SARS-CoV-2 Omicron (B.1.1.529) infection in rhesus macaques, hamsters, and BALB/c mice with severe lung histopathological damage.

Since the first SARS-CoV-2 outbreak in late 2019, the SARS-CoV-2 genome has harbored multiple mutations, especially spike protein mutations. The currently fast-spreading Omicron variant that manifests without symptoms or with upper respiratory diseases has been recognized as a serious global public health problem. However, its pathological mechanism is largely unknown. In this work, rhesus macaques, hamsters, and BALB/C mice were employed as animal models to explore the pathogenesis of Omicron (B.1.1.529). Notably, Omicron (B.1.1.529) infected the nasal turbinates, tracheae, bronchi, and lungs of hamsters and BALB/C mice with higher viral loads than in those of rhesus macaques. Severe histopathological damage and inflammatory responses were observed in the lungs of Omicron (B.1.1.529)-infected animals. In addition, viral replication was found in multiple extrapulmonary organs. Results indicated that hamsters and BALB/c mice are potential animal models for studies on the development of drugs/vaccines and therapies for Omicron (B.1.1.529).

Ultrahigh-Resolution Photon-Counting Detector CT of the Lungs: Association of Reconstruction Kernel and Slice Thickness With Image Quality.

BACKGROUND. Prior work has shown improved image quality for photon-counting detector (PCD) CT of the lungs compared with energy-integrating detector CT. A paucity of the literature has compared PCD CT of the lungs using different reconstruction parameters. OBJECTIVE. The purpose of this study is to the compare the image quality of ultra-high-resolution (UHR) PCD CT image sets of the lungs that were reconstructed using different kernels and slice thicknesses. METHODS. This retrospective study included 29 patients (17 women and 12 men; median age, 56 years) who underwent noncontrast chest CT from February 15, 2022, to March 15, 2022, by use of a commercially available PCD CT scanner. All acquisitions used UHR mode (1024 × 1024 matrix). Nine image sets were reconstructed for all combinations of three sharp kernels (BI56, BI60, and BI64) and three slice thicknesses (0.2, 0.4, and 1.0 mm). Three radiologists independently reviewed reconstructions for measures of visualization of pulmonary anatomic structures and pathologies; reader assessments were pooled. Reconstructions were compared with the clinical reference reconstruction (obtained using the BI64 kernel and a 1.0-mm slice thickness [BI641.0-mm]). RESULTS. The median difference in the number of bronchial divisions identified versus the clinical reference reconstruction was higher for reconstructions with BI640.4-mm (0.5), BI600.4-mm (0.3), BI640.2-mm (0.5), and BI600.2-mm (0.2) (all p < .05). The median bronchial wall sharpness versus the clinical reference reconstruction was higher for reconstructions with BI640.4-mm (0.3) and BI640.2-mm (0.3) and was lower for BI561.0-mm (-0.7) and BI560.4-mm (-0.3) (all p < .05). Median pulmonary fissure sharpness versus the clinical reference reconstruction was higher for reconstructions with BI640.4-mm (0.3), BI600.4-mm (0.3), BI560.4-mm (0.5), BI640.2-mm (0.5), BI600.2-mm (0.5), and BI560.2-mm (0.3) (all p < .05). Median pulmonary vessel sharpness versus the clinical reference reconstruction was lower for reconstructions with BI561.0-mm (-0.3), BI600.4-mm (-0.3), BI560.4-mm (-0.7), BI640.2-mm (-0.7), BI600.2-mm (-0.7), and BI560.2-mm (-0.7). Median lung nodule conspicuity versus the clinical reference reconstruction was lower for reconstructions with BI561.0-mm (-0.3) and BI560.4-mm (-0.3) (both p < .05). Median conspicuity of all other pathologies versus the clinical reference reconstruction was lower for reconstructions with BI561.0 mm (-0.3), BI560.4-mm (-0.3), BI640.2-mm (-0.3), BI600.2-mm (-0.3), and BI560.2-mm (-0.3). Other comparisons among reconstructions were not significant (all p > .05). CONCLUSION. Only the reconstruction using BI640.4-mm yielded improved bronchial division identification and bronchial wall and pulmonary fissure sharpness without a loss in pulmonary vessel sharpness or conspicuity of nodules or other pathologies. CLINICAL IMPACT. The findings of this study may guide protocol optimization for UHR PCD CT of the lungs.

Evaluation of the Trachea and Bronchi in COVID-19 Patients Using the 3-dimensional Reconstruction Method.

OBJECTIVE: To evaluate changes in the trachea and bronchi of COVID-19 patients using the 3-dimensional reconstruction images obtained from chest CT (computed tomography) scans. STUDY DESIGN: An observational study. Place and Duration of the Study: Departments of Anatomy and Radiology, Faculty of Medicine, Lokman Hekim University, Ankara, Turkey, between March 2021 and January 2022. METHODOLOGY: There were 150 COVID-19 patients in the acute period and 150 individuals as the control group. The CT images were transferred to Mimics software, and a 3-dimensional reconstruction was performed. COVID-19 patients were grouped separately by gender, and their total lung severity score was classified as absent (Grade 0), mild (Grade 1), moderate (Grade 2), and severe (Grade 3). RESULTS: The cross-sectional area and diameter of the right upper lobar bronchus decreased as the grade increased (p<0.05 and p<0.001, respectively). The circumference of the right upper lobar bronchus and the cross-sectional area and circumference of the left lower lobar bronchus were found to be narrower in Grade 1-2-3 COVID-19 patients compared to those of the control group (p<0.01, p<0.05, and p<0.05, respectively). The cross-sectional area, circumference, and diameter of the middle lobar bronchus were found to be narrower in Grade 3 COVID-19 patients (p<0.05, p<0.05, and p<0.05, respectively). CONCLUSION: Although mostly independent of the grade increase, narrowing of the trachea and bronchi was observed in COVID-19 patients in the acute period. Further research is required with to reveal whether the narrowings are permanent. KEY WORDS: COVID-19, Trachea, Bronchus, 3-dimensional reconstruction.

Optimization of Primary Human Bronchial Epithelial 3D Cell Culture with Donor-Matched Fibroblasts and Comparison of Two Different Culture Media.

In vitro airway models are increasingly important for pathomechanistic analyses of respiratory diseases. Existing models are limited in their validity by their incomplete cellular complexity. We therefore aimed to generate a more complex and meaningful three-dimensional (3D) airway model. Primary human bronchial epithelial cells (hbEC) were propagated in airway epithelial cell growth (AECG) or PneumaCult ExPlus medium. Generating 3D models, hbEC were airlifted and cultured on a collagen matrix with donor-matched bronchial fibroblasts for 21 days comparing two media (AECG or PneumaCult ALI (PC ALI)). 3D models were characterized by histology and immunofluorescence staining. The epithelial barrier function was quantified by transepithelial electrical resistance (TEER) measurements. The presence and function of ciliated epithelium were determined by Western blot and microscopy with high-speed camera. In 2D cultures, an increased number of cytokeratin 14-positive hbEC was present with AECG medium. In 3D models, AECG medium accounted for high proliferation, resulting in hypertrophic epithelium and fluctuating TEER values. Models cultured with PC ALI medium developed a functional ciliated epithelium with a stable epithelial barrier. Here, we established a 3D model with high in vivo-in vitro correlation, which has the potential to close the translational gap for investigations of the human respiratory epithelium in pharmacological, infectiological, and inflammatory research.

A novel antiviral formulation containing caprylic acid inhibits SARS-CoV-2 infection of a human bronchial epithelial cell model.

A novel proprietary formulation, ViruSAL, has previously been demonstrated to inhibit diverse enveloped viral infections in vitro and in vivo. We evaluated the ability of ViruSAL to inhibit SARS-CoV-2 (severe acute respiratory syndrome coronavirus-2) infectivity, using physiologically relevant models of the human bronchial epithelium, to model early infection of the upper respiratory tract. ViruSAL potently inhibited SARS-CoV-2 infection of human bronchial epithelial cells cultured as an air-liquid interface (ALI) model, in a concentration- and time-dependent manner. Viral infection was completely inhibited when ViruSAL was added to bronchial airway models prior to infection. Importantly, ViruSAL also inhibited viral infection when added to ALI models post-infection. No evidence of cellular toxicity was detected in ViruSAL-treated cells at concentrations that completely abrogated viral infectivity. Moreover, intranasal instillation of ViruSAL to a rat model did not result in any toxicity or pathological changes. Together these findings highlight the potential for ViruSAL as a novel and potent antiviral for use within clinical and prophylactic settings.

The establishment of COPD organoids to study host-pathogen interaction reveals enhanced viral fitness of SARS-CoV-2 in bronchi.

Chronic obstructive pulmonary disease (COPD) is characterised by airflow limitation and infective exacerbations, however, in-vitro model systems for the study of host-pathogen interaction at the individual level are lacking. Here, we describe the establishment of nasopharyngeal and bronchial organoids from healthy individuals and COPD that recapitulate disease at the individual level. In contrast to healthy organoids, goblet cell hyperplasia and reduced ciliary beat frequency were observed in COPD organoids, hallmark features of the disease. Single-cell transcriptomics uncovered evidence for altered cellular differentiation trajectories in COPD organoids. SARS-CoV-2 infection of COPD organoids revealed more productive replication in bronchi, the key site of infection in severe COVID-19. Viral and bacterial exposure of organoids induced greater pro-inflammatory responses in COPD organoids. In summary, we present an organoid model that recapitulates the in vivo physiological lung microenvironment at the individual level and is amenable to the study of host-pathogen interaction and emerging infectious disease.

Smoking modulates different secretory subpopulations expressing SARS-CoV-2 entry genes in the nasal and bronchial airways.

SARS-CoV-2 infection and disease severity are influenced by viral entry (VE) gene expression patterns in the airway epithelium. The similarities and differences of VE gene expression (ACE2, TMPRSS2, and CTSL) across nasal and bronchial compartments have not been fully characterized using matched samples from large cohorts. Gene expression data from 793 nasal and 1673 bronchial brushes obtained from individuals participating in lung cancer screening or diagnostic workup revealed that smoking status (current versus former) was the only clinical factor significantly and reproducibly associated with VE gene expression. The expression of ACE2 and TMPRSS2 was higher in smokers in the bronchus but not in the nose. scRNA-seq of nasal brushings indicated that ACE2 co-expressed genes were highly expressed in club and C15orf48+ secretory cells while TMPRSS2 co-expressed genes were highly expressed in keratinizing epithelial cells. In contrast, these ACE2 and TMPRSS2 modules were highly expressed in goblet cells in scRNA-seq from bronchial brushings. Cell-type deconvolution of the gene expression data confirmed that smoking increased the abundance of several secretory cell populations in the bronchus, but only goblet cells in the nose. The association of ACE2 and TMPRSS2 with smoking in the bronchus is due to their high expression in goblet cells which increase in abundance in current smoker airways. In contrast, in the nose, these genes are not predominantly expressed in cell populations modulated by smoking. In individuals with elevated lung cancer risk, smoking-induced VE gene expression changes in the nose likely have minimal impact on SARS-CoV-2 infection, but in the bronchus, smoking may lead to higher viral loads and more severe disease.

Evaluation of alpaca tracheal explants as an ex vivo model for the study of Middle East respiratory syndrome coronavirus (MERS-CoV) infection.

Middle East respiratory syndrome coronavirus (MERS-CoV) poses a serious threat to public health. Here, we established an ex vivo alpaca tracheal explant (ATE) model using an air-liquid interface culture system to gain insights into MERS-CoV infection in the camelid lower respiratory tract. ATE can be infected by MERS-CoV, being 103 TCID50/mL the minimum viral dosage required to establish a productive infection. IFNs and antiviral ISGs were not induced in ATE cultures in response to MERS-CoV infection, strongly suggesting that ISGs expression observed in vivo is rather a consequence of the IFN induction occurring in the nasal mucosa of camelids.

Replication of SARS-CoV-2 Omicron BA.2 variant in ex vivo cultures of the human upper and lower respiratory tract.

BACKGROUND: The Omicron BA.2 sublineage has replaced BA.1 worldwide and has comparable levels of immune evasion to BA.1. These observations suggest that the increased transmissibility of BA.2 cannot be explained by the antibody evasion. METHODS: Here, we characterized the replication competence and respiratory tissue tropism of three Omicron variants (BA.1, BA.1.1, BA.2), and compared these with the wild-type virus and Delta variant, in human nasal, bronchial and lung tissues cultured ex vivo. FINDINGS: BA.2 replicated more efficiently in nasal and bronchial tissues at 33°C than wild-type, Delta and BA.1. Both BA.2 and BA.1 had higher replication competence than wild-type and Delta viruses in bronchial tissues at 37°C. BA.1, BA.1.1 and BA.2 replicated at a lower level in lung parenchymal tissues compared to wild-type and Delta viruses. INTERPRETATION: Higher replication competence of Omicron BA.2 in the human upper airway at 33°C than BA.1 may be one of the reasons to explain the current advantage of BA.2 over BA.1. A lower replication level of the tested Omicron variants in human lung tissues is in line with the clinical manifestations of decreased disease severity of patients infected with the Omicron strains compared with other ancestral strains. FUNDING: This work was supported by US National Institute of Allergy and Infectious Diseases and the Theme-Based Research Scheme under University Grants Committee of Hong Kong Special Administrative Region, China.

Cell response analysis in SARS-CoV-2 infected bronchial organoids.

The development of an in vitro cell model that can be used to study severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) research is expected. Here we conducted infection experiments in bronchial organoids (BO) and an BO-derived air-liquid interface model (BO-ALI) using 8 SARS-CoV-2 variants. The infection efficiency in BO-ALI was more than 1,000 times higher than that in BO. Among the bronchial epithelial cells, we found that ciliated cells were infected with the virus, but basal cells were not. Ciliated cells died 7 days after the viral infection, but basal cells survived after the viral infection and differentiated into ciliated cells. Fibroblast growth factor 10 signaling was essential for this differentiation. These results indicate that BO and BO-ALI may be used not only to evaluate the cell response to SARS-CoV-2 and coronavirus disease 2019 (COVID-19) therapeutic agents, but also for airway regeneration studies.

LTSP: long-term slice propagation for accurate airway segmentation.

PURPOSE: Bronchoscopic intervention is a widely used clinical technique for pulmonary diseases, which requires an accurate and topological complete airway map for its localization and guidance. The airway map could be extracted from chest computed tomography (CT) scans automatically by airway segmentation methods. Due to the complex tree-like structure of the airway, preserving its topology completeness while maintaining the segmentation accuracy is a challenging task. METHODS: In this paper, a long-term slice propagation (LTSP) method is proposed for accurate airway segmentation from pathological CT scans. We also design a two-stage end-to-end segmentation framework utilizing the LTSP method in the decoding process. Stage 1 is used to generate a coarse feature map by an encoder-decoder architecture. Stage 2 is to adopt the proposed LTSP method for exploiting the continuity information and enhancing the weak airway features in the coarse feature map. The final segmentation result is predicted from the refined feature map. RESULTS: Extensive experiments were conducted to evaluate the performance of the proposed method on 70 clinical CT scans. The results demonstrate the considerable improvements of the proposed method compared to some state-of-the-art methods as most breakages are eliminated and more tiny bronchi are detected. The ablation studies further confirm the effectiveness of the constituents of the proposed method and the efficacy of the framework design. CONCLUSION: Slice continuity information is beneficial to accurate airway segmentation. Furthermore, by propagating the long-term slice feature, the airway topology connectivity is preserved with overall segmentation accuracy maintained.

Bronchial epithelia from adults and children: SARS-CoV-2 spread via syncytia formation and type III interferon infectivity restriction.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections initiate in the bronchi of the upper respiratory tract and are able to disseminate to the lower respiratory tract, where infections can cause an acute respiratory distress syndrome with a high degree of mortality in elderly patients. We used reconstituted primary bronchial epithelia from adult and child donors to follow the SARS-CoV-2 infection dynamics. We show that, in epithelia from adult donors, infections initiate in multiciliated cells and spread within 24 to 48 h throughout the whole epithelia. Syncytia formed of ciliated and basal cells appeared at the apical side of the epithelia within 3 to 4 d and were released into the apical lumen, where they contributed to the transmittable virus dose. A small number of reconstituted epithelia were intrinsically more resistant to virus infection, limiting virus spread to different degrees. This phenotype was more frequent in epithelia derived from children versus adults and correlated with an accelerated release of type III interferon. Treatment of permissive adult epithelia with exogenous type III interferon restricted infection, while type III interferon gene knockout promoted infection. Furthermore, a transcript analysis revealed that the inflammatory response was specifically attenuated in children. Taken together, our findings suggest that apical syncytia formation is an underappreciated source of virus propagation for tissue or environmental dissemination, whereas a robust type III interferon response such as commonly seen in young donors restricted SARS-CoV-2 infection. Thus, the combination of interferon restriction and attenuated inflammatory response in children might explain the epidemiological observation of age-related susceptibility to COVID-19.

Cytopathological Findings in Bronchoalveolar Lavage from Patients with COVID-19.

Information on cellular analysis of bronchoalveolar lavage (BAL) in patients with COVID-19 is limited. Some studies have described an increase in lymphocyte percentage or exuberant plasmacytosis. Some reports addressed the importance of molecular testing on BAL samples to confirm COVID-19 pneumonia, in clinically highly suspected patients with consecutive negative nasopharyngeal swab results. In addition to atypical lymphocytes in the peripheral blood, morphologic findings of atypical lymphocytes in BAL were also reported in a few patients. The objective of this study was to describe the cytopathic characteristics identified, any data presented here are descriptives and intended to trigger further research. Three general aspects have been evaluated in each sample: reactive changes, virus-related pathological changes, and differential leukocyte count. Seventeen samples were collected. All samples were negative for malignancy, with an inflammatory background, predominantly lymphohistiocytic in 5 samples, histiocytic in 9, and 3 with predominantly neutrophilic. Hemosiderin-laden macrophages were observed in 12/17. Nonspecific reactive cell changes were identified in 4 samples, including bronchial, alveolar, and reserve cell hyperplasia. Virus-related pathological changes were observed in 14 samples, such as loss of nuclear chromatin pattern, lymphocytes with atypical nuclei, nuclear and cytoplasmic inclusions, multinucleations in bronchial cells and macrophages, or multinucleated giant cells. The identification of multinucleated giant cells could represent a cytopathic effect induced by the virus, at the same time the nuclear clearance of pneumocytes as a possible direct effect. BAL is a procedure aimed at obtaining cells from the respiratory tract that can provide valuable and rapid information. It is important to collect and describe as many cytopathological findings as possible, which can provide relevant information for future studies.

Treatment of a pneumatocele in a COVID-19 patient with endobronchial valves.

A man in his 40s was admitted to his local hospital 6 days after the first vague symptoms of COVID-19. His general condition deteriorated, and he was treated in the intensive care unit but did not require mechanical ventilation. During his recovery, he experienced a cough spell, after which his dyspnoea recurred and rapidly increased. CT pulmonary angiogram showed a 10×18 cm cavitary lesion with an air-fluid level and surrounding atelectasis of the right lower lobe. A one-way valve mechanism had developed, leading to the formation of a pneumatocele. The patient was treated by occlusion of all bronchial segments of the right lower lobe with endobronchial valves, and the pneumatocele was evacuated with a pigtail catheter. The valves were removed 4 weeks after insertion, and the right lower lobe re-expanded. Six months after treatment, the patient had recovered completely and almost regained his former lung function.

Efficacy Analysis of DSA-Guided Bronchial Arterial Chemoembolization Interventional Therapy in Patients with Middle-Advanced Primary Bronchial Lung Cancer.

Objective: To investigate the clinical efficacy of digital subtraction angiography- (DSA-) guided bronchial arterial chemoembolization (BACE) in patients with primary bronchial lung cancer. Methods: A total of 178 patients with primary intermediate and advanced bronchial lung cancer admitted to our hospital from February 2019 to March 2020 were selected as the subjects, and they were divided into control group (84 cases) and observation group (94 cases) according to the different chemotherapy regimens adopted by the patients. The control group was treated with traditional perfusion chemotherapy, and the observation group was treated with DSA-guided BACE interventional therapy, treated for 4 cycles, and followed up until the end of June 2021. The short-term clinical efficacy, hemoptysis remission, and incidence of adverse reactions were compared between the two groups. The mortality and recurrence rates between the two groups from treatment to the end of follow-up were counted, and the quality of life after treatment and 1 year after treatment were compared. Results: The short-term remission rate (73.40% vs. 58.33%), disease control rate (93.62% vs. 84.52%), hemoptysis remission rate (75.00% vs. 41.51%), the quality of life after chemotherapy cycle (90.86 ± 2.55 vs. 78.04 ± 2.21), and the quality of life after 1 year of follow-up (85.68 ± 2.23 vs. 70.27 ± 1.72) in the observation group were significantly higher than those in the control group, and the difference was statistically significant (P < 0.05). The incidence of adverse reactions (9.57% vs. 20.24%), mortality (10.64% vs. 21.43%), and recurrence rate (11.70% vs. 27.38%) during the follow-up period in the observation group were significantly lower than those in control group, and the differences were statistically significant (P < 0.05). Conclusion: DSA-guided BACE interventional therapy for patients with primary middle-advanced bronchial lung cancer has significant efficacy, which can not only reduce the mortality and recurrence rate of patients but also improve the quality of life of patients, with fewer adverse reactions and high safety, which is worthy of promotion.

Differential Effect of SARS-CoV-2 Spike Glycoprotein 1 on Human Bronchial and Alveolar Lung Mucosa Models: Implications for Pathogenicity.

BACKGROUND: The SARS-CoV-2 spike protein mediates attachment of the virus to the host cell receptor and fusion between the virus and the cell membrane. The S1 subunit of the spike glycoprotein (S1 protein) contains the angiotensin converting enzyme 2 (ACE2) receptor binding domain. The SARS-CoV-2 variants of concern contain mutations in the S1 subunit. The spike protein is the primary target of neutralizing antibodies generated following infection, and constitutes the viral component of mRNA-based COVID-19 vaccines. METHODS: Therefore, in this work we assessed the effect of exposure (24 h) to 10 nM SARS-CoV-2 recombinant S1 protein on physiologically relevant human bronchial (bro) and alveolar (alv) lung mucosa models cultured at air-liquid interface (ALI) (n = 6 per exposure condition). Corresponding sham exposed samples served as a control. The bro-ALI model was developed using primary bronchial epithelial cells and the alv-ALI model using representative type II pneumocytes (NCI-H441). RESULTS: Exposure to S1 protein induced the surface expression of ACE2, toll like receptor (TLR) 2, and TLR4 in both bro-ALI and alv-ALI models. Transcript expression analysis identified 117 (bro-ALI) and 97 (alv-ALI) differentially regulated genes (p ≤ 0.01). Pathway analysis revealed enrichment of canonical pathways such as interferon (IFN) signaling, influenza, coronavirus, and anti-viral response in the bro-ALI. Secreted levels of interleukin (IL) 4 and IL12 were significantly (p < 0.05) increased, whereas IL6 decreased in the bro-ALI. In the case of alv-ALI, enriched terms involving p53, APRIL (a proliferation-inducing ligand) tight junction, integrin kinase, and IL1 signaling were identified. These terms are associated with lung fibrosis. Further, significantly (p < 0.05) increased levels of secreted pro-inflammatory cytokines IFNγ, IL1ꞵ, IL2, IL4, IL6, IL8, IL10, IL13, and tumor necrosis factor alpha were detected in alv-ALI, whereas IL12 was decreased. Altered levels of these cytokines are also associated with lung fibrotic response. CONCLUSIONS: In conclusion, we observed a typical anti-viral response in the bronchial model and a pro-fibrotic response in the alveolar model. The bro-ALI and alv-ALI models may serve as an easy and robust platform for assessing the pathogenicity of SARS-CoV-2 variants of concern at different lung regions.

SARS-CoV-2 Omicron variant replication in human bronchus and lung ex vivo.

The emergence of SARS-CoV-2 variants of concern with progressively increased transmissibility between humans is a threat to global public health. The Omicron variant of SARS-CoV-2 also evades immunity from natural infection or vaccines1, but it is unclear whether its exceptional transmissibility is due to immune evasion or intrinsic virological properties. Here we compared the replication competence and cellular tropism of the wild-type virus and the D614G, Alpha (B.1.1.7), Beta (B.1.351), Delta (B.1.617.2) and Omicron (B.1.1.529) variants in ex vivo explant cultures of human bronchi and lungs. We also evaluated the dependence on TMPRSS2 and cathepsins for infection. We show that Omicron replicates faster than all other SARS-CoV-2 variants studied in the bronchi but less efficiently in the lung parenchyma. All variants of concern have similar cellular tropism compared to the wild type. Omicron is more dependent on cathepsins than the other variants of concern tested, suggesting that the Omicron variant enters cells through a different route compared with the other variants. The lower replication competence of Omicron in the human lungs may explain the reduced severity of Omicron that is now being reported in epidemiological studies, although determinants of severity are multifactorial. These findings provide important biological correlates to previous epidemiological observations.

Imiquimod Boosts Interferon Response, and Decreases ACE2 and Pro-Inflammatory Response of Human Bronchial Epithelium in Asthma.

Background: Both anti-viral and anti-inflammatory bronchial effects are warranted to treat viral infections in asthma. We sought to investigate if imiquimod, a TLR7 agonist, exhibits such dual actions in ex vivo cultured human bronchial epithelial cells (HBECs), targets for SARS-CoV-2 infectivity. Objective: To investigate bronchial epithelial effects of imiquimod of potential importance for anti-viral treatment in asthmatic patients. Methods: Effects of imiquimod alone were examined in HBECs from healthy (N=4) and asthmatic (N=18) donors. Mimicking SARS-CoV-2 infection, HBECs were stimulated with poly(I:C), a dsRNA analogue, or SARS-CoV-2 spike-protein 1 (SP1; receptor binding) with and without imiquimod treatment. Expression of SARS-CoV-2 receptor (ACE2), pro-inflammatory and anti-viral cytokines were analyzed by RT-qPCR, multiplex ELISA, western blot, and Nanostring and proteomic analyses. Results: Imiquimod reduced ACE2 expression at baseline and after poly(I:C) stimulation. Imiquimod also reduced poly(I:C)-induced pro-inflammatory cytokines including IL-1ß, IL-6, IL-8, and IL-33. Furthermore, imiquimod increased IFN-ß expression, an effect potentiated in presence of poly(I:C) or SP1. Multiplex mRNA analysis verified enrichment in type-I IFN signaling concomitant with suppression of cytokine signaling pathways induced by imiquimod in presence of poly(I:C). Exploratory proteomic analyses revealed potentially protective effects of imiquimod on infections. Conclusion: Imiquimod triggers viral resistance mechanisms in HBECs by decreasing ACE2 and increasing IFN-ß expression. Additionally, imiquimod improves viral infection tolerance by reducing viral stimulus-induced epithelial cytokines involved in severe COVID-19 infection. Our imiquimod data highlight feasibility of producing pluripotent drugs potentially suited for anti-viral treatment in asthmatic subjects.

Local and systemic responses to SARS-CoV-2 infection in children and adults.

It is not fully understood why COVID-19 is typically milder in children1-3. Here, to examine the differences between children and adults in their response to SARS-CoV-2 infection, we analysed paediatric and adult patients with COVID-19 as well as healthy control individuals (total n = 93) using single-cell multi-omic profiling of matched nasal, tracheal, bronchial and blood samples. In the airways of healthy paediatric individuals, we observed cells that were already in an interferon-activated state, which after SARS-CoV-2 infection was further induced especially in airway immune cells. We postulate that higher paediatric innate interferon responses restrict viral replication and disease progression. The systemic response in children was characterized by increases in naive lymphocytes and a depletion of natural killer cells, whereas, in adults, cytotoxic T cells and interferon-stimulated subpopulations were significantly increased. We provide evidence that dendritic cells initiate interferon signalling in early infection, and identify epithelial cell states associated with COVID-19 and age. Our matching nasal and blood data show a strong interferon response in the airways with the induction of systemic interferon-stimulated populations, which were substantially reduced in paediatric patients. Together, we provide several mechanisms that explain the milder clinical syndrome observed in children.