Monitoring of tumor promotion and progression in a mouse model of inflammation-induced colon cancer with magnetic resonance colonography

Abstract

Early detection of precancerous tissue has significantly improved survival of most cancers including colorectal cancer (CRC). Animal models designed to study the early stages of cancer are valuable for identifying molecular events and response indicators that correlate with the onset of disease. The goal of this work was to investigate magnetic resonance (MR) colonography in a mouse model of CRC on a clinical MR imager. Mice treated with azoxymethane and dextran sulfate sodium were imaged by serial MR colonography (MRC) from initiation to euthanasia. Magnetic resonance colonography was obtained with both T1- and T2-weighted images after administration of a Fluorinert enema to remove residual luminal signal and intravenous contrast to enhance the colon wall. Individual tumor volumes were calculated and validated ex vivo. The Fluorinert enema provided a clear differentiation of the lumen of the colon from the mucosal lining. Inflammation was detected 3 days after dextran sulfate sodium exposure and subsided during the next week. Tumors as small as 1.2 mm(3) were detected and as early as 29 days after initiation. Individual tumor growths were followed over time, and tumor volumes were measured by MR imaging correlated with volumes measured ex vivo. The use of a Fluorinert enema during MRC in mice is critical for differentiating mural processes from intraluminal debris. Magnetic resonance colonography with Fluorinert enema and intravenous contrast enhancement will be useful in the study of the initial stages of colon cancer and will reduce the number of animals needed for preclinical trials of prevention or intervention.

Figure 1

Timeline for AOM/DSS treatment. Day 0: mice were injected with a single dose of AOM. Days 7 to 12: mice were treated with of 0, 1%, or 2% DSS in drinking water. Days 28 to 33: some mice were treated with a second cycle of DSS. Mice were imaged by MRI multiple times throughout the study. MR indicates the imaging time points used to generate Figure 3.

Figure 2

Magnetic resonance imaging shows inflammation 3 days after DSS treatment. T2W and T1W image sequences applied in coronal plane without and with intravenous contrast. Colons were not distended with air or enema. Normal colon (A), inflamed colon 3 days after DSS exposure ended (B), colon recovering from inflammation 9 days after DSS exposure ended (C; T2W only). Red arrows show the anus (A), the rectum (R), sigmoid colon (SC), descending colon (DC), and the transverse colon (TC); white arrowheads show fat (F); white arrows show muscle (M), stool (ST), stomach (S), and kidney (K); and red arrowheads in panel B show luminal contrast agent. (D) Corresponding whole-mount H&E-stained section of colon tissue confirms inflammation 3 days after DSS: normal colon (top panels) and inflamed colon 3 days after DSS ended (bottom panels). Scale bar, 1 mm.

Figure 3

Fluorinert is used to distend and reduce signal from the lumen of the colon. Representative images from MRI sequences applied in coronal plane: (A) T2W, T1W, and T1W with contrast-enhanced coronal images in a healthy colon after Fluorinert enema; (B) T2W coronal images in a tumor-bearing colon: before enema (left panel) and with Fluorinert enema (right panel). (C) Sagittal (top panels) and axial (bottom panels) views after MPR of postcontrast T1 images: normal colon (left panels) and tumor-bearing colon (right panels). Subpanels a-a, b-b, c-c, and d-d indicate the same location in each view. Arrowhead shows healthy mucosal lining (a-a and b-b) and damaged mucosal lining (c-c and d-d).

Figure 4

Magnetic resonance colonography is useful for monitoring the progression of inflammation-induced colon carcinogenesis. (A) Mice were imaged with T2W images followed by T1W images before and after contrast. Representative coronal images from T1W MRI sequences after contrast are shown. Except for control, images are from the same mouse imaged on days 15, 21, 29, 54, and 63 after AOM injection. Untreated mouse was injected with AOM but did not receive DSS and was imaged on day 39. Day 15 shows inflammation (contrast bleeding into lumen). Day 20 shows that inflammation has resolved and contrast enhancement of mucosal lining (arrowhead). Day 29 shows beginning of tumor growth (white arrows). Day 50 shows that the same tumors have grown (white arrows) and new tumors have formed. Day 60 shows mouse just before euthanasia. (B) Axial views after multiplanar reformatting of postcontrast T1 images.

Figure 5

Comparison of MRC with histopathology. Optical photography of the colon ex vivo shows multiple tumors. (A–C) Axial MPR and corresponding ex vivo images: axial view from MPR (A), optical photography of the same colon shortly after MRI (B), and H&E-stained tissues (C). Locations of tumors were estimated from MRI with ImageJ software. Tissue slices were taken at the corresponding distance from the rectum in the extracted colon. Red arrows indicate estimated location for acquisition of images. Hematoxylin and eosin from untreated control animals are from distal (left) and proximal (right) colon. (D–F) Tumors depicted in coronal view correspond to tumors seen in excised colon with lumen exposed: coronal MRI (D), photograph of the exposed lumen (E), and H&E-stained tissue (F). Tissue slices were taken at the corresponding distance from the rectum in the extracted colon (red arrows). Scale bar in H&E, 2 mm.

Figure 6

Tumor volumes measured from MRI correlate with volumes measured by caliper. (A) Tumor volume determined from MRI matches closely to the tumor volume measured after necropsy. The volumes of eight tumors from two animals were compared. (B) Individual tumor growth rates were followed over time. Four individual tumor volumes from the same mouse were calculated at days 35 and 58 after AOM. Volumes were calculated from multiple imaging sections using ImageJ and compared with digital measurements from optical images of H&E stains or with ex vivo caliper measurements, where vol = πr²h.