Micro-CT analysis of paraffin embedded lung tissue : is small airway obstruction an early feature of COPD?

Rationale: Airflow obstruction, the hallmark characteristic of Chronic Obstructive Pulmonary Disease (COPD) has long been attributed to a combination of small airways disease and emphysematous destruction, however, the relative role of each pathological feature has not been well understood. McDonough et al., recently reported a significant reduction in terminal bronchiolar number in end-stage COPD compared to healthy controls. Further, the study reported that the loss of terminal bronchioles occurred in regions of lung with and without emphysema, posing the question, which pathological event occurs first? Hypothesis: Narrowing and obliteration of small airways precedes emphysematous destruction in COPD. Methods: Lung samples obtained from patients undergoing surgical treatment for lung cancer were formalin inflated, sliced and sampled prior to paraffin embedding (FFPE). A Nikon Metrology micro-CT scanner was used to scan these FFPE cores, and volumetric data sets were examined to determine the number of terminal bronchioles/ml (TB/ml) and presence of emphysema using mean linear intercept (Lm). Semi-automatic segmentation enabled 3D reconstruction of the airways to characterize structure and calibre, and scout for regions of interest that were sectioned and stained with Movat’s Pentachrome, for analysis of airway morphology. Results: Using a novel combination of multidetector-CT, micro-CT and histology, this study demonstrates that micro-CT scans of FFPE cores provide adequate contrast to determine Lm values that were validated by histology. We report that the number of terminal bronchioles/ml is significantly decreased from 6.2 TB/ml in the controls, to 4.6 TB/ml in mild/moderate COPD patients. When the number of TB/ml is compared to Lm we demonstrate that terminal bronchioles are destroyed in tissues where no emphysema is present. These lesions were confirmed by histology, using the micro-CT images to precisely locate the airway lesions, enabling efficient sectioning of the FFPE cores for further characterization. Conclusions: Treatment of COPD has traditionally focused on patients with severe disease and no current pharmacological therapies have been shown to affect long term, lung function decline. Our findings suggest that irreversible pathological events occur in the early stages of disease and emphasize the importance of early diagnosis and intervention to modify the progression of this debilitating respiratory disorder.Medicine, Faculty ofAnesthesiology, Pharmacology and Therapeutics, Department ofGraduat