Journal: Frontiers in Immunology

Article Title: SARS-CoV-2 evolution enhances endocytic uptake while preserving TMPRSS2-dependent fusion

doi: 10.3389/fimmu.2025.1736891

Figure Lengend Snippet: Susceptibility of the nine different SARS-CoV-2 strains to inhibitors of TMPRSS2-mediated fusion and cathepsin-mediated endocytosis. SARS-CoV-2 European wild-type (B.1.1), variant-of-concern (VOC) Alpha (B1.1.7 Q27*K68*, B.1.1.7 Q27*), Delta (AY.33), and Omicron (BA.1.17.2, BA.1.1, BA.2.9, BE.1.1, BA.5.1) were propagated in four human cell lines (Calu-3, Caco-2, A549 hACE2+/TMPRSS2+ , HEK293T) in the presence of increasing concentrations (0.024-100 µM) of camostat, an inhibitor of TMPRSS2-mediated viral direct fusion with cellular membrane (green curve), aloxistatin, an inhibitor of cathepsin-mediated viral endocytic uptake (pink curve), and a 1:1 mixture of both inhibitors (black curve). Viral load was determined in cell culture supernatants 48h p.i. using RT-qPCR and are presented as log 10 RNA copies/ml. Controls included cells infected with SARS-CoV-2 in the absence of inhibitors (“virus control”, VC), and cells fixed with 4% paraformaldehyde and exposed to SARS-CoV-2 (“background control”, BG), shown as dashed horizontal lines. Data show mean and standard error of three independent experiments.

Article Snippet: SARS-CoV-2 strains were propagated in human Calu-3, Caco-2 (CLS Cell Lines Service, Eppelheim, Germany), A549 hACE2+/TMPRSS2+ (InvivoGen, San Diego, US), and HEK293T cell lines using DMEM (Gibco, Waltham, MA) supplemented with 1% streptomycin/penicillin and 10% fetal calf serum (Pan Biotech, Aidenbach, Germany).

Techniques: Variant Assay, Membrane, Cell Culture, Quantitative RT-PCR, Infection, Virus, Control

Journal: Frontiers in Immunology

Article Title: SARS-CoV-2 evolution enhances endocytic uptake while preserving TMPRSS2-dependent fusion

doi: 10.3389/fimmu.2025.1736891

Figure Lengend Snippet: Degree of direct TMRPSS2-mediated fusion and cathepsin-mediated endocytic uptake of the nine different SARS-CoV-2 strains in four different cell lines. Calculation of the reduction in viral load induced by aloxistatin (pink columns) and camostat (green columns) at the non-toxic concentration of 25 µM as % inhibition induced by the mixture of the two inhibitors for Calu-3 cells (A) , HEK293T cells (B) , Caco-2 cells (C) and A549 hACE2+/TMPRSS2+ cells (D) . A higher percentage indicates a stronger role of the respective mode of entry for the respective virus strain. In A549 hACE2+/TMPRSS2+ cells, strains BA.1.17.2 and BA.1.1 were non-replicative (n.r.). In HEK293T cells, virus replication was observed for strains BE.1.1 and BA.5.1 only. Statistics was performed using two-way ANOVA with Šidák’s correction to account for multiple comparisons. *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001.

Article Snippet: SARS-CoV-2 strains were propagated in human Calu-3, Caco-2 (CLS Cell Lines Service, Eppelheim, Germany), A549 hACE2+/TMPRSS2+ (InvivoGen, San Diego, US), and HEK293T cell lines using DMEM (Gibco, Waltham, MA) supplemented with 1% streptomycin/penicillin and 10% fetal calf serum (Pan Biotech, Aidenbach, Germany).

Techniques: Concentration Assay, Inhibition, Virus

Journal: Frontiers in Immunology

Article Title: Acetylsalicylic acid disrupts SARS-CoV-2 spike protein glycosylation and selectively impairs binding to ACE2

doi: 10.3389/fimmu.2025.1706997

Figure Lengend Snippet: ASA prevents S1-induced lung injury, fibrosis and inflammation in hACE2-KI mice. (A, B) Representative images of lung sections stained with H&E (A) and Masson’s trichrome (B) from mice receiving intratracheal instillation of vehicle, 15 μg S1 pre-treated overnight with vehicle (S1), 15 μg S1 pre-treated overnight with ASA 20 mg/L (ASA-treated S1) or 15 μg S1 pre-treated overnight with vehicle followed by ASA 20 mg/L administered immediately afterward through the same route (S1+ASA) at 7 days (n=3 per group). Scale bars: 100 µm for H&E and 20 µm for Masson’s trichrome staining. (C-E) Representative images and quantification of fibronectin ( C , red), MAC2 + macrophages ( D , red), and GR1 + neutrophils ( E , red) in lung tissue sections of mice receiving intratracheal instillation of vehicle, S1, ASA-treated S1 or S1+ASA at 7 days (n=3 per group). Lung structures and nuclei were counterstained with WGA lectin (green) and DAPI (blue), respectively. Scale bar: 20 µm. Data are expressed as % of fibronectin fluorescence area per high power field at ×63 magnification (% area/field) and the average number of MAC2 + or GR1 + cells per high power field at ×63 magnification. For all panels, results are shown as mean ± SEM and were analyzed with Tukey’s multiple comparison test. *p-value<0.05, **p-value<0.01, and ***p-value<0.001 vs Vehicle; °°p-value<0.01, and °°°p-value<0.001 vs S1; ## p-value<0.01, and ### p-value<0.001 vs ASA-treated S1.

Article Snippet: Then, wells were incubated with 0.1 μg/mL ACE2 with C-term human Fc tag (Invivogen, fc-hace2) for 1 hour at room temperature in PBS 1X.

Techniques: Staining, Fluorescence, Comparison, Significance Assay

Journal: ACS Omega

Article Title: A Tetrapodal Tryptophan Derivative with Multiple Exposed Free Carboxylic Acids Blocks Host Cell Entry of Omicron SARS-Cov-2 and Respiratory Syncytial Virus

doi: 10.1021/acsomega.5c10442

Figure Lengend Snippet: Evaluation of the antiviral activity and toxicity of different compounds of our in-house collection against VSV pseudotyped with BA.2 (A) and BA.4/5 S (B) proteins. VSV pseudotyped with the indicated S proteins was preincubated with the different compounds at 20 μM, followed by the infection of A549-ACE2-TMPRSS2 cells. Viral infection was monitored by counting GFP-expressing cells. Toxicity was assessed via the analysis of cell confluency. Bars indicate the mean and SE for n = 3. Individual data points are shown as circles.

Article Snippet: A549-ACE2-TMPRSS2 cells (catalog a549-hace2tpsa) were purchased from InVivoGen.

Techniques: Activity Assay, Infection, Expressing

Journal: ACS Omega

Article Title: A Tetrapodal Tryptophan Derivative with Multiple Exposed Free Carboxylic Acids Blocks Host Cell Entry of Omicron SARS-Cov-2 and Respiratory Syncytial Virus

doi: 10.1021/acsomega.5c10442

Figure Lengend Snippet: Compound 2 blocks the entry of RSV into cells. Time-of-addition experiment with 2 treatment at 4 μM prior to and during infection ( Preincubation and Pre - + Post-incubation ) or following infection ( Post-incubation ) of A549 cells with RSV-A encoding the fluorescent protein mKate2 (see ). Viral infection was quantified at 24 h post infection by examination of mKate2 fluorescence.

Article Snippet: A549-ACE2-TMPRSS2 cells (catalog a549-hace2tpsa) were purchased from InVivoGen.

Techniques: Infection, Incubation, Fluorescence