Fig 3. ACE2 protein expression increases with age in ventilated patients.

A) Representative images of lung stained for ACE2 from a 40-year-old man with acute lung injury are shown using a 1x objective (left, scale bar 3mm) with the region outlined by the red dashed box magnified at low power (middle, scale bar 200μm) and a second field at high power (right, scale bar 50μm). Clusters of reactive AT2 cells (red arrowheads) are present along the alveolar septum which exhibit low level ACE2 expression. B) Representative images of lung stained for ACE2 from a 67-year-old man with acute lung injury superimposed on fibrosing interstitial lung disease are shown using a 1x objective (left, scale bar 3mm) with the region outlined by the red dashed box magnified at low power (middle, scale bar 200μm) and a second field at high power (right, scale bar 50μm). Numerous reactive AT2 cells (red arrowheads) exhibiting nucleomegaly and abundant cytoplasm can be seen demonstrating strong ACE2 staining. C) Quantitative IHC for ACE2 was carried out on samples from ventilated patients (n = 12 samples from 11 patients). Total ACE2 expression from 5 low power fields is plotted relative to the patient’s age at the time of specimen collection. A linear fit to the data is indicated by the dashed line with the 95% confidence interval highlighted in grey. D) The same specimens quantitated in (C) were normalized by cellularity and the average ACE2 expression per cell is plotted along with a linear fit to the data and its 95% confidence interval. The red arrow in (C) and (D) indicates a patient providing 1 sample from the left lung and 1 sample from the right lung during the same procedure, utilized as a control for intra-individual reproducibility. The green and blue circle in (C) and (D) indicate the staining intensity of the samples depicted in Fig 5A and 5B, respectively. Sections were stained for ACE2 using DAB and counterstained with hematoxylin.