Each datapoint represents a single cell.  All major cell types of the small airway epithelium (SAE), including basal cells, intermediate cells, club cells, mucus cells, and ciliated cells, express ACE2.  Because of technology limitations, only 1-2% of all of the tested cells were found to express ACE2.  It is thought that cells expressing lower amounts of the enzyme went uncounted, and these reported data are an underrepresentation of the number of cells that express ACE2.  

ACE2 is Expressed in Major Cell Types in the Small Airway Epithelium

RNA samples were obtained by brushing tracheal basal cells (BCs) on day 0.  Cell colonies were then created on an air–liquid interface culture, and additional samples were taken every 7 days.  ACE2 expression levels were determined using Affymetrix HG-U133 Plus 2.0 microarrays.  It was found that as primary basal cells, which are the precursors for other epithelial cells, differentiate into these said cells, their ACE2 expression significantly increases (p < 10^-5).  

ACE2 Expression Increases in Primary Epithelium Basal Cells Throughout Differentiation

Epithelium samples were obtained by brushing the trachea, large airway epithelium (LAE), and small airway epithelium (SAE).  Expression was quantified using Affymetrix HG-U133 Plus 2.0 microarrays.  It was observed that all parts of the tracheobronchial tree express ACE2 in normal health with the trachea and LAE expressing it more than the SAE.  

ACE2 is Expressed in All Parts of the Tracheobronchial Tree in Healthy Nonsmokers

- expression in the tracheobronchial tree
- expression in primary basal cells
- expression by cell type in the small airway

Michigan State University

One of the primary manifestations of COVID-19 is respiratory disease.  ACE2 expression was assessed in human airway epithelial tissue to better understand how SARS-CoV-2 interacts. with these cells. Studies included:

- ACE2 expression in healthy nonsmokers
- ACE2 expression in smokers
- ACE2 expression modulation by air pollution
- ACE2 expression modulated by miRNA (miR-1246)
- ACE2 expression in basal cells from the large and small airway epithelia of healthy nonsmokers

ACE2 Expression in Human Airway Epithelial Tissue

Data were generated by the O’Beirne research group.  The relationship between ACE2 expression in the small airway epithelium (SAE) and average monthly pollution levels in New York City, using data from the EPA, was assessed.  There was no difference reported between expression and pollution levels in nonsmokers (white dots), smokers (gray dots), or a combination of the two groups.  It should be noted that the maximum 30-day mean PM2.5 concentration was 18 μg/m^3 which is considered safe by the EPA.  

ACE2 Expression in the Small Airway Epithelium is Not Associated with Air Pollution in New York City

- Smoking Upregulates Angiotensin-Converting Enzyme-2 Receptor: A Potential Adhesion Site for Novel Coronavirus SARS-CoV-2 (Covid-19)

Clinical imaging data suggest that the small airway epithelium (SAE) in an important SARS-CoV-2 binding site.  
- expression versus nonsmokers
- expression versus nonsmokers by sex

ACE2 Expression in the Airway Epithelium of Smokers

ACE2 Expression in the Airway Epithelium of Healthy Nonsmokers

A main feature of symptomatic COVID-19 disease is respiratory infection. Severe damage to the lungs during respiratory distress may cause pulmonary fibrosis, necessitating lung transplants. Lasting damages to the lungs may occur even in recovering patients with mild infections. SARS-CoV-2 targets the following cell types:

- Type I pneumocytes (AT1) 
- Type II pneumocytes (AT2) 
- alveolar macrophages

ACE2 expression is primarily seen in AT2 cells which have been shown to be involved in the reproduction and transmission of the virus.  Viral targeting of alveolar macrophages is evidenced by N antigen expression.

Effects of COVID-19 on the Lungs

miRNAs that are homologous to the 3' end of the ACE2 gene were assessed using the data set from the Wang research group to determine if ACE2 expression was modulated by smoking.  miR-1246 is complementary to the  3′ untranslated region (3′UTR) of ACE2 mRNA.  Predicted base pairing was assessed using TargetScanHuman 7.2 (http://www.targetscan.org/vert_72/).  Affymetirx miRNA 2.0 arrays and TaqMan PCR reveal that miR-1246 expression is significantly decreased in smokers compared to nonsmokers (**P < 0.01 and ***P < 0.001).  This miRNA may be involved in the mechanism that regulates ACE2 expression in the SAE. 

miR-1246 Levels May Modulate ACE2 Expression Levels in the Small Airway Epithelium

BCi-NS1.1 cells were derived from a single basal cell from the large airway epithelium (LAE) of a healthy nonsmoker, and hSABCi-NS1.1 cells were derived from a single basal cell from the small airway epithelium (SAE) of a healthy nonsmoker.  Expression levels were determined using HiSeq 4000 RNA-seq data at day 0 (baseline) and after 28 days on an air-liquid interface (ALI).  Notable findings include:

- Both cell lines showed an increase in ACE2 expression after 28 days of differentiation
- ADAM10, ADAM17, FURIN, CTSL, and PI4KB expression was relatively maintained throughout differentiation in both cell lines
- TMPRSS2 was notably less expressed in undifferentiated LAE basal cells than in SAE basal cells, and their expression increased.  However, its expression was maintained in SAE basal cells.  
- TPRSS11A and TMPRSS11D expression remained low in both LAE and SAE cells throughout differentiation

Because these cell lines are immortal, they may continue to be useful in SARS-CoV-2 research - especially for early drug screening.  

ACE2 and Other Cellular Proteins Related to B-Coronavirus Infection Expression Increases in Basal Cells from the Large Airway Epithelium Throughout Differentiation

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