Immunohistochemical and Transcriptional Analysis of SARS-CoV-2 Entry Factors and Renin-Angiotensin-Aldosterone System Components in Lethal COVID-19
<b><i>Introduction:</i></b> Since angiotensin converting enzyme-2 (ACE2) was discovered as an essential entry factor of SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2), there has been conflicting evidence regarding the role of renin-angiotensin-aldosterone system (RAAS) in COVID-19. This study elucidates pulmonary expression patterns SARS-CoV-2 entry factors (ACE2 and transmembrane protease serine subtype 2, TMPRSS2) and RAAS components in lethal COVID-19. <b><i>Methods:</i></b> Lung tissue from COVID-19 autopsies (<i>n</i> = 27) and controls (<i>n</i> = 23) underwent immunohistochemical staining for RAAS components (angiotensin receptors 1 and 2, ACE2 and Mas-receptor) and bradykinin receptors 1 and 2. Staining of individual cellular populations (alveolar pneumocytes [ALV], desquamated cells [DES] and endothelium [END]) was measured by a binary scale (positive/negative). SARS-CoV-2 was detected using immunohistochemistry against nucleocapsid protein, <i>in-situ</i> hybridization and quantitative reverse transcriptase polymerase chain reaction. Gene expression profiling for <i>ACE2, ACE</i> and <i>TMPRSS2</i> was performed. <b><i>Results:</i></b> Subtle differences were observed when comparing COVID-19 patients and controls not reaching statistical significance, such as a higher incidence of ACE2-positivity in END (52% vs. 39%) but lower positivity in ALVs (63% vs. 70%) and an overall downregulation of <i>ACE2</i> gene expression (0.25 vs. 0.55). However, COVID-19 patients with RAAS inhibitor (RAASi) intake had significantly shorter hospitalization times (5 vs. 12 days), higher viral loads (57,517 vs. 15,980/10<sup>6</sup> RNase P-gene copies) and decreased <i>ACE/ACE2</i>-expression ratios (4.58 vs. 11.07) than patients without. <i>TMPRSS2</i> expression was significantly (1.76-fold) higher in COVID-19 patients than controls. <b><i>Conclusion:</i></b> Our study delineates the heterogeneous expression patterns of RAAS components in the lungs, which vary amongst cellular populations, and implies that COVID-19 patients with RAASi-intake present with a more rapid disease progression, although this requires further investigation.