Representative images of original and 2D-reconstructed lung sections for the quantification and the mapping of pulmonary tissue density by means of digital automatic analysis.
Masson trichrome-stained images (A, E) and their respective 2D-reconstructed images (C, G) correspond to a lung section from saline control (A, C) and BLM-treated (0.75 mg/kg) lungs (E, G). Panels B, D, F, H show details from their original stained (B, F) and 2D-reconstructed (D, H) images at higher magnification. Lung tissue density was determined from thousands of micro-tiles crisscrossing entire lung sections. For mapping thousands tissue density values throughout lung section density values were graded in 20 classes of increasing values (I) and pseudocolours were assigned from blue (low density values) to yellow (high density values) according to their classification. Note that high density values (yellow) were restricted in alveolar parenchyma of BLM-treated lung (G, H) and located in fibrotic lesions evidenced in the respective original stained image (E, F). The frequency of tissue density (I) was determined from the classification of the whole unitary density values obtained in each lung section (A, E). HDFm index corresponds to the sum of the frequencies of the highest tissue density (classes 12 to 20) expressed in fibrotic foci. The mean tissue density (Dm) (J) was evaluated for each lung section from thousands of micro-tiles. Scale bars: 1 mm (A, C, E, G), 100 μm (B, D, F, H).