Ex vivomicro-computed tomography analysis of bleomycin-induced lung fibrosis for preclinical drug evaluation

Abstract

Research into the pathogenesis underlying the development of idiopathic pulmonary fibrosis is hampered by a repertoire of animal models that fail to recapitulate all the features of the human disease. Better use and understanding of what the animal models represent may improve clinical predictability. We interrogated ex vivo micro-computed tomography (CT) as a novel end-point measure in the mouse model of bleomycin-induced lung fibrosis (BILF), and to evaluate a therapeutic dosing regimen for preclinical drug evaluation.A detailed characterisation of BILF was performed using standard end-point measures (lung hydroxyproline and histology). High resolution micro-CT (,13.7 mm voxel size) was evaluated for quantifying the extent and severity of lung fibrosis.The period from 14 to 28 days following bleomycin instillation represents progression of established fibrosis. A therapeutic dosing regimen during this period was validated using a transforming growth factor-b receptor-1 kinase inhibitor, and micro-CT provided a highly sensitive and quantitative measure of fibrosis. Moreover, fibrotic lesions did not completely resolve, but instead persisted for o6 months following a single insult with bleomycin.Ex vivo micro-CT analysis of BILF allows robust evaluation of therapeutic dosing once fibrosis is already well established, requiring fewer mice than conventional biochemical end-points. @ERSpublications Micro-CT has improved out understanding of the bleomycin model of fibrotic lung disease for preclinical drug evaluation

Quantification of Pulmonary Fibrosis in a Bleomycin Mouse Model Using Automated Histological Image Analysis

“…Hence, in vivo micro-CT resolutions are much lower compared to the high resolution (3.5 μm/pixel) used in the automated histological analysis. An improved micro-CT analysis using higher sensitivity involving high resolution scanning [13,34,38,39] might also be used to enhance accurate quantification of low level of fibrotic alterations. High resolutions (1–2 μm/pixel) have also been achieved with micro-CT analysis on ex vivo fixed lungs [34].…”

“…In contrast, the automated analysis is applied to the entire lung section. It should be noted that micro-CT high resolutions are associated with sometimes prohibitively long scan/reconstruction times of several hours per sample precluding high throughput [13,34]. In contrast the automated analysis acquisition/analysis time is much shorter in the range of 1 hour for 100 entire lung sections.…”

“…In contrast the automated analysis acquisition/analysis time is much shorter in the range of 1 hour for 100 entire lung sections. Ex vivo micro-CT may also be used for a 3D evaluation of severity of lung fibrosis providing a more sensitive and quantitative measure as well as a marked reduction of scanning time [13]. …”

Claudin-18–mediated YAP activity regulates lung stem and progenitor cell homeostasis and tumorigenesis

“…Lungs were fixed and inflated with 4% paraformaldehyde (PFA) at 20 cmH 2 O pressure overnight and then incubated through a serial ethanol gradient (50%, 70%, 80%, 90%, and 100%) followed by incubation with 100% hexamethyldisilazane overnight before air drying (80). Lung specimens were scanned at an isotropic resolution of 10 microns at 45 kVp, 200 mAs (81).…”

Review of the British Thoracic Society Winter Meeting 2014, 3-5 December, London, UK

“…In the first session, Professor Clive Page (London) described the usefulness of in vivo models to identify new drugs for respiratory disease, while Dr Chris Scotton (Exeter) spoke about novel ways of using in vivo imaging in murine models of lung disease,9 and Professor Stephen Renshaw (Sheffield) described how the zebrafish, which does not have lungs, is a useful model for studying pulmonary inflammation 10. In the second session, Professor Eric White (Michigan, USA) discussed novel 3D cell culture models for studying fibrotic lung disease, Professor Donna Davies (Southampton) described how novel tissue-engineered constructs with microfluidics platforms to deliver inflammatory cells will be used to develop novel therapeutic treatments for asthma, and Dr Carmel Nanthakumar (London) discussed how novel in vitro platforms are being used in industry to develop new anti-fibrotic drugs.…”