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Micro-CT Imaging with a Hepatocyte-selective Contrast Agent for Detecting Liver Metastasis in Living Mice

Posted Sep 30, 2008 Updated Apr 7, 2023
By Hye Won Kim MD , Quan-Yu Cai MD , Hong Yeoung Jun , Kwon Su Chon PhD , Seung Hoon Park MD , Seung Jae Byun MD , Myung Soo Lee MD , Jae Min Oh MD , Hun Soo Kim MD , Kwon-Ha Yoon MD
8 min read

Rationale and Objectives

Micro-computed tomography (CT) is a important tool for longitudinal imaging of tumor development. The detection and monitoring of tumors in the liver in live animals using micro-CT is challenging. We evaluated the feasibility of high-resolution micro-CT enhanced with a hepatocyte-selective contrast agent for detecting liver metastases in a live murine model.

Materials and Methods

Hepatic metastases were induced in 10 BALB/C mice. Two mice each were randomly selected on days 3, 5, 7, 10, and 13 after CT26 colon adenocarcinoma cells were injected into the portal vein; micro-CT imaging was performed at 10 minutes and 4 hours after intravenous administration of a hepatocyte-selective contrast agent at a dose of 0.4 mL/mouse. The attenuation values of the normal liver and the tumors were obtained. The number of metastases was counted and their sizes were measured on the micro-CT images. Gross or histopathologic evaluation was performed for correlating the liver tumors with the micro-CT images.

Results

A total of 74 separate tumor sites larger than 300 μm in diameter were detected on pathologic examination of the mice that were sacrificed 7 days after cell injection. On micro-CT, 66 of 74 tumors were detected (83.8%). The smallest tumor detected on micro-CT was 300 μm. There were eight false-negative readings on micro-CT. The sizes of the individual liver metastases measured by micro-CT and on the excised specimen were highly correlated ( P < .001). The correlation between the CT scan measurement and the actual measurement was r = 0.8354 ( P < .0001).

Conclusions

High-resolution micro-CT enhanced with a hepatocyte-selective contrast agent can be a promising tool for detecting liver metastases in a live murine model.

The liver is a common site of metastases from various primary tumors, and so it is an important area of metastasis research ( ). Rodent models that mimic the development of metastatic disease in the human liver are increasingly being recognized as powerful tools for the development of anticancer drugs and for evaluating the efficacy of novel therapeutics in the preclinical setting ( ). Noninvasive longitudinal imaging of rodent models would decrease experimental variability and provide a more accurate assessment of metastatic progression and the efficacy of therapeutic interventions ( ). Several longitudinal imaging modalities are now used, including magnetic resonance imaging, x-ray computed tomography (CT), positron emission tomography, and fluorescent and bioluminescent imaging.

Micro-CT has been applied in biomedical research mainly for studying bones and lung disease, for which the natural contrast between bone and air and the surrounding soft tissues are provided ( ). Micro-CT increased the spatial resolution noninvasively and it allowed very high precision for localization of bony changes. It has been proposed by some researchers that small-animal x-ray CT is an important tool for longitudinal imaging of tumor development, for visualizing blood vessels and angiogenesis, and for following the response of tumors to preclinical therapeutic intervention ( ). However, the detection and monitoring of tumor in the liver in live animals using micro-CT is still challenging because of the poor natural contrast between the tumor and the liver parenchyma. When performing micro-CT imaging for the liver of small animals, it is not possible to use the conventional water-soluble contrast agents that are used in humans because they clear rapidly from the blood.

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Methods and materials

Tumor Model

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Micro-CT System

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Micro-CT Imaging

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Image Analysis

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Histopathologic Correlation

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Statistical Analysis

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Results

Histopathologic Findings of the Metastatic Liver Tumors

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Figure 1, Photographs of the hematoxylin and eosin staining of the hepatic metastasis murine model (40× magnification). (a) Some necrotic areas were found on the peripheral areas of the livers at day 3 after cell injection. (b) A small tumor (<300 μm) was observed and the necrotic areas had shrunk at day 5 after cell injection. (c) Discrete tumors larger than 300 μm were observed and the necrotic areas were completely dissolved at day 7 after cell injection. (d) Multiple innumerable tumors were observed at day 10 after cell injection.

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Liver Parenchyma and Tumor Attenuation on the Micro-CT Images

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Figure 2, Serial micro–computed tomography (CT) images acquired after a hepatocyte-specific contrast agent injection. The baseline image before contrast agent injection shows the lack of contrast between soft tissues (a) . On the micro-CT image with 10-minute scan, the pulmonary and hepatic vessels were well visualized as bright lines (b) . Four-hour scan shows high attenuation of the liver parenchyma, which was reached a maximum value of contrast enhancement (c) .

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Table 1

Mean CT Number and Standard Deviation Measured in the Livers and Tumors of the Mice

Time (h) Liver Parenchyma Mean ± SD (HU) Tumor Mean ± SD (HU) Statistical Analysis ( P value) Precontrast 77.8 ± 5.76 71.7 ± 7.73 .132062 10 min 126.3 ± 25.15 80.82 ± 21.72 .011595 4 h 191.14 ± 34.24 84.05 ± 14.28 .00021

CT: computed tomography; SD: standard deviation; HU: Hounsfield units.

Figure 3, Contrast enhancement of the liver parenchyma, the inferior vena cava (IVC), and the tumor at 10 minutes and 4 hours after contrast agent injection. The graph represents the significant difference of contrast enhancement between the liver parenchyma and tumor at 4 hours after contrast agent injection. HU, Housefield Units.

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Micro-CT Images Correlated with the Histopathology

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Figure 4, Micro–computed tomographic image at day 13 after cell injection. After injection of a hepatocyte-specific contrast agent, numerous low-density nodules in the liver ( arrows ) were detected (a) . On gross morphology of the liver, innumerable metastatic nodules were found at the surface of the liver (b) .

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Table 2

Detectability of the Experimental Liver Tumors

Tumor Size No. on Micro–CT No. on Pathology Detectability >500 μm 32 32 100% 300–500 μm 34 42 80.9% Total 66 74 83.8%

CT: computed tomography.

Figure 5, Micro–computed tomographic image (a) at day 7 after cell injection. After injection of a hepatocyte-specific contrast agent, a low-density nodule of 500 μm in size is seen ( arrow ) in the right lobe of the liver, which was correlated with histopathology (b) .

Figure 6, Micro–computed tomographic image (a) at day 7 after cell injection after injection of a hepatocyte-specific contrast agent shows a low-density nodule of 300 μm in size ( arrow ) in the left lobe of the liver, which was correlated with the histopathology (b) .

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Figure 7, Regression analysis shows that the correlation between the computed tomographic (CT) scan measurement and the actual measurement was r = 0.8354 ( P < .001). Units: micrometers.

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Discussion

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Academic Radiology, Volume 15, Issue 10
Journals General Radiology
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