15-Hydroxyprostaglandin dehydrogenase is an in vivo suppressor of colon tumorigenesis

Abstract

15-Hydroxyprostaglandin dehydrogenase (15-PGDH) is a prostaglandin-degrading enzyme that is highly expressed in normal colon mucosa but is ubiquitously lost in human colon cancers. Herein, we demonstrate that 15-PGDH is active in vivo as a highly potent suppressor of colon neoplasia development and acts in the colon as a required physiologic antagonist of the prostaglandin-synthesizing activity of the cyclooxygenase 2 (COX-2) oncogene. We first show that 15-PGDH gene knockout induces a marked 7.6-fold increase in colon tumors arising in the Min (multiple intestinal neoplasia) mouse model. Furthermore, 15-PGDH gene knockout abrogates the normal resistance of C57BL/6J mice to colon tumor induction by the carcinogen azoxymethane (AOM), conferring susceptibility to AOM-induced adenomas and carcinomas in situ. Susceptibility to AOM-induced tumorigenesis is mediated by a marked induction of dysplasia, proliferation, and cyclin D1 expression throughout microscopic aberrant crypt foci arising in 15-PGDH null colons and is concomitant with a doubling of prostaglandin E(2) in 15-PGDH null colonic mucosa. A parallel role for 15-PGDH loss in promoting the earliest steps of colon neoplasia in humans is supported by our finding of a universal loss of 15-PGDH expression in microscopic colon adenomas recovered from patients with familial adenomatous polyposis, including adenomas as small as a single crypt. These models thus delineate the in vivo significance of 15-PGDH-mediated negative regulation of the COX-2 pathway and moreover reveal the particular importance of 15-PGDH in opposing the neoplastic progression of colonic aberrant crypt foci.

Fig. 1.

Colon tumor induction by AOM. (a) Tumor development in 15-PGDH +/+ (n = 21), +/− (n = 40), and −/− (n = 24) C57BL/6J mice. Diamonds indicate mice without tumors. Boxes designate mice with colon tumors, with box size proportional to tumor number. Yellow fill designates tubular adenomas, and red fill designates tumors with high-grade dysplasia (also termed “carcinoma in situ”). ∗, P < 0.0001 for increased total colon tumors, P < 0.0008 for increased carcinoma in situ tumors, −/− vs. +/+ mice. Western blot assay of colonic 15-PGDH and actin are shown below each of the mice genotypes. (b) Gross morphology of a tumor (arrow) in 15-PGDH−/− mouse colon (Lower) compared with colon of a 15-PGDH+/+ mouse (Upper). (c and d) Representative histopathology of AOM-induced tumors. (c) Adenomatous polyp. (Scale bar, 100 μm.) (d) High-grade dysplasia (equivalently termed carcinoma in situ). (Scale bar, 200 μm for low-power field, with Inset magnified to same scale as c.)

Fig. 2.

Tumor induction in the APC^+/Min mouse. (a and b) Shown are the number of tumors per mouse in a combined cohort of APC^+/MinPGDH^+/+ and APC^+/MinPGDH^+/− mice (total n = 21) in which 15-PGDH is present (Present) vs. APC^+/MinPGDH^−/− mice (n = 13) in which 15-PGDH is absent (Absent). (a) Colon tumors. ∗, P < 0.0001 for increased colon tumors in 15-PGDH-absent mice. (b) Small intestinal tumors. ∗, P < 0.0001 for increased small intestinal tumors in 15-PGDH-absent mice. (c) Gross morphology of tumors (arrows) in a representative APC^+/MinPGDH^−/− mouse colon (Lower) compared with colon of an APC^+/MinPGDH^+/+ mouse (Upper). (d) Representative histopathology of a colon tumor from an APC^+/MinPGDH^−/− mouse. (Scale bar, 200 μm, with Inset as in Fig. 1d.)

Fig. 3.

ACF induction by AOM. (a and b) Diamonds indicate numbers of ACF observed for each colon examined from 15-PGDH +/+ (n = 10), +/− (n = 8), and −/− (n = 10) mice. Horizontal bars designate group means. Error bars denote SEMs. (a) Total ACF. ∗, P < 0.007 for increase in ACF in 15-PGDH −/− mice. (b) Large ACF (four or more crypts). ∗, P < 0.0001 for increase in large ACF in 15-PGDH−/− mice. (c) Methylene blue-stained ACF (bracketed) encompassing four crypts. (Scale bar, 100 μm.) (d) Percentage of ACF from 15-PGDH +/+ mice (open bars) vs. 15-PGDH −/− mice (filled bars) exhibiting moderate to severe dysplasia (+/+, n = 23 ACF from eight mice; −/−, n = 16 ACF from nine mice), nuclear cyclin D1 (+/+, n = 22 ACF from eight mice; −/−, n = 14 ACF from eight mice), and Ki-67 staining of the upper half of the crypt (+/+, n = 21 ACF from eight mice; −/−, n = 12 ACF from six mice). ∗, P < 0.003 for increased dysplasia, P < 0.0008 for increased cyclin D1, and P < 0.0001 for increased Ki-67. (e–g) A representative aberrant crypt focus (bracketed) from a 15-PGDH +/+ (Left) vs. a −/− (Right) mouse, with serial sections stained for histology (e), cyclin D1 (f), and Ki-67 (g). Arrows indicate regions of positive staining.

Fig. 4.

15-PGDH loss in FAP. (a) Graphical display of 15-PGDH immunostaining intensity (0 to 3+) in nine FAP patients providing 41 normal colonic mucosal samples (green bars) and 126 colon adenomas of sizes from >10 mm to fewer than eight crypts, with additional bar colors denoting adenoma lesions of different size classes. Bar heights denote the number of samples at each staining intensity level within each of the groups of different-sized adenomas, with sample numbers also tabulated beneath each grouping. (b) Photomicrograph demonstrating loss of 15-PGDH immunostaining in a single-crypt-sized adenoma vs. presence of 15-PGDH expression in surrounding normal epithelium. (Scale bar, 100 μm.)