stat1 knockout stat1 hep 2 cells  (ATCC)

Journal: npj Viruses

Article Title: STAT1 signaling controls cholesterol metabolism in epithelial cells and RSV-induced syncytia formation

doi: 10.1038/s44298-026-00173-w

Figure Lengend Snippet: A Sequence alignment of wild-type and STAT1 −/− to STAT1 gene of the human genome (GrCh38.p14). The red bar indicates the binding site of sgRNAs. The red star indicates the 17 bp deletion on the second allele. B Western blot of whole cell lysates of wild-type and STAT1 −/− HEp-2 cells targeting STAT1 (top) and β-Actin (bottom) as loading control. C Bright-field IncuCyte live-cell images of wild-type (left) and STAT1 −/− HEp-2 (right) at 72 hours (h) post-seeding. Images representative of three independent experiments (10 technical replicates). Scale bar indicated next to images. D Quantification of cell eccentricity for wild-type and STAT1 −/− HEp-2 cells at 72 h post-seeding based on live cell images. Eccentricity was assessed at approximately 60% confluence, which was reached at 72 h post-seeding for both cell types. Minimum area, minimum eccentricity, and Hole fill were set to 600 μm 2 , 0.3, and 1000 μm 2 , respectively. Mean ± SD from three independent experiments, with 10 replicates each, are shown. Statistical analysis: Unpaired t-test. E Quantitative analysis of IncuCyte live-cell images for cells per well, with images taken every 3 h for 4.5 days. Mean ± SD of three independent experiments (10 technical replicates) is shown, with non-linear regressions applied for wild-type (green) and STAT1 −/− (red) HEp-2 cells. Statistical analysis: Mann-Whitney test for comparison of ranks. F Volcano plot demonstrating the number of differentially expressed genes between wild-type and STAT1 −/− HEp-2 cells with thresholds p < 0.05 and |Log2 Fold change | ≥ 1. G Bar plot of Gene Set Enrichment Analysis of Gene Ontology. The plot shows significantly enriched biological processes in STAT1 −/− compared to wild-type HEp-2 cells.

Article Snippet: HEp-2 (ATCC, CCL-23) and STAT1 knockout (STAT1 −/− ) HEp-2 cells were cultured in Minimum Essential medium with Earl’s salts (Capricorn Scientific, MEM-A) supplemented with 10% fetal bovine serum (FBS, Gibco), 1% penicillin/streptomycin (P/S, Capricorn Scientific), 1% non-essential amino acids (Gibco), and 1% GlutaMAX (Gibco) at 37 °C, 5% CO 2 .

Techniques: Sequencing, Binding Assay, Western Blot, Control, MANN-WHITNEY, Comparison

Journal: npj Viruses

Article Title: STAT1 signaling controls cholesterol metabolism in epithelial cells and RSV-induced syncytia formation

doi: 10.1038/s44298-026-00173-w

Figure Lengend Snippet: A Viral replication kinetics of RSV-A-0594 (dashed lines) and rRSV-A-0594-eGFP (solid lines)-infected wild-type (green) and STAT1 −/− (red) HEp-2 cells (MOI 0.05). Mean ± SD of three independent experiments (four technical replicates) is shown. Statistical analysis: Two-way ANOVA with Fisher’s LSD test. B Fluorescence images of rRSV-A-0594-eGFP-infected (MOI 0.05) wild-type (top) and STAT1 −/− (bottom) Hep-2 cells at 48 hours post-infection (hpi). Images representative of three independent experiments (two technical replicates). Scale bar indicated next to images. C Graph showing the comparison of viral transcript numbers from RNA-Seq analysis between wild-type and STAT1 −/− HEp-2 cells from 0 to 72 hpi with thresholds set at p < 0.05 and |Log2 Fold change | ≥ 1. D Presatovir-based fusion inhibition assay on wild-type (green) and STAT1 −/− (red) HEp-2 cells at 72 hpi with rRSV-A-0594-eGFP (MOI 0.05). Fluorescence intensity (eGFP) relative to untreated, infected cells is shown. Solid lines indicate mean values, while dashed lines above and below each curve indicate the corresponding standard deviation (Mean ± SD). Horizontal dashed line and the right graph indicate Half Maximal Inhibitory Concentration (IC 50 ) of wild-type (green) and STAT1 −/− (red) HEp-2 based on non-linear regression fitted to graphs. Mean ± SD of three independent experiments (eight technical replicates) is shown. Statistical analysis for IC 50 comparison: Unpaired t-test with Welsh’s correction.

Article Snippet: HEp-2 (ATCC, CCL-23) and STAT1 knockout (STAT1 −/− ) HEp-2 cells were cultured in Minimum Essential medium with Earl’s salts (Capricorn Scientific, MEM-A) supplemented with 10% fetal bovine serum (FBS, Gibco), 1% penicillin/streptomycin (P/S, Capricorn Scientific), 1% non-essential amino acids (Gibco), and 1% GlutaMAX (Gibco) at 37 °C, 5% CO 2 .

Techniques: Infection, Fluorescence, Comparison, RNA Sequencing, Inhibition, Standard Deviation, Concentration Assay

Journal: npj Viruses

Article Title: STAT1 signaling controls cholesterol metabolism in epithelial cells and RSV-induced syncytia formation

doi: 10.1038/s44298-026-00173-w

Figure Lengend Snippet: A UpSet plot demonstrating differentially expressed genes that are shared and unique to infected wild-type and STAT1 −/− HEp-2 after comparison to respective uninfected controls with thresholds p < 0.05 and |Log2 Fold change | ≥ 1. B Cleveland Dot Plot of Gene Set Enrichment Analysis (GSEA) of Gene Ontology comparing processes enriched in both infected wild-type and STAT1 −/− HEp-2 cells based on Normalized Enrichment Scores (NES). C Quantification of total free cholesterol by luminescent enzymatic assay in uninfected wild-type (green) and STAT1 −/− (red) HEp-2 cells. Statistical analysis: Mann-Whitney test. D Quantification of total free cholesterol by luminescent enzymatic assay in wild-type and STAT1 −/− HEp-2 cells, which were uninfected (grey), treated with 5 mM methyl-beta-cyclodextrin (MβCD, orange), infected at MOI 0.05 (light red), infected at MOI (dark red), or treated with 100 ng/ml IFN-α2 and 50 ng/mL IFN-γ at 24 and 48 hpi. Only significant differences between untreated and treated cells within a cell type or within treatments between cell types are shown. Statistical analysis: two-way ANOVA with Fisher’s LSD test. Mean ± SD of four independent experiments (two technical replicates) shown for ( C , D ).

Article Snippet: HEp-2 (ATCC, CCL-23) and STAT1 knockout (STAT1 −/− ) HEp-2 cells were cultured in Minimum Essential medium with Earl’s salts (Capricorn Scientific, MEM-A) supplemented with 10% fetal bovine serum (FBS, Gibco), 1% penicillin/streptomycin (P/S, Capricorn Scientific), 1% non-essential amino acids (Gibco), and 1% GlutaMAX (Gibco) at 37 °C, 5% CO 2 .

Techniques: Infection, Comparison, Enzymatic Assay, MANN-WHITNEY

Journal: npj Viruses

Article Title: STAT1 signaling controls cholesterol metabolism in epithelial cells and RSV-induced syncytia formation

doi: 10.1038/s44298-026-00173-w

Figure Lengend Snippet: Representative fluorescence images of rRSV-A-0594-eGFP-infected wild-type ( A ) and STAT1 −/− ( B ) HEp-2 cells (MOI 0.05) at 48 hpi. Cells were left untreated (first row), treated with 50 μM Gemfibrozil (GFZ, second row), treated with 5 μM methyl-β-cyclodextrin (MβCD) and 50 μM GFZ (third row), or with 10 μM 25-hydroxycholesterol (25-HC, bottom panel). Free cholesterol in plasma membranes was visualized by Filipin III staining (0.05 mg/ml), and DNA was visualized by Propidium iodide (PI) staining (5 μg/ml). Images representative of three independent experiments (two technical replicates). Scale bar indicated next to images.

Article Snippet: HEp-2 (ATCC, CCL-23) and STAT1 knockout (STAT1 −/− ) HEp-2 cells were cultured in Minimum Essential medium with Earl’s salts (Capricorn Scientific, MEM-A) supplemented with 10% fetal bovine serum (FBS, Gibco), 1% penicillin/streptomycin (P/S, Capricorn Scientific), 1% non-essential amino acids (Gibco), and 1% GlutaMAX (Gibco) at 37 °C, 5% CO 2 .

Techniques: Fluorescence, Infection, Clinical Proteomics, Staining

Journal: npj Viruses

Article Title: STAT1 signaling controls cholesterol metabolism in epithelial cells and RSV-induced syncytia formation

doi: 10.1038/s44298-026-00173-w

Figure Lengend Snippet: A Fluorescence images from IncuCyte live-cell imaging of rRSV-A-0594-eGFP-infected wild-type (left) and STAT1 −/− (right) HEp-2 cells (MOI 0.05), which were left untreated (left column), treated with 50 μM Gemfibrozil (GFZ, middle column), or with 10 μM 25-hydroxycholesterol (25-HC, right column) at 12, 24, 36, and 48 hpi. Images representative of three independent (two technical replicates) experiments. Scale bar indicated next to images. B Quantification of syncytia size by IncuCyte live-cell imaging for green (fluorescent) area per image relative to green object count per image of rRSV-A-0594-eGFP-infected wild-type (solid lines) and STAT1 −/− (dashed lines) HEp-2 cells (MOI 0.05), either untreated (red), treated with 50 μM GFZ (blue), or 10 μM 25-HC (green). Images of cells were taken every hour for up to 48 hpi. Mean ± SEM of eight independent experiments (two technical replicates) is shown. Statistical analysis was conducted on the overall distribution of syncytia size during the course of infection using the Friedman test with Dunn’s multiple-comparison test.

Article Snippet: HEp-2 (ATCC, CCL-23) and STAT1 knockout (STAT1 −/− ) HEp-2 cells were cultured in Minimum Essential medium with Earl’s salts (Capricorn Scientific, MEM-A) supplemented with 10% fetal bovine serum (FBS, Gibco), 1% penicillin/streptomycin (P/S, Capricorn Scientific), 1% non-essential amino acids (Gibco), and 1% GlutaMAX (Gibco) at 37 °C, 5% CO 2 .

Techniques: Fluorescence, Live Cell Imaging, Infection, Comparison

Journal: npj Viruses

Article Title: STAT1 signaling controls cholesterol metabolism in epithelial cells and RSV-induced syncytia formation

doi: 10.1038/s44298-026-00173-w

Figure Lengend Snippet: Confocal microscopy images of RSV-A-0594-infected wild-type ( A ) and STAT1 −/− ( B ) HEp-2 cells (MOI 0.05) at 48 hpi. Infected cells were left untreated (top), treated with 50 μM Gemfibrozil (GFZ, middle), or 10 μM 25-hydroxycholesterol (25-HC, bottom). Cells were stained for RSV postfusion F protein on the cell surface. Intracellular actin was visualized by ActinRed staining and nuclei by NucBlue staining. Images representative of three independent experiments (two technical replicates). Scale bar indicated next to images.

Article Snippet: HEp-2 (ATCC, CCL-23) and STAT1 knockout (STAT1 −/− ) HEp-2 cells were cultured in Minimum Essential medium with Earl’s salts (Capricorn Scientific, MEM-A) supplemented with 10% fetal bovine serum (FBS, Gibco), 1% penicillin/streptomycin (P/S, Capricorn Scientific), 1% non-essential amino acids (Gibco), and 1% GlutaMAX (Gibco) at 37 °C, 5% CO 2 .

Techniques: Confocal Microscopy, Infection, Staining

Journal: npj Viruses

Article Title: STAT1 signaling controls cholesterol metabolism in epithelial cells and RSV-induced syncytia formation

doi: 10.1038/s44298-026-00173-w

Figure Lengend Snippet: Western blot of membrane fractions and total cell lysates in uninfected (‘Mock’) and rRSV-A-0594-eGFP-infected (MOI 0.5) wild-type ( A ) and STAT1 −/− HEp-2 ( B ) HEp-2 cells at 48 hpi. Infected cells were left untreated, treated with 50 μM Gemfibrozil (GFZ), treated with 5 μM methyl-β-cyclodextrin (MβCD) and 50 μM GFZ, or with 10 μM 25-hydroxycholesterol (25-HC). Membrane fractions were stained for prefusion F protein, postfusion F protein, and Na + -K + -ATPase (loading control). Total cell lysates were stained for total F protein (prefusion and postfusion conformation) and β-Actin (loading control). Images representative of three independent experiments.

Article Snippet: HEp-2 (ATCC, CCL-23) and STAT1 knockout (STAT1 −/− ) HEp-2 cells were cultured in Minimum Essential medium with Earl’s salts (Capricorn Scientific, MEM-A) supplemented with 10% fetal bovine serum (FBS, Gibco), 1% penicillin/streptomycin (P/S, Capricorn Scientific), 1% non-essential amino acids (Gibco), and 1% GlutaMAX (Gibco) at 37 °C, 5% CO 2 .

Techniques: Western Blot, Membrane, Infection, Staining, Control