Micro-computed tomography of pulmonary fibrosis in mice induced by adenoviral gene transfer of biologically active transforming growth factor-β1

Abstract

Background: Micro-computed tomography (micro-CT) is a novel tool for monitoring acute and chronic disease states in small laboratory animals. Its value for assessing progressive lung fibrosis in mice has not been reported so far. Here we examined the importance of in vivo micro-CT as non-invasive tool to assess progression of pulmonary fibrosis in mice over time.

Methods: Pulmonary fibrosis was induced in mice by intratracheal delivery of an adenoviral gene vector encoding biologically active TGF-β1 (AdTGF-β1). Respiratory gated and ungated micro-CT scans were performed at 1, 2, 3, and 4 weeks post pulmonary adenoviral gene or control vector delivery, and were then correlated with respective histopathology-based Ashcroft scoring of pulmonary fibrosis in mice. Visual assessment of image quality and consolidation was performed by 3 observers and a semi-automated quantification algorithm was applied to quantify aerated pulmonary volume as an inverse surrogate marker for pulmonary fibrosis.

Results: We found a significant correlation between classical Ashcroft scoring and micro-CT assessment using both visual assessment and the semi-automated quantification algorithm. Pulmonary fibrosis could be clearly detected in micro-CT, image quality values were higher for respiratory gated exams, although differences were not significant. For assessment of fibrosis no significant difference between respiratory gated and ungated exams was observed.

Conclusions: Together, we show that micro-CT is a powerful tool to assess pulmonary fibrosis in mice, using both visual assessment and semi-automated quantification algorithms. These data may be important in view of pre-clinical pharmacologic interventions for the treatment of lung fibrosis in small laboratory animals.

Figure 1

Respiratory ungated in vivo micro-CT at 1, 2, 3 and 4 weeks after intratracheal instillation of control vector or AdTGFβ1. (A) Lung micro-CT of mice treated with the control vector AdDL70-3. (B) Lung micro-CT of mice treated with AdTGF-β1. Only minor changes are seen in the control group, the consolidation in the fibrosis group is illustrated with pronounced micro-CT consolidation at 2 and 3 weeks. Representative images are shown.

Figure 2

Correlation of visual consolidation assessment and histological assessment by Ashcroft scoring in animals after treatment with control vector AdDL70-3 (A, showing very low Ashcroft histology scores (i.e. no fibrosis), and low visual consolidation assessment scores) or AdTGF-β1 (B). Mean consolidation assessment values of the 3 observers are plotted against Ashcroft scores of each animal in respiratory gated and respiratory ungated exams, as indicated. Linear interpolation is given for respiratory gated and ungated exams, respectively. Note the different scaling.

Figure 3

Semiautomated region growing segmentation for assessment of pulmonary consolidation 3 weeks after instillation of control vector AdDL70-3 (A-C) or AdTGF-β1 (D-E). Aerated lung volume is used as an inverse surrogate marker for pulmonary fibrosis. (A, D) Axial micro-CT. (B, E) Placement of seed points for the region growing segmentation in the aerated lung. Seed points are enlarged for better detection. (C, F) Segmentation result shows aerated lung volume in white.

Figure 4

Correlation of aerated lung volume determined by region growing segmentation with histological assessment of lung fibrosis by Ashcroft scoring in mice treated with control vector (A, showing very low Ashcroft histology scores (i.e. no fibrosis) and normal aerated lung volume) or AdTGF-β1 (B). Aerated lung volumes are plotted against Ashcroft scores for each animal in respiratory gated and respiratory ungated exams, as indicated. Linear interpolation is given for respiratory gated and ungated exams, respectively. Note the different scaling.