The contrasting role of nasopharyngeal angiotensin converting enzyme 2 (ACE2) transcription in SARS-CoV-2 infection: A cross-sectional study of people tested for COVID-19 in British Columbia, Canada

Abstract

Background: Angiotensin converting enzyme 2 (ACE2) protein serves as the host receptor for SARS-CoV-2, with a critical role in viral infection. We aim to understand population level variation of nasopharyngeal ACE2 transcription in people tested for COVID-19 and the relationship between ACE2 transcription and SARS-CoV-2 viral load, while adjusting for expression of: (i) the complementary protease, Transmembrane serine protease 2 (TMPRSS2), (ii) soluble ACE2, (iii) age, and (iv) biological sex. The ACE2 gene was targeted to measure expression of transmembrane and soluble transcripts.

Methods: A cross-sectional study of n = 424 "participants" aged 1-104 years referred for COVID-19 testing was performed in British Columbia, Canada. Patients who tested positive for COVID-19 were matched by age and biological sex to patients who tested negative. Viral load and host gene expression were assessed by quantitative reverse-transcriptase polymerase chain reaction. Bivariate analysis and multiple linear regression were performed to understand the role of nasopharyngeal ACE2 expression in SARS-CoV-2 infection.

Findings: Analysis showed no association between age and nasopharyngeal ACE2 transcription in those who tested negative for COVID-19 (P = 0•092). Mean relative transcription of transmembrane (P = 0•00012) and soluble (P<0•0001) ACE2 isoforms, as well as TMPRSS2 (P<0•0001) was higher in COVID-19-negative participants than COVID--19 positive ones, yielding a negative correlation between targeted host gene expression and positive COVID-19 diagnosis. In bivariate analysis of COVID-19-positive participants, transcription of transmembrane ACE2 positively correlated with SARS-CoV-2 viral RNA load (B = 0•49, R²=0•14, P<0•0001), transcription of soluble ACE2 negatively correlated (B= -0•85, R²= 0•26, P<0•0001), and no correlation was found with TMPRSS2 transcription (B= -0•042, R²=<0•10, P = 0•69). Multivariable analysis showed that the greatest viral RNA loads were observed in participants with high transmembrane ACE2 transcription (Β= 0•89, 95%CI: [0•59 to 1•18]), while transcription of the soluble isoform appears to protect against high viral RNA load in the upper respiratory tract (Β= -0•099, 95%CI: [-0•18 to -0•022]).

Interpretation: Nasopharyngeal ACE2 transcription plays a dual, contrasting role in SARS-CoV-2 infection of the upper respiratory tract. Transcription of the transmembrane ACE2 isoform positively correlates, while transcription of the soluble isoform negatively correlates with viral RNA load after adjusting for age, biological sex, and transcription of TMPRSS2.

Funding: This project (COV-55) was funded by Genome British Columbia as part of their COVID-19 rapid response initiative.

Keywords: ACE2 transcription; COVID-19; Cross-sectional study; Nasopharynx; SARS-CoV-2; Soluble ACE2; Viral RNA load.

Fig. 1

Relationship between age and nasopharyngeal transmembrane ACE2 transcription in unmatched COVID-19-negative participants. Boxplots of transmembrane ACE2 transcription by 10-year age categories in unmatched COVID-19-negative participants between the ages of 19 and 98 (n = 198); boxes represent the Q1-Q3 interquartile range, whiskers represent 1.5x the Q1 or Q3 and horizontal lines the median transmembrane ACE2 transcription by age category. Participants who tested negative younger than 19 or older than 98 were excluded based on n<10 observations per age group. No difference was detected in mean transmembrane ACE2 transcription among age categories (ANOVA, P = 0•092).

Fig. 2

Relative nasopharyngeal transcription of targeted host genes by COVID-19 test result. Gene transcription is portrayed in kernel density plots stratified by COVID-19 test result. Probability densities of relative host gene transcription are shown by positive (red, transmembrane ACE2; blue, soluble ACE2; yellow, TMPRSS2) and negative COVID-19 test results (gray). Levene's test was used to detect non-equal variance in gene transcription for all host targets between COVID-19-negative and -positive participants. A two-tailed, paired t-test was used to examine mean difference in host gene transcription by COVID-19 test result assuming unequal variance: (a) transmembrane ACE2 (P = 0•00,012), (b) soluble ACE2 (P<0•0001) and (c) TMPRSS2 (P<0•0001).

Fig. 3

The association between transmembrane ACE2 transcription and SARS-CoV-2 RNA load in nasopharyngeal tissue differs by the amount of soluble ACE2 transcription. Soluble ACE2 transcription was categorized into low, mean and high levels to demonstrate the relationship. The low category (gray) represents soluble ACE2 transcription one standard deviation or greater below the mean transcription (n = 74). The mean category (orange) codes for the mean soluble ACE2 transcription at zero standard deviations. The high category (blue) indicates soluble ACE2 transcription one standard deviation or greater above mean transcription (n = 80). Shaded areas represent 95% confidence intervals, solid lines represent Β-coefficients for simple slopes derived from multiple linear regression. Figure S4 visualizes the corresponding effect of soluble ACE2 transcription +/- 2 SD or greater from the mean; estimates are reported in Table 2.