Journal: Human Vaccines & Immunotherapeutics

Article Title: Mucosal immunization with an adenoviral vector expressing a prefusogenic F protein protects the upper and lower respiratory tracts of cotton rats against live respiratory syncytial virus challenge

doi: 10.1080/21645515.2026.2623573

Figure Lengend Snippet: ChAd68-PreF induces strong neutralization antibody titers against RSV A and RSV B subtypes. serum 60% plaque-reduction neutralization titers (PRNT 60 ) against RSV A2 (A), RSV B (B) and RSV Long (C) determined at days 0, 56 and 61. Symbols represent individual cotton rats; bars show mean ± SD. Statistics: data were log-transformed and analyzed by two-way repeated measures ANOVA with Tukey’s multiple-comparisons test, considering treatment group and time as factors (* p < .05; *** p < .001; **** p < .0001).

Article Snippet: Neutralization assays in cotton rats were performed using the following strains: RSV A2 (ATCC VR-1540), RSV B (strain 18537; ATCC VR-1580PQ), and RSV Long (ATCC VR-26).

Techniques: Neutralization, Transformation Assay

Journal: Microbiology Spectrum

Article Title: Cathepsin S contributes to influenza-induced lung injury by driving inflammation, promoting apoptosis, and disrupting epithelial barrier integrity

doi: 10.1128/spectrum.01128-25

Figure Lengend Snippet: Pathogenesis dynamics and transcriptomic changes during influenza infection in mice. ( A ) Left: the PCA for the sequencing samples at 5 dpi with various infection dose. Right: the PCA for the sequencing samples with 100 or 1,000 PFU PR8 IAVs infection during the time post-infection (1, 3, 5, and 7 dpi). ( B ) Volcano plots of the DEGs between PBS and IAVs treated groups (10 1 , 10 2 , 10 3 , and 10 4 PFU, respectively). The abscissa represents the log 2 FC, the ordinate represents the −log 10 ( P adj). Red points are the upregulated DEGs defined by the absolute value of FC > 2 and P adj < 0.05, while blue points are the downregulated DEGs. ( C and D ) Heatmap clustering of key extracellular-matrix-related genes from overlap DEGs across various infection doses at 5 dpi ( C ) and across multiple timepoints in the 100 PFU infection group ( D ). Expression levels are shown as log 2 -transformed FPKM values after centralization correction, highlighting the top 30 genes with significant changes relative to the PBS group. ( E ) WGCNA module identification and trait correlation, positive and negative correlations are indicated by red and blue colors, respectively. ( F ) Gene interaction network within the ME-darkred module, where node color depth indicates the number of connections. The central area highlights candidate hub genes.

Article Snippet: The Influenza A/Puerto Rico/8/1934 (H1N1) (PR8) virus (ATCC, VR-95PQ) was propagated in 9–11-day-old embryonated chicken eggs and titrated in MDCK cells using either plaque assays or 50% tissue culture infective dose (TCID 50 ) assays ( , ).

Techniques: Infection, Sequencing, Expressing, Transformation Assay

Journal: Microbiology Spectrum

Article Title: Cathepsin S contributes to influenza-induced lung injury by driving inflammation, promoting apoptosis, and disrupting epithelial barrier integrity

doi: 10.1128/spectrum.01128-25

Figure Lengend Snippet: CTSS is upregulated by influenza infection in mouse lung. ( A ) CTSS mRNA expression in lung tissues from mice ( n = 3 per group) inoculated with either PBS or 100 PFU of PR8 virus, quantified by RT-qPCR. ( B ) Western blotting analysis of proteins in lung tissues from mice inoculated with PBS or 100 PFU of PR8 virus. ( C ) CTSS activity in BALF and lung homogenates from mice inoculated with PBS or 100 PFU of PR8, measured using a CTSS Activity Assay Kit. ( D ) Left panel: comparison of CTSS mRNA expression levels in the lungs of mice infected with 5 PFU and 100 PFU of IAV, determined by RT-qPCR. Right panel: western blotting analysis of activated CTSS protein levels in lung tissues from mice infected with 5 PFU and 100 PFU of PR8. ( E ) Temporal profile of CTSS mRNA expression across various days post-infection, measured via RT-qPCR (left), and western blotting analysis of activated CTSS protein levels across different timepoints post-infection (right). Data are represented as mean ± standard deviation (SD). **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

Article Snippet: The Influenza A/Puerto Rico/8/1934 (H1N1) (PR8) virus (ATCC, VR-95PQ) was propagated in 9–11-day-old embryonated chicken eggs and titrated in MDCK cells using either plaque assays or 50% tissue culture infective dose (TCID 50 ) assays ( , ).

Techniques: Infection, Expressing, Virus, Quantitative RT-PCR, Western Blot, Activity Assay, Comparison, Standard Deviation

Journal: Microbiology Spectrum

Article Title: Cathepsin S contributes to influenza-induced lung injury by driving inflammation, promoting apoptosis, and disrupting epithelial barrier integrity

doi: 10.1128/spectrum.01128-25

Figure Lengend Snippet: The effects of CTSS inhibitor LY3000328 on influenza infection in mice. ( A ) Schematic diagram of the experimental design. C57BL/6J mice ( n = 5 per group) received intraperitoneal injection of 30 mg/kg LY3000328 or a placebo vehicle twice daily for 5 days. Four hours after the initial dose, mice were intranasally inoculated with 100 PFU of PR8 virus. A mock control group received the vehicle orally twice daily and PBS intranasally. Body weight was monitored daily. Mouse lungs were collected at 5 dpi for the analyses of viral titers, histopathology, and gene expression. ( B ) CTSS activity in lung homogenate among different groups. ( C ) Viral titers in lung homogenates determined using the TCID 50 assay in MDCK cells. ( D ) Body weight change of mice across the experimental period. ( E ) Survival curve of mice across the experimental period. ( F ) Histopathological examination of mouse lungs performed by H&E staining. ( G ) Analysis of mRNA expression levels of cytokines in lung tissues by RT-qPCR. Data are represented as mean ± SD. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, non-significant.

Article Snippet: The Influenza A/Puerto Rico/8/1934 (H1N1) (PR8) virus (ATCC, VR-95PQ) was propagated in 9–11-day-old embryonated chicken eggs and titrated in MDCK cells using either plaque assays or 50% tissue culture infective dose (TCID 50 ) assays ( , ).

Techniques: Infection, Injection, Virus, Control, Histopathology, Gene Expression, Activity Assay, Staining, Expressing, Quantitative RT-PCR

Journal: Microbiology Spectrum

Article Title: Cathepsin S contributes to influenza-induced lung injury by driving inflammation, promoting apoptosis, and disrupting epithelial barrier integrity

doi: 10.1128/spectrum.01128-25

Figure Lengend Snippet: Influenza infection activates CTSS expression in lung epithelial cells (A549). ( A ) CTSS mRNA expression in A549 cells following infection with PR8 virus at MOIs of 0, 0.1, 0.5, and 1. ( B ) Protein levels of pro-CTSS and activated CTSS in A549 cells 24 hpi with serial MOIs of PR8 virus, assessed by western blotting. ( C ) CTSS activity in the supernatant and cell lysate of A549 cells infected with indicated MOI of PR8 virus at 24 hpi, measured using a CTSS Activity Assay Kit. Data are presented as mean ± SD from three independent experiments. ***, P < 0.001; ****, P < 0.0001; ns, non-significant.

Article Snippet: The Influenza A/Puerto Rico/8/1934 (H1N1) (PR8) virus (ATCC, VR-95PQ) was propagated in 9–11-day-old embryonated chicken eggs and titrated in MDCK cells using either plaque assays or 50% tissue culture infective dose (TCID 50 ) assays ( , ).

Techniques: Infection, Expressing, Virus, Western Blot, Activity Assay

Journal: Microbiology Spectrum

Article Title: Cathepsin S contributes to influenza-induced lung injury by driving inflammation, promoting apoptosis, and disrupting epithelial barrier integrity

doi: 10.1128/spectrum.01128-25

Figure Lengend Snippet: CTSS does not affect PR8 virus replication but reduces cytokine expression in A549 cells. A549 cells were transfected with two different CTSS-siRNAs (siCTSS #1 and siCTSS #2) or a scramble siRNA (siSCR) and then infected with 0.25 MOI of PR8 virus. ( A ) Left panel: representative western Blot analysis of CTSS and viral proteins (HA and NP) expression at 24 hpi, Right panel: intensity scanning of the bands from the left, quantified with data from three independent repeats. ( B ) Viral M gene expression at 24 hpi was quantified by RT-qPCR. ( C ) Viral titers at specified timepoints were measured using the TCID 50 assay. ( D ) The changes in mRNA expression levels of CTSS, TNF-α, RANTES, and IP-10 at 24 hpi were assessed by RT-qPCR. Data are presented as mean ± SD. Two to three independent experimental repeats were conducted for each assay. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, non-significant.

Article Snippet: The Influenza A/Puerto Rico/8/1934 (H1N1) (PR8) virus (ATCC, VR-95PQ) was propagated in 9–11-day-old embryonated chicken eggs and titrated in MDCK cells using either plaque assays or 50% tissue culture infective dose (TCID 50 ) assays ( , ).

Techniques: Virus, Expressing, Transfection, Infection, Western Blot, Gene Expression, Quantitative RT-PCR

Journal: Microbiology Spectrum

Article Title: Cathepsin S contributes to influenza-induced lung injury by driving inflammation, promoting apoptosis, and disrupting epithelial barrier integrity

doi: 10.1128/spectrum.01128-25

Figure Lengend Snippet: CTSS is released into the cytoplasm during influenza infection, thereby regulating apoptosis and epithelial barrier integrity. ( A ) Cell death was assessed by measuring LDH release from non-targeting and CTSS-targeting siRNA transfected A549 cells following PR8 infection at 24 hp i, using an LDH Cytotoxicity Assay Kit (Roche, USA, 91963) according to the manufacturer’s instructions. ( B ) TUNEL staining and quantification of positive cells in non-targeting and CTSS-targeting siRNA-transfected A549 cells after PR8 virus infection at 24 hpi. Representative images (left panel) and statistical data (right panel) are shown. ( C ) Protein levels of CTSS, LAMP1, NP, and HA and PARP1 in the cytoplasm, lysosome, and supernatant of A549 cells infected with PR8 (MOI = 0.1) at 0, 6, 12, and 24 hpi were determined by western blotting. ( D ) CTSS activity in the cytoplasm, lysosome, and supernatant of A549 cells infected with PR8 virus was measured using a CTSS Activity Assay Kit. ( E ) Expression of CTSS, intercellular tight junctions, and cell death markers in non-targeting and CTSS-targeting siRNA transfected A549 cells following PR8 virus infection at 24 hpi was determined by western blotting. Data are represented as mean ± SD. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

Article Snippet: The Influenza A/Puerto Rico/8/1934 (H1N1) (PR8) virus (ATCC, VR-95PQ) was propagated in 9–11-day-old embryonated chicken eggs and titrated in MDCK cells using either plaque assays or 50% tissue culture infective dose (TCID 50 ) assays ( , ).

Techniques: Infection, Transfection, LDH Cytotoxicity Assay, TUNEL Assay, Staining, Virus, Western Blot, Activity Assay, Expressing