APC-dependent suppression of colon carcinogenesis by PPARgamma

Abstract

Activation of PPARgamma by synthetic ligands, such as thiazolidinediones, stimulates adipogenesis and improves insulin sensitivity. Although thiazolidinediones represent a major therapy for type 2 diabetes, conflicting studies showing that these agents can increase or decrease colonic tumors in mice have raised concerns about the role of PPARgamma in colon cancer. To analyze critically the role of this receptor, we have used mice heterozygous for Ppargamma with both chemical and genetic models of this malignancy. Heterozygous loss of PPARgamma causes an increase in beta-catenin levels and a greater incidence of colon cancer when animals are treated with azoxymethane. However, mice with preexisting damage to Apc, a regulator of beta-catenin, develop tumors in a manner insensitive to the status of PPARgamma. These data show that PPARgamma can suppress beta-catenin levels and colon carcinogenesis but only before damage to the APC/beta-catenin pathway. This finding suggests a potentially important use for PPARgamma ligands as chemopreventative agents in colon cancer.

Figure 1

Histopathology and PPARγ expression in the colons of untreated Pparγ^+/+ and Pparγ^+/− mice. (a) Hematoxylin/eosin (H&E) staining of colons from Pparγ^+/+ Pparγ^+/− mice. Representative H&E staining from one mouse of each genotype. (b) Expression of PPARγ from protein lysates isolated from the colonic epithelium of Pparγ^+/+ and Pparγ^+/− mice. Protein lysates were obtained from the colonic epithelium of three mice per genotype. A representative immunoblot is shown. (c) Immunohistochemistry for PPARγ in the colons of (i) Pparγ^+/+ and (ii) Pparγ^+/− mice. Two different sets of immunohistochemistry for PPARγ: one Pparγ^+/+ and one Pparγ^+/− per set embedded in the same frozen section are shown.

Figure 2

Survival of Pparγ^+/+ and Pparγ^+/− mice after treatment with azoxymethane. (a) Survival of mice treated with 3.5 mg/kg azoxymethane. Mice not displaying rectal bleeding or morbidity were euthanized at 50 weeks after the last azoxymethane injections. P = 0.03, log-rank test. (b) Survival of mice treated with 7.0 mg/kg azoxymethane. P = 0.001, log-rank test. ⋄, Pparγ^+/+ mice; ●, Pparγ^+/− mice.

Figure 3

Pathology of tumors arising in mice after azoxymethane treatment. H&E staining of tumors arising in the colons of Pparγ^+/+ (Left) and Pparγ^+/− (Right) mice. (Magnification = ×10.) (Inset) Representative H&E sections. (Magnification = ×50.)

Figure 4

Expression of β-catenin in the colonic epithelium of untreated mice. (a) Expression of β-catenin from protein lysate isolated from the colonic epithelium of Pparγ^+/+ and Pparγ^+/− mice before carcinogen treatment. Protein lysates were obtained from the colonic epithelium of three mice per genotype. A representative immunoblot is shown. Actin levels are shown as a loading control. (b) Immunohistochemistry for β-catenin in the colons of (i) Pparγ^+/+ and (ii) Pparγ^+/− mice before carcinogen treatment. Two different sets of immunohistochemistry for β-catenin, consisting of one Pparγ^+/+ and one Pparγ^+/− per set embedded in the same frozen section are shown.

Figure 5

Expression of β-catenin from tumors arising in azoxymethane-treated mice. (a) Expression of β-catenin in protein lysates from tumors arising in Pparγ^+/+ and Pparγ^+/− mice after azoxymethane treatment. Representative immunoblots of two different tumors from different mice of each genotype are shown. Actin levels are shown as a loading control. (b) Expression of β-catenin in tumors arising in Pparγ^+/+ and Pparγ^+/− mice by immunohistochemistry.

Figure 6

β-catenin levels in the colonic epithelium and survival of Apc^+/1638N:Pparγ^+/+ Apc^+/1638N:Pparγ^+/+ mice. (a) Expression of β-catenin from protein lysate isolated from the colonic epithelium of Pparγ^+/+ and Pparγ^+/− mice on an Apc^+/1638N background. Protein lysates were obtained from the colonic epithelium of three mice per genotype. A representative immunoblot is shown. Actin levels are shown as a loading control. (b) Survival of Pparγ^+/+ and Pparγ^+/− mice on an Apc^+/1638N background. ⋄, Apc^+/1638N:Pparγ^+/+ mice; ●, Apc^+/1638N:Pparγ^+/− mice. P = 0.25, log-rank test.