To our knowledge, this is the first report on ACE2 expression in Blacks versus Whites from normal epithelial cell. We noted that Blacks had lower levels of both ACE2 and TMPRSS2 expression. While the ACE2 approached but did not quite achieve conventional statistical significance, reassuringly using a multivariable model (with age, gender), it crossed the statistical significance threshold. Furthermore, ACE2 closely mirroring with TMPRSS2 which did achieve statistical significance in both the univariable and multivariable analyses.
The epidemiological data for racial disparities in Covid-19 is compelling. For instance, in a large cohort study in Louisiana, Blacks with Covid-19 had a relative risk of 1.96 (95% confidence interval 1.62–2.37) of being hospitalized when compared to Whites after adjusting for covariates (obesity, age, sex, Charlson Comorbidity Index score, residence in a low-income area, and insurance plan) . Similarly, Azar and colleagues noted that in 1052 confirmed cases, non-Hispanic Blacks had a 2.7-fold higher risk of hospitalization than non-Hispanic Whites even after adjusting for age, gender, comorbidities, and income . However, it is possible that there may be residual confounding that might occur from socioeconomic factors (living conditions, essential worker designation etc.). Racial health disparities (cancer, hypertension, etc.) are generally related to both biological and socioeconomic factors .
ACE2 is well established as the molecular receptor for the novel Coronavirus. Using transgenic mice, it has been shown that eliminating ACE2 markedly suppressed infections of Covid-19. Thus, at first pass, ACE 2 data may seem paradoxical. However, there is emerging evidence that ACE2 downregulation may potentiate the severity of inflammation which is a trigger towards ARDs and thus mortality from Covid-19. ACE 2 is responsible for degrading a variety of pro-inflammatory peptides such as angiotensin II. Increased angiotensin II leads to enhanced inflammation including macrophage activation with concomitant elaboration of interleukin 6, tumor necrosis factor alpha, and other inflammatory cytokines . Moreover, ACE2 also generates angiotensin 1–7 which has salutary effects interacting with the Mas receptor to counteracting the pro-inflammatory effects of angiotensin II . Rivellese and colleagues point out that ACE2 deficiency may occur in the elderly providing one potential mechanism for the higher mortality from Covid-19 . Indeed, in experimental models of pneumonia, ACE2 was protective against severe inflammation induced by acid . Furthermore, the powerful survival advantage effects of immune suppressants including the corticosteroid dexamethasone underscore the hyper-immune response . As pointed out by Zamai, the role of ACE is complex with an “Adverse Axis” and the ACE2 “Protective Axis” with Covid-19 infection disrupting the Yin-Yang leading to the vasoconstriction, inflammation etc. that characterizes serious/lethal infections .
One important caveat is that the epidemiology suggests that both the case fatality rate and prevalence of infection are higher. Our data suggests that ACE2 suppression may increase severity but would not explain increased prevalence of Covid-19 in Blacks. Indeed, since ACE2 is necessary for Covid-19 access to epithelium, this downregulation in Blacks would be predicted to and loss of ACE2 seems to decrease risk of infection. Similarly, since TMPRSS2 acts on the coronavirus Spike protein to allow virus internalization and thus suppression would also be anticipated to decrease risk of infection. Interestingly, ACE2 rectal levels did not parallel other Covid-19 severity risk factors (obesity, age, smoking status etc.) leading to the speculation of distinct modalities involved in racial differences .
The mechanisms of rectal mucosal ACE2 downregulations in Blacks is unclear. The determinants of ACE2 levels may be influenced by genetics (e.g., single-nucleotide polymorphisms) and epigenetics (e.g., methylation) both of which have shown to be common themes in racially distinct gene expression. From a genetic perspective, there are a number of single-nucleotide polymorphisms (SNPs) that have been shown to racially segregate . SNPs can lead to altered protein levels through alterations in regulatory units, splice sites etc. Additionally, ACE2 levels have been shown to be impacted by epigenetics including methylation . For SNPs which are clearly racially mediated, several lines of evidence indicate that racially mediated methylation is noted in cancer disparity studies .
These findings are consistent with data from Chen et al. where aging, a key risk factor for severity, was associated with a decreased tissue ACE2 level . Our data is in contradistinction to ACE2 increase in sputum of asthmatics which showed higher ACE2 expression . However, in this study, once patients were given inhaled corticosteroids, the racial difference abated suggesting this was a result of inflammatory rather than epithelial expression but this was removed with inhaled steroids suggesting non-epithelial drivers .
For the use of rectal biopsies as a sample of convenience, we acknowledge that bronchial epithelium might be more relevant. However, GI manifestations are frequent (~ 20% in Covid-19 infections) and includes diarrhea, vomiting, and anorexia. Additionally, ~ 50% of patients have viral RNA detectable in feces. Moreover, there is evidence of GI manifestations frequently parallel in overall disease . With regard to ACE2, its expression at both the mRNA and protein level in intestinal tissue is strongly relative to many other sites [8, 22]. While it is logical to extrapolate that the processes that drive ACE2 expression (SNPs, methylation) in the GI tract should have a more ubiquitous impact, this needs to be empirically corroborated. In this regard, some recent data suggests that factors that may impact ACE2 levels (age, obesity etc.) impact multiple epithelial sites . Moreover, recent single cell RNA sequencing study showed co-expression of ACE2 and TMPRSS2 mirrored in several tissue types including respiratory tree and colon . These authors also demonstrate that TMPRSS2 was only expressed in a subset of ACE2-positive cells supporting the role of other proteases and indicating the primacy of ACE2 for Covid-19 infection .
Other limitations include the modest sample size which precluded univariable (p = 0.07) analysis achieving statistical significance, but the multivariable analysis is reassuring. Other putative confounders such as hypertension or medications which may modulate ACE2 expression (e.g., ARBs have been shown to increase cardiac ACE2 levels) were unable to be assessed . In addition, the relative importance of these biological factors versus socioeconomic remains to be determined . Blacks are known to have many more underlying medical issues associated with Covid-19 susceptibility (obesity, diabetes, hypertension, etc.) . While our data used multivariable analysis to address confounders, we were only able to adjust for a limited number of variables leading to the possibility of unmeasured confounders. From a socioeconomic perspective, issues like more frequency of needing to work (essential workers in pandemic) and less ability to socially distance also may play a role in this disparity. However, there is precedence for biological factors involved with Covid-19 severity. For instance, a recent genome-wide association study indicated that blood types are associated with severity risk of Covid-19 . Importantly, it is well established that there are significant differences in blood types by race . It needs to be reiterated that our data does not indicate the relative contributions of biological versus socioeconomic factors in the higher mortality of Covid-19 in Blacks. Indeed, it is conceivable that socioeconomic and biological factors may act in combination to foster Covid-19 disparities through the chronic downregulation of ACE2 expression leading to more severe sequelae of infection.
In summary, we demonstrate that ACE2 levels are decreased in Blacks providing the potential insights in biological underpinnings of Covid-19 racial disparities. This may offer new vistas in mitigating Covid-19 toll on vulnerable populations (i.e., potential druggable pathways) which needs to be implemented in conjunction with efforts targeting the socioeconomic and co-segregating risk factors (obesity, hypertension, smoking etc.) which are likely contributors to these disparities.