Abstract
BackgroundUnderstanding the three-dimensional (3-D) micro-architecture of lung tissue can provide insights into the pathology of lung disease. Micro computed tomography (µCT) has previously been used to elucidate lung 3D histology and morphometry in fixed samples that have been stained with contrast agents or air inflated and dried. However, non-destructive microstructural 3D imaging of formalin-fixed paraffin embedded (FFPE) tissues would facilitate retrospective analysis of extensive tissue archives of lung FFPE lung samples with linked clinical data.MethodsFFPE human lung tissue samples (n = 4) were scanned using a Nikon metrology µCT scanner. Semi-automatic techniques were used to segment the 3D structure of airways and blood vessels. Airspace size (mean linear intercept, Lm) was measured on µCT images and on matched histological sections from the same FFPE samples imaged by light microscopy to validate µCT imaging.ResultsThe µCT imaging protocol provided contrast between tissue and paraffin in FFPE samples (15mm x 7mm). Resolution (voxel size 6.7 µm) in the reconstructed images was sufficient for semi-automatic image segmentation of airways and blood vessels as well as quantitative airspace analysis. The scans were also used to scout for regions of interest, enabling time-efficient preparation of conventional histological sections. The Lm measurements from µCT images were not significantly different to those from matched histological sections.ConclusionWe demonstrated how non-destructive imaging of routinely prepared FFPE samples by laboratory µCT can be used to visualize and assess the 3D morphology of the lung including by morphometric analysis.
Highlights
The airways, vasculature and lymphatics of the human lung form a complex interrelated three dimensional (3D) network that structurally ranges from macroscopic to microscopic scales
Airspace size was measured on μCT images and on matched histological sections from the same formalin-fixed paraffin embedded (FFPE) samples imaged by light microscopy to validate μCT imaging
We demonstrated how non-destructive imaging of routinely prepared FFPE samples by laboratory μCT can be used to visualize and assess the 3D morphology of the lung including by morphometric analysis
Summary
The airways, vasculature and lymphatics of the human lung form a complex interrelated three dimensional (3D) network that structurally ranges from macroscopic to microscopic scales. Macroscopic structures greater than 2.3 mm in diameter can be visualized in vivo using clinical multi-detector computed tomography (MDCT) scanners [1]. For diseases such as asthma and chronic obstructive pulmonary disease (COPD) the small airways with internal diameters of less than 2mm with no cartilage support [2,3] are thought to be the major site of airflow obstruction. The use of μCT has previously been to image air-inflated and fixed whole murine lung [8] or human lung samples [3,9] visualising small airways and alveolar structures without physically sectioning the sample. Non-destructive microstructural 3D imaging of formalin-fixed paraffin embedded (FFPE) tissues would facilitate retrospective analysis of extensive tissue archives of lung FFPE lung samples with linked clinical data
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