scholarly journals SARS-CoV-2 Omicron spike mediated immune escape, infectivity and cell-cell fusion

2021 ◽  
Author(s):  
Bo Meng ◽  
Isabella Ferreira ◽  
Adam Abdullahi ◽  
Steven A Kemp ◽  
Niluka Goonawardane ◽  
...  
Keyword(s):  
Marked Reduction ◽  
Immune Escape ◽  
Lung Epithelial Cells ◽  
Lung Cells ◽  
Wild Type ◽  
Short Term ◽  
Neutralising Antibodies ◽  
Cleavage Efficiency ◽  
Lung Epithelial ◽  
Vectored Vaccine

The Omicron variant emerged in southern Africa in late 2021 and is characterised by multiple spike mutations across all spike domains. Here we show that the Omicron spike confers very significant evasion of vaccine elicited neutralising antibodies that is more pronounced for ChAdOx-1 adenovirus vectored vaccine versus BNT162b2 mRNA vaccine. Indeed neutralisation of Omicron was not detectable for the majority of individuals who had received two doses of ChAdOx-1. Third dose mRNA vaccination rescues neutralisation in the short term. Despite three mutations predicted to favour spike S1/S2 cleavage, observed cleavage efficiency is lower than for wild type Wuhan-1 D614G and Delta. We demonstrate significantly lower infectivity of lung organoids and Calu-3 lung cells expressing endogenous levels of ACE2 and TMPRSS2 but similar infection as compared to Delta when using H1299 lung epithelial cells. Importantly, fusogenicity of the Omicron spike is impaired, leading to marked reduction in syncitia formation. These observations indicate that Omicron has gained immune evasion properties whilst possibly modulating properties associated with replication and pathogenicity.

scholarly journals Delta spike P681R mutation enhances SARS-CoV-2 fitness over Alpha variant

2021 ◽  
Author(s):  
Yang Liu ◽  
Jianying Liu ◽  
Bryan A. Johnson ◽  
Hongjie Xia ◽  
Zhiqiang Ku ◽  
...  
Keyword(s):  
Human Lung ◽  
Lung Epithelial Cells ◽  
Wild Type ◽  
Competition Assay ◽  
Furin Cleavage ◽  
Cleavage Efficiency ◽  
Furin Cleavage Site ◽  
Lung Epithelial ◽  
The World ◽  
Alpha Variant

SARS-CoV-2 Delta variant has rapidly replaced the Alpha variant around the world. The mechanism that drives this global replacement has not been defined. Here we report that Delta spike mutation P681R plays a key role in the Alpha-to-Delta variant replacement. In a replication competition assay, Delta SARS-CoV-2 efficiently outcompeted the Alpha variant in human lung epithelial cells and primary human airway tissues. Delta SARS-CoV-2 bearing the Alpha-spike glycoprotein replicated less efficiently than the wild-type Delta variant, suggesting the importance of Delta spike in enhancing viral replication. The Delta spike has accumulated mutation P681R located at a furin cleavage site that separates the spike 1 (S1) and S2 subunits. Reverting the P681R mutation to wild-type P681 significantly reduced the replication of Delta variant, to a level lower than the Alpha variant. Mechanistically, the Delta P681R mutation enhanced the cleavage of the full-length spike to S1 and S2, leading to increased infection via cell surface entry. In contrast, the Alpha spike also has a mutation at the same amino acid (P681H), but the spike cleavage from purified Alpha virions was reduced compared to the Delta spike. Collectively, our results indicate P681R as a key mutation in enhancing Delta variant replication via increased S1/S2 cleavage. Spike mutations that potentially affect furin cleavage efficiency must be closely monitored for future variant surveillance.

scholarly journals Role of Lung Epithelial Cells in Defense against Klebsiella pneumoniae Pneumonia

2002 ◽  
Vol 70 (3) ◽  
pp. 1075-1080 ◽  
Author(s):  
Guadalupe Cortés ◽  
Dolores Álvarez ◽  
Carles Saus ◽  
Sebastián Albertí
Keyword(s):  
Epithelial Cells ◽  
Klebsiella Pneumoniae ◽  
Type Strain ◽  
Pathogenic Bacteria ◽  
Wild Type Strain ◽  
Capsular Polysaccharide ◽  
Defense Mechanism ◽  
Lung Epithelial Cells ◽  
Wild Type ◽  
Lung Epithelial

ABSTRACT The airway epithelium represents a primary site for the entry of pathogenic bacteria into the lungs. It has been suggested for many respiratory pathogens, including Klebsiella pneumoniae, that adhesion and invasion of the lung epithelial cells is an early stage of the pneumonia process. We observed that poorly encapsulated K. pneumoniae clinical isolates and an isogenic unencapsulated mutant invaded lung epithelial cells more efficiently than highly encapsulated strains independent of the K type. By contrast, the unencapsulated mutant was completely avirulent in a mouse model of pneumonia, unlike the wild-type strain, which produced pneumonia and systemic infection. Furthermore, the unencapsulated mutant bound more epithelially produced complement component C3 than the wild-type strain. Our results show that lung epithelial cells play a key role as a host defense mechanism against K. pneumoniae pneumonia, using two different strategies: (i) ingestion and control of the microorganisms and (ii) opsonization of the microorganisms. Capsular polysaccharide avoids both mechanisms and enhances the virulence of K. pneumoniae.

scholarly journals The P681H mutation in the Spike glycoprotein confers Type I interferon resistance in the SARS-CoV-2 alpha (B.1.1.7) variant

2021 ◽  
Author(s):  
Maria Jose Lista ◽  
Helena Winstone ◽  
Harry Wilson ◽  
Adam Dyer ◽  
Suzanne Pickering ◽  
...  
Keyword(s):  
Immune Escape ◽  
Transmembrane Protein ◽  
Lung Epithelial Cells ◽  
Type I ◽  
Spike Glycoprotein ◽  
Global Response ◽  
Lung Epithelial ◽  
Interferon Resistance ◽  
The Uk

Variants of concern (VOCs) of severe acute respiratory syndrome coronavirus type-2 (SARS-CoV-2) threaten the global response to the COVID-19 pandemic. The alpha (B.1.1.7) variant appeared in the UK became dominant in Europe and North America in early 2021. The Spike glycoprotein of alpha has acquired a number mutations including the P681H mutation in the polybasic cleavage site that has been suggested to enhance Spike cleavage. Here, we show that the alpha Spike protein confers a level of resistance to the effects of interferon-β (IFNβ) in lung epithelial cells. This correlates with resistance to restriction mediated by interferon-induced transmembrane protein-2 (IFITM2) and a pronounced infection enhancement by IFITM3. Furthermore, the P681H mutation is necessary for comparative resistance to IFNβ in a molecularly cloned SARS-CoV-2 encoding alpha Spike. Overall, we suggest that in addition to adaptive immune escape, mutations associated with VOCs also confer replication advantage through adaptation to resist innate immunity.

scholarly journals Drug repurposing screens reveal FDA approved drugs active against SARS-Cov-2

2020 ◽  
Author(s):  
Mark Dittmar ◽  
Jae Seung Lee ◽  
Kanupriya Whig ◽  
Elisha Segrist ◽  
Minghua Li ◽  
...  
Keyword(s):  
Drug Sensitivity ◽  
Drug Repurposing ◽  
Vero Cells ◽  
Low Ph ◽  
Lung Epithelial Cells ◽  
Lung Cells ◽  
Clinical Implementation ◽  
Lung Epithelial ◽  
Approved Drugs ◽  
Fda Approved Drugs

AbstractThere are an urgent need for antivirals to treat the newly emerged SARS-CoV-2. To identify new candidates we screened a repurposing library of ~3,000 drugs. Screening in Vero cells found few antivirals, while screening in human Huh7.5 cells validated 23 diverse antiviral drugs. Extending our studies to lung epithelial cells, we found that there are major differences in drug sensitivity and entry pathways used by SARS-CoV-2 in these cells. Entry in lung epithelial Calu-3 cells is pH-independent and requires TMPRSS2, while entry in Vero and Huh7.5 cells requires low pH and triggering by acid-dependent endosomal proteases. Moreover, we found 9 drugs are antiviral in lung cells, 7 of which have been tested in humans, and 3 are FDA approved including Cyclosporine which we found is targeting Cyclophilin rather than Calcineurin for its antiviral activity. These antivirals reveal essential host targets and have the potential for rapid clinical implementation.

scholarly journals IL-35 subunit EBI3 alleviates bleomycin-induced pulmonary fibrosis via suppressing DNA enrichment of STAT3

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Donghong Chen ◽  
Guofeng Zheng ◽  
Qing Yang ◽  
Le Luo ◽  
Jinglian Shen
Keyword(s):  
Gene Expression ◽  
Pulmonary Fibrosis ◽  
Regulatory Mechanism ◽  
Lung Epithelial Cells ◽  
Wild Type ◽  
Qrt Pcr ◽  
Lung Epithelial ◽  
Gene Expression Analyses ◽  
Human And Mouse

Abstract Background IL-35 subunit EBI3 is up-regulated in pulmonary fibrosis tissues. In this study, we investigated the pathological role of EBI3 in pulmonary fibrosis and dissected the underlying molecular mechanism. Methods Bleomycin-induced pulmonary fibrosis mouse model was established, and samples were performed gene expression analyses through RNAseq, qRT-PCR and Western blot. Wild type and EBI3 knockout mice were exposed to bleomycin to investigate the pathological role of IL-35, via lung function and gene expression analyses. Primary lung epithelial cells were used to dissect the regulatory mechanism of EBI3 on STAT1/STAT4 and STAT3. Results IL-35 was elevated in both human and mouse with pulmonary fibrosis. EBI3 knockdown aggravated the symptoms of pulmonary fibrosis in mice. EBI3 deficiency enhanced the expressions of fibrotic and extracellular matrix-associated genes. Mechanistically, IL-35 activated STAT1 and STAT4, which in turn suppressed DNA enrichment of STAT3 and inhibited the fibrosis process. Conclusion IL-35 might be one of the potential therapeutic targets for bleomycin-induced pulmonary fibrosis.

scholarly journals The Spike D614G mutation increases SARS-CoV-2 infection of multiple human cell types

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Zharko Daniloski ◽  
Tristan X Jordan ◽  
Juliana K Ilmain ◽  
Xinyi Guo ◽  
Gira Bhabha ◽  
...  
Keyword(s):  
Human Cell ◽  
Human Lung ◽  
Cell Types ◽  
Lung Epithelial Cells ◽  
Site Directed Mutagenesis ◽  
Wild Type ◽  
Protein Variant ◽  
Lung Epithelial ◽  
Minimal Difference ◽  
Novel Variant

A novel variant of the SARS-CoV-2 virus carrying a point mutation in the Spike protein (D614G) has recently emerged and rapidly surpassed others in prevalence. This mutation is in linkage disequilibrium with an ORF1b protein variant (P314L), making it difficult to discern the functional significance of the Spike D614G mutation from population genetics alone. Here, we perform site-directed mutagenesis on wild-type human-codon-optimized Spike to introduce the D614G variant. Using multiple human cell lines, including human lung epithelial cells, we found that the lentiviral particles pseudotyped with Spike D614G are more effective at transducing cells than ones pseudotyped with wild-type Spike. The increased transduction with Spike D614G ranged from 1.3- to 2.4-fold in Caco-2 and Calu-3 cells expressing endogenous ACE2 and from 1.5- to 7.7-fold in A549ACE2 and Huh7.5ACE2 overexpressing ACE2. Furthermore, trans-complementation of SARS-CoV-2 virus with Spike D614G showed an increased infectivity in human cells. Although there is minimal difference in ACE2 receptor binding between the D614 and G614 Spike variants, the G614 variant is more resistant to proteolytic cleavage, suggesting a possible mechanism for the increased transduction.

Abstract 28: Extracellular Matrix Coating Enhances Nanoparticle Uptake by Lung Epithelial Cells

2015 ◽  
Vol 117 (suppl_1) ◽  
Author(s):  
Primana Punnakitikashem ◽  
Priya Ravikumar ◽  
Jinglei Wu ◽  
Kytai Nguyen ◽  
Connie Hsia ◽  
...  
Keyword(s):  
Gene Expression ◽  
Extracellular Matrix ◽  
Epithelial Cells ◽  
Cellular Uptake ◽  
Lung Tissue ◽  
Protein Release ◽  
Lung Epithelial Cells ◽  
Lung Cells ◽  
Significant Difference ◽  
Lung Epithelial

Introduction: Rapid uptake of drug-loaded nanoparticles (NPs) by lung cells is critical for effective pulmonary delivery of therapeutic agents because of the rapid pulmonary clearance mechanisms. We tested the possibility that coating NPs with extracellular matrix (ECM) derived from lung tissue enhances nanoparticles uptake by lung cells. Methods and Results: Fresh adult porcine lung tissue obtained from a local slaughterhouse was decellularized using detergent (sodium dodecyl sulfate) and then enzymatically digested into a soluble solution. The double emulsion method was utilized to fabricate core-shell poly(lactic-co-glycolic) (PLGA) nanoparticles loaded with bovine serum albumin (BSA) for protein release studies, 6-coumarin for cellular uptake studies, or human erythropoietin receptor (hEPOR) cDNA co-expressing green fluorescent protein (GFP) for in vivo gene expression studies. The ECM was coated onto the nanoparticle surface by physical adsorption using the ECM solution (100 μg/ml). There is no significant difference in the diameter, blood compatibility or cell toxicity between coated and uncoated NPs. ECM-coated NPs showed slower protein release rate than uncoated NPs as the ECM coating hindered protein diffusion into the solution. ECM-coated NPs showed significantly higher cellular uptake by human lung epithelial cells than collagen-coated or uncoated NPs. In addition, ECM-coated and uncoated NPs loaded with hEPOR-GFP cDNA were aerosolized and delivered by inhalation into rat lung. Following single inhalation using uncoated NPs, GFP expression in lung tissue progressively increased for up to 21 days. Using the ECM-coated NPs EPOR expression peaked at 14 days, then declined thereafter. Conclusions: Coating NPs with lung-derived ECM markedly enhances NP uptake by lung cells, delays the release of encapsulated protein or DNA, and shortens the duration of peak tissue gene expression compared to uncoated NPs. This NP formulation may be useful where more precise timing of delayed payload release is desired.

scholarly journals Transcriptome Sequencing Data Sets of Human Lung Epithelial Cells in the Course of Francisella tularensis Infection

2020 ◽  
Vol 9 (42) ◽  
Author(s):  
Adi Bercovich-Kinori ◽  
Ma’ayan Israeli ◽  
Theodor Chitlaru ◽  
Inbar Cohen-Gihon ◽  
Ofir Israeli ◽  
...  
Keyword(s):  
Francisella Tularensis ◽  
Human Lung ◽  
Severe Disease ◽  
Lung Epithelial Cells ◽  
Lung Cells ◽  
Data Sets ◽  
Transcriptome Data ◽  
Sequencing Data ◽  
Content Type ◽  
Lung Epithelial

ABSTRACT Francisella tularensis is a highly infectious intracellular bacterium representing the causative agent of tularemia, a severe disease which requires prompt antibacterial intervention for mitigating its potential high mortality. Inhaled bacteria interact with lung cells belonging to various subpopulations, including those of the epithelium. As of today, the host epithelial response to inhalational infection with F. tularensis is poorly understood. Here, we announce host transcriptome data sets which systematically address the human epithelial response to F. tularensis at different time points postinfection.

Reduction of BCNU toxicity to lung cells by high-level expression ofO6-methylguanine-DNA methyltransferase

2001 ◽  
Vol 280 (4) ◽  
pp. L755-L761 ◽  
Author(s):  
Min Wu ◽  
Mark R. Kelley ◽  
W. Kent Hansen ◽  
William J. Martin
Keyword(s):  
Dna Repair ◽  
Survival Rate ◽  
Epithelial Cells ◽  
Dna Methyltransferase ◽  
A549 Cells ◽  
Lung Epithelial Cells ◽  
Lung Cells ◽  
Cytotoxic Effects ◽  
Methylguanine Dna Methyltransferase ◽  
Lung Epithelial

1,3-Bis(2-chloroethyl)-1-nitrosourea (BCNU) is an important cause of pulmonary toxicity. BCNU alkylates DNA at the O6position of guanine. O6-methylguanine-DNA methyltransferase (MGMT) is a DNA repair protein that removes alkyl groups from the O6position of guanine. To determine whether overexpression of MGMT in a lung cell reduces BCNU toxicity, the MGMT gene was transfected into A549 cells, a lung epithelial cell line. Transfected A549 cell populations demonstrated high levels of MGMT RNA, MGMT protein, and DNA repair activity. The overexpression of MGMT in lung epithelial cells provided protection from the cytotoxic effects of BCNU. Control A549 cells incubated with 100 μM BCNU had a cell survival rate of 12.5 ± 1.2%; however, A549 cells overexpressing MGMT had a survival rate of 71.8 ± 2.7% ( P < 0.001). We also demonstrated successful transfection of MGMT into human pulmonary artery endothelial cells and a primary culture of rat type II alveolar epithelial cells with overexpression of MGMT, resulting in significant protection from BCNU toxicity. These data suggest that overexpression of DNA repair proteins such as MGMT in lung cells may protect the lung cells from cytotoxic effects of cancer chemotherapy drugs such as BCNU.

scholarly journals Lung Epithelial TRPA1 Transduces the Extracellular ROS into Transcriptional Regulation of Lung Inflammation Induced by Cigarette Smoke: The Role of Influxed Ca2+

2015 ◽  
Vol 2015 ◽  
pp. 1-16 ◽  
Author(s):  
An-Hsuan Lin ◽  
Meng-Han Liu ◽  
Hsin-Kuo Ko ◽  
Diahn-Warng Perng ◽  
Tzong-Shyuan Lee ◽  
...  
Keyword(s):  
Nadph Oxidase ◽  
Cigarette Smoke ◽  
Lung Inflammation ◽  
Lung Epithelial Cells ◽  
Wild Type ◽  
Intracellular Ros ◽  
Lung Epithelial ◽  
Intracellular Ca ◽  
Acetyl Cysteine

The mechanism underlying the inflammatory role of TRPA1 in lung epithelial cells (LECs) remains unclear. Here, we show that cigarette smoke extract (CSE) sequentially induced several events in LECs. The Ca2+influx was prevented by decreasing extracellular reactive oxygen species (ROS) with the scavenger N-acetyl-cysteine, removing extracellular Ca2+with the chelator EGTA, or treating with the TRPA1 antagonist HC030031. NADPH oxidase activation was abolished by its inhibitor apocynin, EGTA, or HC030031. The increased intracellular ROS was halted by apocynin, N-acetyl-cysteine, or HC030031. The activation of the MAPKs/NF-κB signaling was suppressed by EGTA, N-acetyl-cysteine, or HC030031. IL-8 induction was inhibited by HC030031 or TRPA1 siRNA. Additionally, chronic cigarette smoke (CS) exposure in wild-type mice induced TRPA1 expression in LECs and lung tissues. In CS-exposuretrpa1−/−mice, the increased BALF level of ROS was similar to that of CS-exposure wild-type mice; yet lung inflammation was lessened. Thus, in LECs, CSE may initially increase extracellular ROS, which activate TRPA1 leading to an increase in Ca2+influx. The increased intracellular Ca2+contributes to activation of NADPH oxidase, resulting in increased intracellular ROS, which activate the MAPKs/NF-κB signaling leading to IL-8 induction. This mechanism may possibly be at work in mice chronically exposed to CS.

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