6 weeks and 4, 12, and 24 months). Glyceraldehyde-3-phosphate dehydrogenase (GAPDH)-specific antibody served as a control for the loading of equal quantities. A representative immunoblot is shown. (G) Immunoblot densitometric analysis of hepatic IQGAP1 expression in wild-type and Iqgap2−/− mice of various ages. Data are presented as means ± SEM of the integrated optical densities of bands. n = 3 for each age group.