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

Abstract

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.

Effects of SPA4 peptide on lipopolysaccharide‐disrupted lung epithelial barrier, injury, and function in a human cell system and mouse model of lung injury

“…Our results demonstrated that SPA4 peptide treatment significantly improved the LPS‐disrupted TEER and lung alveolar epithelial barrier function in a dose‐ and time‐dependent manner (100 μM SPA4 peptide; p < 0.05 and p = 0.0753 at 24 and 48 h, respectively, Figure 2 ). Since the H441 cells express an increased level of TLR4 even at basal level [Supplemental Figure in (Jeon et al, 2016 ), and (Rahman et al, 2021 )], it is likely that an increased treatment dose (100 μM) was needed for restoration of the epithelial barrier.…”

Insight into the pulmonary molecular toxicity of heated tobacco products using human bronchial and alveolar mucosa models at air–liquid interface

“…The detailed protocol and details of cellular differentiation (Club cells, goblet cells, basal cells, ciliated cells, etc.) of the bro-ALI model has been described previously and the model has been in several studies 34 , 35 , 37 – 41 .…”

“…The alv-ALI model was developed using NCI-H441 (ATCC HTB-174; derived from the pericardial fluid of a patient with papillary adenocarcinoma of the lung) cell line and the detailed protocol and model characteristics have been described recently 35 , 37 . The NCI-H441 cells, representative of human type II pneumocytes, express constitutively the mRNA and protein of the major surfactant apo-protein.…”

Adverse effects of COVID-19 mRNA vaccines: the spike hypothesis

“…Reportedly, S protein (apart from the LNP-mRNA platform discussed earlier) mediates proinflammatory and/or injury (of different etiology) responses in various human cell types [102][103][104], and ACE2-mediated infection of human bronchial epithelial cells with S protein pseudovirions induced inflammation and apoptosis [105]. Also, S protein promoted an inflammatory cytokine IL-6/IL-6Rinduced trans signaling response and alarmin secretion in human endothelial cells, along with increased oxidative stress, induction of inflammatory paracrine senescence, and higher levels of leucocyte adhesion [106].…”