Early humoral defense under the radar: microvascular-epithelial cooperation at airways infection in asthma and health.

to the editor: I read with interest the review by Ramakrishnan et al. (1) discussing why asthma, against expectations, seems without increased risk for severe COVID-19 in several studies. Like currently acknowledged frontiers in innate defense (2, 3), the authors focus on cell mechanisms (1). This Letter discusses an additional possibility.

First it needs to be acknowledged that the well-reported possibility that persons with asthma are at reduced risk (1) is not generally valid (4). Significantly increased risk for developing severe COVID-19 has been demonstrated, especially for patients with severe uncontrolled asthma and a history of hospitalization due to exacerbations and systemic steroid treatment in recent years (4). The focus of the present discussion remains on airways defense in both health and asthma.

LACK OF ATTENTION TO PHYSIOLOGY OF ENDOTHELIAL-EPITHELIAL BARRIER FUNCTIONS IN VIVO

Physiological cooperation between the superficial airway microcirculation and the epithelial lining is rarely included in schemes depicting immune mechanisms. One consequence is a lack of attention to roles of biologically active, plasma-derived molecules in early mucosal defense. Yet, in environmentally challenged airways, perhaps particularly in asthmatic bronchi (5), swift microvascular-epithelial transmission of nonsieved plasma peptides/proteins contributes prompt antimicrobial opportunities (6, 7).

At the primary site of respiratory infections, the human conducting airways (8), nonsieved plasma-derived macromolecules are transmitted across an intact epithelial lining without impeding the normal airway barrier against cenvironmental challenges (5–7). This direction-dependent asymmetry is a crucial, but overlooked (7), operational feature of the airway pseudostratified epithelial lining. Bedside and in vivo data suggest that exudation of plasma-derived peptides/proteins at the airway level has capacity to reduce progression of COVID-19 into severe pulmonary stages and beyond.

DISTINCT FROM THE PULMONARY CIRCULATION, THE AIRWAYS MICROCIRCULATION ACCOUNTS FOR HUMORAL INNATE IMMUNE RESPONSES

As a significant pathogenic component of severe COVID-19, the pulmonary circulation leaks fluid and protein across injured endothelial and alveolar linings (3, 6). This acute respiratory distress syndrome (ARDS)-like lung plasma leak condition is entirely distinct from the plasma exudation responses that operate at airways defense (5–7, 9). Hence, extravasation of plasma from the systemic subepithelial microcirculation of human conducting airways must not be confused with poorly reversible, pathogenic leak of plasma from the low-pressure pulmonary microcirculation.

In human nasal and tracheobronchial airways, a ubiquitous subepithelial distribution of postcapillary venules, carrying systemic oxygenized blood, offers opportunities for highly localized vascular permeability responses to mucosal challenges. Depending on type and level of environmental challenge, the venules will leak plasma and the overlying epithelium promptly transmits extravasated plasma, for brief, biphasic, or quite prolonged but still well-controlled lengths of time (5, 6). Experimental data indicate that the extravasated plasma itself, by slightly increasing the hydrostatic pressure that impacts on basolateral epithelial aspects, opens valve-like paracellular pathways for its own transmission across the pseudostratified epithelial lining (5, 7). Actively regulated, venular endothelial gap formation thus caters for localized occurrence and tightly controlled duration of plasma exudation responses in conducting airways (6).

MICROVASCULAR-EPITHELIAL EXUDATION OF NONSIEVED PLASMA-DERIVED PEPTIDES/PROTEINS

Mimicked in guinea pig trachea, human nasal and tracheobronchial airways harbor a profuse superficial microcirculation positioned just beneath a pseudostratified epithelium (5, 10). Environmental challenge of these airways with autacoids, allergens, and viruses has revealed these basic mucosal plasma exudation features: prompt, dose-dependent, glucocorticoid-insensitive, reversible, repeatable exudation of nonsieved plasma-derived peptides/proteins. Nonsieved plasma exudation means joint mucosal surface appearance of well-known circulating defense molecules belonging to coagulation, complement, natural antibody, and more systems (11), including cathelicidins (12).

Exuded plasma, rather than popular local cell production, is a source of many important mucosal surface proteins in asthma including complement and IgM (5, 6). Although the humoral source was not acknowledged, it is of interest that roles of IgM and complement in resisting SARS-CoV-2 infection have been briefly discussed (reviewed in Ref. 6). As with antiviral effects of interferon-β, the authors have emphasized that antiviral actions of complement would be important in the early phase of respiratory infection whereas inflammation may be worsened by its involvement in late-stage COVID-19 (6). At advanced hyperinflammatory stages of COVID-19, with leakage of plasma from the pulmonary circulation, additional exudation of plasma in the conducting airways would be an undesirable response. Changing from contributing to early viral resistance, the airways plasma exudation response may then become a component of the host’s poor tolerance to the viral infection. It is of note that exaggerated plasma exudation in severe noninfectious asthma may contribute in several ways to severity of this disease (5).

A CRUCIAL ASYMMETRY OF THE AIRWAY PSEUDOSTRATIFIED EPITHELIAL BARRIER

Swift epithelial transmission of bulk plasma molecules, in human and guinea pig airways, occurs without injury and without compromising the epithelial absorption barrier (5–7, 11, 13–16). Consistent with such dramatic, unidirectional penetrability of the pseudostratified epithelial lining, operating when approached from beneath, lymphatic protein transport may not increase (10) and edema has not been induced at mucosal challenge-induced plasma exudation (5). An increase in hydrostatic pressure of less than 5 cmH2O, impacting basolateral aspects of an intact in situ pseudostratified airway epithelium, reversibly and repeatably creates such plasma exudation-like, para-epithelial movement of macromolecules to the mucosal surface, and this occurs without reducing the normal epithelial barrier function (5, 13, 15). Such asymmetry of the epithelial barrier, operating in human conducting airways and guinea pig trachea, is crucial for the defense potential of environmental challenge-induced plasma exudation responses (7).

It seems unfortunate that established ideas centering around “leaky epithelial barriers” (based on bench data without consideration of actual bedside data) have totally eclipsed the physiologic barrier asymmetry of the pseudostratified epithelium that lines our conducting airways (7).

LOCAL PLASMA EXUDATION ASSOCIATES WITH HUMAN AIRWAYS INFECTION UNTIL RESOLUTION

Distinct from size-dependent passive diffusion in human healthy airways (6), nonsieved plasma proteins move to airway mucosal surfaces at infection (6), in asthma already at baseline without infection (5, 6, 9). From onset to resolution, rhinovirus 16 and coronavirus229E inoculation-induced cold has been associated with human nasal exudation of plasma macromolecules (determined as albumin and fibrinogen) (6, 17–19). The cold may then be followed by a period of plasma exudation hyperresponsiveness suggestive of increased humoral defense alert (6, 17). Potentially reflecting such hyperresponsiveness, influenza AB caused very large sputum levels of fibrinogen in asthma (18). Based on available data it seems likely that influenza and common cold, localized to nasal and tracheobronchial airways, associate with plasma exudation although the intensity of the latter varies between individuals (6).

The baseline plasma exudation in asthmatic airways, although greater than in health, may only be a marginal contribution to the volume of airway surface liquids. Its highly localized occurrence also would not perfectly coincide with sites where inhaled viruses are being deposited. However, the exudative responsiveness to viral infection could be increased in the already diseased airways (6, 18) and thus improve defense. Judging from experiences with common cold viruses, airways plasma exudation responses start only when symptomatic infection is evident (6, 17). Thus, despite belonging to early defense responses, airways plasma exudation may not reduce initial take rate of infections (17).

The innate humoral defense of conducting airways may be described as a viral resistance mechanism that is proresolution, operating to limit the progress of viral infection. This is an uncharted defense mechanisms territory. Hence, research is warranted to determine to what extent the plasma exudation response occurs and to what extent it may combat airways infections including those caused by SARS-CoV-2.

HAS ASTHMA BEEN IDENTIFIED AS A RISK FACTOR FOR SEVERE DISEASE IN PREVIOUS VIRAL PANDEMICS?

In 2009, a challenging pandemic of H1N1 influenza occurred causing severe disease in children and adults. As evidenced by numerous publications, the role of asthma for severity of the H1N1 infection received considerable interest. Initially, asthma was identified as a risk factor associated with hospitalization in the 2009 pandemic. However, a thorough analysis of US hospitalized patients demonstrated that severity of the disease in terms of developing pneumonia, needing mechanical ventilation, and death was less in patients with asthma compared with patients without asthma (20).

A recent, comprehensive review by Veerapandian et al. (21) is of interest for the present discussion. The authors cite more than 10 reports, which jointly suggested that asthma was associated with reduced severity of the H1N1 disease. They also discuss in some detail why asthma may not predict a severe outcome as had been argued: there were uncertainties regarding the diagnosis of asthma, especially in young patients, exacerbation of asthma was not distinguished from severity of infection, and associated risk factors, notably obesity, did not receive due consideration.

Veerapandian et al. (21) combine their critical approach with currently acknowledged frontier accounts of asthma mechanisms presenting three colorful and detailed illustrations of airway mucosal response in asthma to allergen and influenza, separately and together. Typically, the subepithelial microcirculation is missing in these figures. Also typically, in their text on asthma pathophysiology, the authors mention leak of plasma merely in relation to edema and disturbed mucociliary transport (21). [I take some responsibility for disseminating such traditional and limited ideas regarding plasma exudation in asthma in the 1980s (22)]. As expected, and adhering to the accepted discourse regarding defense and COVID-19 (1–3), attempts by Veerapandian et al. (21) at explaining reduced infection severity in asthma exclusively deal with cellular mechanisms of defense and repair. To summarize, experiences from the 2009 H1N1 pandemic may support rather than be at variance with the present discussion regarding the possibility of a role of early humoral defense in airways antimicrobial defense in general and in asthma.

DEFENSE, BARRIER, AND REPAIR AT SITES OF PATCHY EPITHELIAL SHEDDING

Epithelial shedding is a feature of asthma. Add to this the established notion of “defect epithelial repair” in this disease (5, 7, 23) and one would expect a dramatic loss of the mucosal barrier. However, this has not happened.

Contrary to the firmly rooted research front idea of defect barrier (7, 23), well-controlled in vivo studies in asthma and allergic rhinitis demonstrate that the human airway mucosal barrier against environmental challenges is maintained in these diseases, or even operationally tighter compared with healthy airways (5, 7). Failed expectations may explain why some of the best-controlled studies, actually demonstrating reduced absorption permeability in asthma, are now available only as abstract (5).

Acknowledged leaders in the fields of airway epithelial barrier and repair in asthma seem to either overlook and refuse to discuss the dichotomy between their bench data-based notions and physiological in vivo demonstrations in patients, or may not fully appreciate the scientific impact of well-controlled human in vivo data in the area of interest (7, 23).

Plasma exudation may actually be involved in maintaining a physical and biological defense barrier at sites of epithelial loss: Asthma-like denudation (= uninjured basement membrane and no bleeding), induced in guinea pig trachea, allowed physiologically well-controlled demonstration of epithelial regeneration milieu/events in vivo (24): protecting denudation spots, restrictedly local plasma exudation promptly and sustainedly produced a biologically powerful defense and repair milieu held in place by a fibrin-fibronectin gel barrier; in this dynamic in vivo setting, all neighboring epithelial phenotypes dedifferentiated into rapidly migrating, tethered regeneration cells; when a new cell barrier of undifferentiated repair epithelium covered the denuded patch, plasma exudation ceased and the fibrin-fibronectin gel was shed (5, 6, 9).

Airways infection with SARS-CoV-2 may cause epithelial injury/shedding (3, 6). Perhaps this is, especially likely in asthmatic bronchi with an already fragile epithelium. Importantly, epithelial loss in asthma seems exceedingly patchy (5, 6, 23). This feature, together with plasma-derived both provisional barrier and milieu, the latter designed for optimal speed of epithelial regeneration as well as for biological defense capacity, may in part explain why asthmatic bronchi have exhibited neither increased absorption of inhaled molecules (5, 7) nor a consistently increased risk for severe COVID-19 (1).

EXAGGERATED EPITHELIAL REGENERATION IN PATHOGENESIS OF ASTHMA

Of note, the speedy epithelial regeneration in vivo came with a catch. Beyond plasma exudation, epithelial regeneration alone evoked eosinophilic-neutrophilic inflammation, hypersecretion as well as epithelial basement membrane and fibrocyte/smooth muscle remodeling (5, 24). These effects agree with known pathology of asthma and data suggest that epithelial shedding regeneration is a contributing factor both for inception of asthma and for worsening of established disease. Sputum analyses have thus demonstrated numerous clusters of shed epithelial cells both in established asthma and, as an independent factor, predicting development of asthma, in wheezing infants (5).

Taken together, experimental in vivo data on pathogenic effects of epithelial regeneration alone and human bedside observations on maintained barrier despite occurrence of epithelial loss (5) suggest the possibility that exaggerated epithelial regeneration, rather than currently established notions of defect epithelial repair and defect epithelial barrier (7, 23), is involved in pathogenesis of asthma. If increased epithelial regeneration associates with SARS-CoV-2 infection, this would be a factor that worsens asthma.

WHY “UNDER THE RADAR?”

A variety of factors may explain why defense capacities of airways plasma exudation have been overlooked (5–7). Perhaps the popular idea of “leaky epithelial barrier” (7) is a major factor since it disregards the asymmetrical perviousness of the pseudostratified epithelial lining of our airways (5, 7). However, without ranking, several more circumstances may be involved:

Unshakable view that airways plasma exudation reflects/causes injury = pathogenic.

Limited understanding of distinctions between bronchial and pulmonary circulations.

Unfortunate confusion with pulmonary plasma leak in severe ARDS-COVID-19.

Unfounded notions of increased airway absorption permeability in exudative asthma.

Misconceptions regarding direction-selective penetrability of human airway epithelial lining.

Since early 1990s: classical clinical research considered obsolete for medical discoveries.

Since early 1990s: low priority to exploratory research on airway-specific in vivo physiology.

Airway plasma exudation not present in popular models (cell cultures and mice).

Plasma exudation = too complex molecular opportunities for “precision immunology.”

Physiological bedside in vivo data may reduce or ruin favored bench-based ideas.

CONCLUSIONS

Human in vivo data indicate ample but variable opportunities for locally exuded plasma protein systems to effect defense on infected airway surfaces, apparently more so in asthma than in health. It is speculated that this humoral response contributes to containing SARS-CoV-2 infections by combatting COVID-19 at the primary site of infection, the conducting airways. Research is warranted to move the early humoral airways response to environmental challenges from intriguing, well-controlled opportunity toward well-defined operational importance in antimicrobial defense.

DISCLOSURES

No conflicts of interest, financial or otherwise, are declared by the author.

AUTHOR CONTRIBUTIONS

C.P. drafted manuscript; edited and revised manuscript; and approved final version of manuscript.