MicroRNA-155 modulation by renin-angiotensin system inhibitors may underlie their enigmatic role in COVID-19

Abstract

Severe acute respiratory coronavirus-2 (SARS-CoV-2) infection course differs between the young and healthy and the elderly with co-morbidities. In the latter a potentially lethal coronavirus disease 2019 (COVID-19) cytokine storm has been described with an unrestrained renin-angiotensin (Ang) system (RAS). RAS inhibitors [Ang converting enzyme inhibitors and Ang II type 1 receptor (AT1R) blockers] while appearing appropriate in COVID-19, display enigmatic effects ranging from protection to harm. MicroRNA-155 (miR-155)-induced translational repression of key cardiovascular (CV) genes (i.e., AT1R) restrains SARS-CoV-2-engendered RAS hyperactivity to tolerable and SARS-CoV-2-protective CV phenotypes supporting a protective erythropoietin (EPO) evolutionary landscape. MiR-155’s disrupted repression of the AT1R 1166C-allele associates with adverse CV and COVID-19 outcomes, confirming its decisive role in RAS modulation. RAS inhibition disrupts this miR-155-EPO network by further lowering EPO and miR-155 in COVID-19 with co-morbidities, thereby allowing unimpeded RAS hyperactivity to progress precariously. Current pharmacological interventions in COVID-19 employing RAS inhibition should consider these complex but potentially detrimental miR-155/EPO-related effects.

Key Words: Angiotensin converting enzyme inhibitors; Angiotensin II type 1 receptor blocker; COVID-19; MicroRNA; Mineralocorticosteroid receptor antagonists; MicroRNA-155; Renin-angiotensin system inhibitors; SARS-CoV-2; Sodium-glucose transporter 2

Core Tip: The role of renin-angiotensin system (RAS) inhibition in coronavirus disease 2019 (COVID-19) remains enigmatic. Novel insights involving interference of microRNA-155 (miR-155) and erythropoietin (EPO) levels support a detrimental role by RAS inhibitors in the elderly and in co-morbidities. Current pharmacological interventions in COVID-19 employing RAS inhibition should consider these complex but potentially detrimental miR-155 effects. Future research should address the knowledge gap on how RAS inhibition affects miR-155 while therapeutic strategies should focus on pharmacological interventions that restore EPO and miR-155 levels and functionality. The use of tissue specific miRNA analogs could become a reality by then.

TO THE EDITOR

Despite the overwhelming involvement of the renin-angiotensin (Ang) system (RAS) in severe acute respiratory coronavirus-2 (SARS-CoV-2) infection and the coronavirus disease 2019 (COVID-19) pathophysiology, there is an ongoing controversy regarding the use of RAS inhibitors (RASi) [Ang converting enzyme (ACE) inhibitors (ACEi) and Ang II type 1 receptor (AT1R) blockers (ARB)] in COVID-19[1]. The proposed effects of pharmacological RAS inhibition in COVID-19 have ranged from protection to harm[2]. Recent studies indicate a lack of benefit and advise against RASi initiation in critically ill patients[3]. Other researchers observed that discontinuing RAS inhibition in COVID-19 patients, although not significantly affecting maximum disease severity, could result in a quicker and more complete recovery[4]. Additional reports indicate that 2^nd-generation ARB may carry a higher risk of hypotension compared to 1^st-generation ARB[5]. Intriguingly, the pathophysiology behind these puzzling findings remains unknown.

In this letter, we will provide novel insights that involve interference of microRNA-155 (miR-155) levels by pharmacological RAS inhibition, potentially shedding light on their enigmatic role in COVID-19.

RAS MODULATION OF HOST IMMUNE AND HEMODYNAMIC RESPONSE

Upon host invasion, the mandatory SARS-CoV-2 spike protein (S) interaction with ACE2 inevitably links the RAS to the host response (Figure 1)[6]. ACE2 is thereby downregulated in that way, leading to a hyperactive host RAS with increased Ang II and reduced Ang1-7 in vascular endothelial cells impairing endothelial nitric oxide (NO) synthase (eNOS) activity and NO generation and bioavailability[6]. How the infected individual is equipped to handle this RAS hyperactivity decides the fate of the infection[7]. In newborns, healthy children, and teenagers, genetically imprinted, physiological RAS hyperactive states, engendered through lower nasal ACE2 expression, higher, age-related ACE activity, and certain RAS single nucleotide polymorphisms, are well documented, supporting the notion that elevated Ang II activity mediates desirable immunological benefits when its potentially detrimental circulatory effects are constrained[7]. In the young, the above-mentioned physiological RAS hyperactive states, are well tolerated, even when further enhanced by SARS-CoV-2, and appear protective as they seem to render the infection mild or asymptomatic in those age groups[6,7]. In contrast, unimpeded RAS stimulation in the elderly, the genetically susceptible, and those with comorbidities [type 2 diabetes mellitus (T2DM), sarcopenia, obesity, smoking, aging, male gender, cardiovascular (CV) and renal disease, cancer, or pharmacological interventions] results in severe and extensive endotheliitis, the hallmark of COVID-19 cytokine storm[7].

Figure 1 Schematic interplay between microRNA-155/renin-angiotensin system/erythropoietin axis and influence of pharmacological renin-angiotensin system inhibition in severe acute respiratory coronavirus-2 infection. Hypoxia of any etiology elicits a hyperactive renin-angiotensin (Ang) system (RAS) and increases erythropoietin (EPO) production directly and via Ang II. MicroRNA-155 (miR-155) tames RAS hyperactivity via Ang II type 1 receptor (AT1R) repression. All these actions, pivotally regulated by miR-155 availability, collectively promote a protective cardiovascular phenotype and EPO evolutionary landscape that mediates severe acute respiratory coronavirus-2 resistance. Ang converting enzyme inhibitors and AT1R blockers block Ang II and the AT1R, respectively, and thereby impede EPO formation. Both also reduce miR-155 levels. ACE: Angiotensin converting enzyme; ACEi: Angiotensin converting enzyme inhibitors; ARB: Angiotensin II type 1 receptor blockers; AT1R: Angiotensin II type 1 receptor; EPO: Erythropoietin; MiR-155: MicroRNA-155; SARS-CoV-2: Severe acute respiratory coronavirus-2.

PIVOTAL EFFECTS OF ERYTHROPOIETIN AND MICRORNA-155 IN SARS-CoV-2 INFECTION

Why SARS-CoV-2 infection outcomes diverge depending on age and comorbidities is unclear. Our detailed pathophysiological analysis of the SARS-CoV-2 infection and COVID-19 progression highlights the prominent roles of erythropoietin (EPO) and miR-155 as crucial and pivotal regulators at the critical juncture between youth and old age/comorbidities[6-8].

Substantial evidence corroborates the observation that in the young and healthy, without comorbidities and/or pharmacological RAS interventions, physiological and SARS-CoV-2-ACE2-binding-engendered RAS hyperactivity elevates Ang II and aldosterone[6]. In addition to their main hemodynamic roles, these hormones are master regulators of erythropoiesis, mediating heightened anemia-independent EPO secretion (Figure 1)[7,9-12]. Age-dependent and anemia-independent EPO elevation, peaking prior to the age of 5 and declining during a child’s development, forms an ancestral early life protective EPO evolutionary landscape. Disproportionately elevated Ang II and EPO levels have been recurrently observed in young children with malaria and are thought to grant the host a fitness advantage against cerebral and other complications while imposing constraints on pathogen adaptation and invasion[13]. Viral infections like SARS-CoV-2 trigger the innate immune system's intracellular pattern recognition receptors (PRR) to sense pathogen-associated molecular patterns and danger-associated molecular patterns. The NLR family pyrin domain containing 3 protein (NLRP3), a well-studied PRR, senses these patterns and triggers the formation of inflammasomes[7]. Dysregulated activation of NLRP3 inflammasomes during acute SARS-CoV-2 infection is linked to severe COVID-19, playing a key role in the massive inflammation seen[7,14,15]. NLRP3 inflammasome activation can be effectively contained through increased EPO engendered via the above-mentioned RAS hyperactivity. Furthermore, EPO interaction with eNOS will result in increased NO generation and bioavailability subsequently inhibiting the imminent endotheliitis, SARS-CoV-2 replication, and its cell entry, ultimately protecting against SARS-CoV-2 infection[7]. In addition, EPO may modulate host systemic immunological response towards increased tolerance, preventing autoimmune pathology as well as contributing to a more apposite inflammatory response to pathogens, thereby lowering the infection burden particularly in young children[7].

As multifunctional miR-155 targets and represses over 241 genes, it regulates crucial aspects of a host’s antiviral, immunological, and CV functions (Table 1)[16]. MiR-155 regulation of genes contributing to atherosclerosis, cardiomyopathy, and hypertension underlies its prominent involvement in the pathogenesis of CV disease (CVD)[16]. By repressing AT1R gene (AGTR1) (the gene coding for AT1R) and downregulating AT1R membrane expression, miR-155 controls the ACE1/Ang II/AT1R axis, thereby restraining Ang II pro-inflammatory effects, and limiting SARS-CoV-2-AT1R-dependent endocytosis (Table 1). Moreover, eNOS dissociates from AT1R, increasing NO synthesis and bioavailability, consequently blocking viral replication and cell entry[6]. Elevated plasma Ang II will now interact with Ang II type 2 receptor and mediate EPO-eNOS/NO vasculoprotective signaling and appropriate NLRP3 inflammasome regulation[6,7,17]. Repression of Arginase2 and E26 transformation-specific sequence-1 improves eNOS substrate (L-arginine) availability and negates Ang II-induced endothelial and vascular inflammation (Table 1)[6]. MiR-155 thus appears to purposefully counteract and tame this SARS-CoV-2-triggered RAS hyperactivity, thereby inducing a cardioprotective phenotype, while simultaneously forming constraints for SARS-CoV-2 to invade, adapt, and replicate[6]. Lack of miR-155 repressive action as seen with the 1166C-allele of the AT1R is associated with adverse CV and SARS-CoV-2 outcomes[6]. In addition, miR-155 mediated BTB and CNC homology 1, basic leucine zipper transcription factor 1 and suppressor of cytokine signaling 1 repressions robustly induce antiviral interferons and cytoprotection, and reduce inflammatory burden (Table 1)[6].

Table 1 Direct gene targets of microRNA-155 relevant to renin-angiotensin system and immune system in severe acute respiratory coronavirus-2 infection.

Gene symbol	Full gene name	Action
AGTR1	Angiotensin II type 1 receptor gene	AGTR1 gene repression downregulates its translation, thereby lowering AT1R membrane expression and downstream signalling, like endogenous AT1R blockade redirecting Ang II towards its alternative receptor Ang II type 2 receptor
ARG2	Arginase2	Repressed translation allows increased L-arginine availability to generate nitric oxide supporting immune cell function and beneficially regulating inflammatory responses in the lung
BACH1	BTB and CNC homology 1, basic leucine zipper transcription factor 1	Translational repression of BACH1 leads to potent anti-inflammatory, cytoprotective, antioxidant programs through heme oxygenase-1
Ets-1	E26 transformation-specific sequence-1	Ets-1 translational repression prevents Ang II/AT1R-mediated gene upregulation involved in inflammation, fibrosis, vascular remodeling, and cardiac hypertrophy
CACNA1C(Cav1.2)	L-type calcium channel	CACNA1C forms part of the LTCC that mediates reactive oxygen species production and calcium influx in vascular smooth muscle cells, causing vasoconstriction and oxidative stress, both important components of vascular aging. These attributes are contained by miR-155-induced translational repression
SOCS1	Suppressor of cytokine signaling 1	SOCS1 negatively regulates type I IFN signaling, thus, its translational repression enhances type I IFN-mediated antiviral response. Erythropoietin’s protective effects in ischemic injury are mediated through the Janus kinase 2/Y343/signal transducers and activators of transcription 5 pathway, enhanced via miR-155-induced SOCS1 repression

AGTR1: Angiotensin II type 1 receptor gene; Ang: Angiotensin; ARG2: Arginase 2 gene; AT1R: Angiotensin II type 1 receptor; BACH1: BTB and CNC homology 1, basic leucine zipper transcription factor 1 gene; Ets-1: E26 transformation-specific sequence-1 gene; IFN: Interferon; LTCC: L-type calcium channel; MiR-155: MicroRNA-155; SOCS1: Suppressor of cytokine signaling 1 gene.

The above combined miR-155 effects lend robust support to the notion that miR-155-RAS-EPO network interactions epitomise a generic and evolutionarily conserved, protective antiviral defence programme engendered through the linkage of the endocrine (RAS and EPO) and immune systems[6,8].

RAS INHIBITION DISTURBS THE MIR-155-RAS-EPO NETWORK

It is well known for almost 3 decades that Ang II increases EPO levels de novo, in a dose-dependent manner, and independent of physiological stimuli for EPO production such as haemorrhage or hypoxia[18,19]. Ang II’s EPO effects are completely abolished by RAS blockade (ACEi/ARB) through inhibition of Ang II formation and AT1R blockade, respectively, potentially resulting in lower hematocrit and/or anemia[19]. In healthy or near healthy states, RAS’s effects on EPO may be unnoticeable and only unveiled when a failing bone marrow necessitates additional stimulatory inputs to sustain erythropoiesis, e.g., immunosuppression and renal or congestive heart failure[19]. RAS blockade (ACEi/ARB) may, in pathological states with low EPO, further impair the protective EPO evolutionary landscape and its tissue-protective effects[6,7].

On the other hand, the effect of RAS blockade (ACEi/ARB) on miR-155 is less well known and warrants further research. Only two publications could be retrieved and both reported significantly reduced miR-155 levels with ACEi/ARB use[20,21]. In situations where miR-155 response is compromised (AGTR1 1166A/C polymorphism and/or its levels are impaired, as in old age, male gender, smoking, and in comorbidities such as obesity, sarcopenia, hypertension, T2DM, CVD, renal disease, or critically ill COVID-19 patients), RAS inhibition may further depress miR-155 levels and interfere with anticipated beneficial antiviral, anti-inflammatory, and immunoregulatory functions, leading to a severe COVID-19 course (Figure 1)[6,20-22]. In addition, RAS inhibition may also not be as multifaceted or as efficient as miR-155-induced AT1R downregulation to mitigate detrimental Ang II effects or a cytokine storm[6]. In COVID-19, initiating pharmacological RAS inhibition, especially with 2^nd-generation ARB, could lead to hypotension and vasoplegia due to potentiation of an already established miR-155-induced AT1R downregulation and impaired Ang II vascular reactivity[5,23]. Finally, recent evidence suggests that AT1R’s effects on CD11c-expressing myeloid cells protect against hypertension, in part through attenuation in T cell recruitment, accumulation, and activation in renal tissue, resulting in blunted inflammatory cytokine production and protection from sodium retention[24]. Blanket RAS blockade potentially deprives COVID-19 patients of the above beneficial immunoregulatory AT1R effects while miR-155-induced AT1R downregulation might be more subtle and better fine-tuned.

CONCLUSION

MiR-155 is the most important microRNA in CV pathology and a crucial regulator of the immune system’s mounting network interactions that represent an evolutionarily conserved, generic antiviral defence programme connecting the endocrine (RAS/Ang II and EPO) and immune systems[6]. In the young and healthy and in the absence of comorbidities, elevated miR-155 levels in SARS-CoV-2 infection are anticipatory and purposeful and, through translational repression of key genes (Table 1), render physiological and infectious RAS hyperactive states CV protection[6]. An adequate supply of Ang II is thereby ensured to sustain elevated EPO production to maintain a protective evolutionary landscape[6,7]. When old age and comorbidities disturb miR-155 levels and responsiveness, initiating or maintaining RAS inhibition may abolish miR-155/EPO network’s control of RAS hyperactivity, resulting in a treacherous COVID-19 course[6].

In the clinical management of COVID-19, substituting RAS inhibition in favor of mineralocorticosteroid receptor (MR) antagonists (spironolactone, eplerenone, and finerenone), may prove effective especially in the elderly, as MR inhibition reverses the significantly depressed basal serum miR-155 levels in the senescent vasculature and subsequently represses the L-type calcium channel (LTCC) subunit Cav1.2 (Table 1)[6,16,25-32]. Verapamil, an LTCC blocker, presents another option, especially in T2DM[33]. Finally, employing sodium-glucose transporter 2 (SGLT-2) inhibitors in T2DM may help raise EPO levels in susceptible patients[19]. SGLT-2 inhibition appeared protective against SARS-CoV-2 infection and against COVID-19 hospitalization with lower risk of adverse CV and kidney outcomes[34-36].

Current pharmacological interventions in COVID-19 employing RAS inhibition should consider these complex but potentially detrimental miRNA-related effects. Future research should address the knowledge gap on how RAS inhibition affects miR-155 levels while therapeutic strategies should focus on pharmacological interventions that restore EPO and miR-155 levels. The use of tissue specific miRNA analogs could become a reality by then.