Severe Asthma PBMCs Show Lower Cytokine Responses to RV-A16

Key Takeaways

• Ex vivo peripheral blood mononuclear cell (PBMC) cultures from adults with severe asthma generally showed lower stimulated cytokine release than cultures from healthy participants.
• CCL2 was the exception to the generally lower RV-A16-induced responses in severe asthma.
• In severe asthma, cytokine responses had both positive and inverse associations with asthma biomarkers, depending on the cytokine.
• In a smaller healthy-donor experiment, replication-competent RV-A16 elicited stronger cytokine responses than ultraviolet-inactivated virus, except for CCL2.

Rhinovirus-associated exacerbations are a concern in severe asthma, but how circulating immune cells respond to viral exposure remains unclear. Measuring cytokines after laboratory viral challenge can examine that cellular response without establishing whether a patient clears infection or experiences an exacerbation.

Investigators enrolled 50 adults with documented severe asthma and 20 healthy, never-smoking participants for the RV-A16 PBMC comparison at Ruhr University Bochum in Germany. Severe asthma met National Institutes of Health (NIH) and Global Initiative for Asthma (GINA) level 5 criteria; the groups were matched by age and sex. Neither group reported infection in the preceding two weeks. Investigators exposed peripheral blood mononuclear cells (PBMCs) ex vivo to rhinovirus A16 (RV-A16) at multiplicities of infection (MOI) of 0.1 and 1, alongside uninfected cultures. Enzyme-linked immunosorbent assay (ELISA) measured supernatant cytokines at 24 hours and seven days. Virus-induced responses and biomarker associations used corresponding uninfected-culture values as the baseline. A smaller healthy-donor experiment compared active with ultraviolet (UV)-inactivated RV-A16.

Induced responses of seven of eight assayed proteins were generally lower in severe-asthma cultures than in healthy cultures, depending on viral dose and incubation time; C-C motif chemokine ligand 2 (CCL2) was the exception. The seven were interferon-alpha (IFNα), interferon-gamma (IFNγ), C-C motif chemokine ligand 5 (CCL5), tumor necrosis factor-alpha (TNFα), interleukin-1 beta (IL-1β), interleukin-6 (IL-6), and interleukin-8 (IL-8). Healthy cells showed induction of all eight proteins, whereas severe-asthma cells showed induction of all except IL-1β. At the higher dose after seven days, TNFα was suppressed in both groups without a significant between-group difference.

Among severe-asthma participants, IFNα, IL-6, and CCL5 responses were inversely associated with fractional exhaled nitric oxide (FeNO), while IL-1β was positively associated with FeNO. IFNγ was positively associated with blood eosinophils and serum interleukin-33 (IL-33); IFNα was positively associated with total serum immunoglobulin E (IgE). No examined response correlated with periostin or thymic stromal lymphopoietin (TSLP). FeNO data were available for 38 participants at 24 hours and 37 at seven days, versus 48 and 47, respectively, for eosinophils, IgE, and IL-33. In healthy-donor cultures, replication-competent RV-A16 yielded viral copies above the inoculum and stronger cytokine responses than UV-inactivated virus, apart from CCL2. The experiment did not test viral replication in severe-asthma cells.

Mixed-cell PBMC cultures cannot identify the cell types or interactions responsible for the observed differences. Cell composition differed between groups, and 11 severe-asthma participants received maintenance oral corticosteroids; neither observation establishes an explanation for the response pattern. Cultures ran at 37°C, unlike the cooler nasal epithelial infection conditions discussed by the investigators, and the principal challenge used RV-A16 alone. Investigators described most biomarker associations as weak. Their proposed connections to viral clearance, exacerbations, and possible drug targets remain hypotheses: patient infection outcomes and treatment effects were not measured.

Investigators considered the differing biomarker associations potentially relevant to asthma phenotypes. The generally lower RV-A16-stimulated responses in severe-asthma circulating cells, with CCL2 as the exception, do not establish a phenotype-specific mechanism or an effect on infection defense.

Clinician Questions

Were patients receiving asthma biologics included in the RV-A16 PBMC comparison?

Adults with severe asthma who supplied PBMCs for the RV-A16 challenge had either never used a monoclonal antibody or had received no monoclonal antibody treatment for at least three months before sampling. The comparison does not establish responses during active biologic treatment.

How did unstimulated PBMC cytokine release differ in severe asthma and healthy controls?

Without RV-A16 exposure, PBMCs from adults with severe asthma released less TNFα, IL-6, IL-8, and IFNγ at one or both assessment times than healthy-control PBMCs, whereas IL-1β was higher. IFNα, CCL2, and CCL5 showed no significant baseline group difference; these are unstimulated findings, distinct from baseline-referenced induced responses.

Why did CCL2 respond to UV-inactivated rhinovirus A16 in healthy-donor PBMC cultures?

UV-inactivated RV-A16 also triggered CCL2 secretion in healthy-donor PBMC cultures. Investigators proposed recognition of incoming viral particles or viral RNA after endocytosis as possible explanations, but did not establish either mechanism.

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