Pivotal role of micro-CT technology in setting up an optimized lung fibrosis mouse model for drug screening

Abstract

Idiopathic pulmonary fibrosis (IPF) is a progressive disease with no curative pharmacological treatment. The most used animal model of IPF for anti-fibrotic drug screening is bleomycin (BLM)-induced lung fibrosis. However, several issues have been reported: the balance among disease resolution, an appropriate time window for therapeutic intervention and animal welfare remain critical aspects yet to be fully elucidated. In this study, C57Bl/6 male mice were treated with BLM via oropharyngeal aspiration (OA) following either double or triple administration. The fibrosis progression was longitudinally assessed by micro-CT every 7 days for 4 weeks after BLM administration. Quantitative micro-CT measurements highlighted that triple BLM administration was the ideal dose regimen to provoke sustained lung fibrosis up to 28 days. These results were corroborated with lung histology and Bronchoalveolar Lavage Fluid cells. We have developed a mouse model with prolonged lung fibrosis enabling three weeks of a curative therapeutic window for the screening of putative anti-fibrotic drugs. Moreover, we have demonstrated the pivotal role of longitudinal micro-CT imaging in reducing the number of animals required per experiment in which each animal can be its own control. This approach permits a valuable decrease in costs and time to develop disease animal models.

A fully automated micro‑CT deep learning approach for precision preclinical investigation of lung fibrosis progression and response to therapy

“…Total expiratory lung volume (V P02 ) was significantly increased on day 7 in the BLM and BLM + NINT groups compared to the SAL control, both in the whole parenchyma and in individual lung lobes, suggesting that this parameter may reflect the inflammatory response to BLM administration, as previously reported [ 20 , 23 ]. V P02 tended to decrease, although non-significantly in the BLM group, at subsequent time-points in the whole and in the left lungs, suggesting that on day 21 the inflammation in the left lung was replaced by fibrotic and scarred tissue.…”

“…Effective preclinical discovery of new drugs requires the development of reliable animal models of human diseases and highly efficient approaches for their detailed investigation. In various animal models, µCT imaging has proven to be a powerful tool to visualize and precisely quantify the dynamic evolution and regional severity of disease [ 20 , 23 ]. However, to take full advantage of this imaging approach, it is necessary to automate the post-processing phase of analysis, including lung segmentation.…”

“…At first, 23 C57bl/6 male mice, 8 to 10-week-old, provided by Envigo (San Pietro al Natisone, Udine, Italy), were acclimatized to the local vivarium conditions (room temperature: 20–24 °C; relative humidity: 40–70%; 12-h light–dark cycle) for at least 5 days, having free access to standard rodent chow and softened tap water. Nineteen mice were then lightly anesthetized with 2.0% isoflurane delivered in a box and administered with bleomycin (Baxter Oncology GmbH) 10 µg/mouse in 50 μl saline (0.9%), while four mice received only 50 μl saline (hereafter designated as “SAL mice”) via oropharyngeal aspiration (OA) using a micropipette [ 20 ]. This procedure was performed on days 0 and 4, as reported in the scheme in Additional file 1 : Fig.…”

“…S1A. The BLM dose utilized in this experiment has been selected to perform routine in-vivo drug screening experiments [ 20 ]. On day 7, twelve BLM-treated mice were randomly selected to receive Nintedanib (hereafter called “BLM + NINT mice”) as the reference tool compound since it has been approved for human IPF treatment [ 21 ].…”

Airway remodeling heterogeneity in asthma and its relationship to disease outcomes

“…More recently, microCT with a spatial resolution of up to 1 μm has enabled volumetric imaging and quantitative assessment of the small conducting airways, respiratory airways, and alveoli ( Haefeli-Bleuer and Weibel, 1988 ; McDonough et al, 2011 ). To date, microCT has been applied to assess the morphometry of the normal human lung and how it is modified in the pathology of chronic obstructive pulmonary disease (COPD) ( McDonough, 2012 ; Koo et al, 2018 ), lung cancer ( Holbrook et al, 2021 ), Idiopathic Pulmonary Fibrosis (IPF) ( Khalajzeyqami et al, 2022 ), cystic fibrosis (CF) ( Wielpütz et al, 2011 ), and bronchiectasis ( Wurnig et al, 2014 ). However one limitation of microCT is that it cannot be used in vivo in humans due to the small field of view and higher radiation dose.…”