Journal: Cell Proliferation

Article Title: FKBP5 Mediates Alveolar Fibroblast Necroptosis During Acute Respiratory Distress Syndrome

doi: 10.1111/cpr.70075

Figure Lengend Snippet: Fibroblast‐specific FKBP5 deletion reduces pulmonary inflammation in septic ARDS. (A) Septic ARDS model using Col1a2 ‐iCre Fkbp5 flox/flox mice. (B) Immunofluorescence of FKBP5 in fibroblasts (200×; scale bar, 50 μm). (C) H&E‐stained lung sections and histopathology scoring (200×; scale bar, 50 μm). (D) FITC‐dextran assay of vascular permeability. (E) Neutrophil percentage in BALF measured by flow cytometry. (F, G) ELISA quantification of TNF‐α, IL‐1β, IL‐6, CXCL1, and CXCL2 in BALF. (H) BALF levels of MPO and cfDNA. (I) TEM of glycocalyx (8000×; scale bar, 500 nm). (J) TEM of tight junctions (20,000×; scale bar, 200 nm). All data are presented as means ± SD from three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; ns, not significant.

Article Snippet: ELISA kits for TNF‐α (EK182, EK282), IL‐1β (EK101B, EK201B), IL‐6 (EK206), CXCL1 (EK196, EK296), and CXCL2 (EK1264, EK214) were sourced from Multi Sciences Biotech (Hangzhou, China).

Techniques: Immunofluorescence, Staining, Histopathology, Permeability, Flow Cytometry, Enzyme-linked Immunosorbent Assay

Journal: Cell Proliferation

Article Title: FKBP5 Mediates Alveolar Fibroblast Necroptosis During Acute Respiratory Distress Syndrome

doi: 10.1111/cpr.70075

Figure Lengend Snippet: FKBP5 in mouse alveolar fibroblasts mediates LPS‐induced cytokine storms via necroptosis. (A) Schematic of enzyme‐digested fibroblast isolation and Scube2 + cell immunofluorescence (100×; scale bar, 100 μm). (B, C) Western blot of NF‐κB pathway proteins and cytokines (IL‐1β, TNF‐α, HMGB1) after LPS treatment (10 μg/mL, 6 h). (D) Volcano plot of differentially expressed genes (DEGs) ( p < 0.05, fold change ≥ 2). (E, F) Heatmaps of differentially expressed proteins. (G) Gene Ontology enrichment analysis of DEGs. (H, I) Western blot validation of necroptosis activation. (J) ELISA of CXCL1 and CXCL2 levels in fibroblast supernatant. All data are presented as means ± SD from three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; ns, not significant.

Article Snippet: ELISA kits for TNF‐α (EK182, EK282), IL‐1β (EK101B, EK201B), IL‐6 (EK206), CXCL1 (EK196, EK296), and CXCL2 (EK1264, EK214) were sourced from Multi Sciences Biotech (Hangzhou, China).

Techniques: Isolation, Immunofluorescence, Western Blot, Biomarker Discovery, Activation Assay, Enzyme-linked Immunosorbent Assay

Journal: Cell Proliferation

Article Title: FKBP5 Mediates Alveolar Fibroblast Necroptosis During Acute Respiratory Distress Syndrome

doi: 10.1111/cpr.70075

Figure Lengend Snippet: FKBP5 regulates necroptosis‐driven cytokine release in human alveolar fibroblasts. (A, B) Western blot analysis of NF‐κB signalling, IL‐1β, and TNF‐α after LPS stimulation (10 μg/mL, 6 h) with or without siRNA Fkbp5 . (C, D) Western blot of necroptosis markers in fibroblasts with siRNA Fkbp5 . (E, F) ELISA analysis of TNF‐α, IL‐1β, CXCL1, and CXCL2 in cell supernatant. All data are presented as means ± SD from three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; ns, not significant.

Article Snippet: ELISA kits for TNF‐α (EK182, EK282), IL‐1β (EK101B, EK201B), IL‐6 (EK206), CXCL1 (EK196, EK296), and CXCL2 (EK1264, EK214) were sourced from Multi Sciences Biotech (Hangzhou, China).

Techniques: Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Cell Proliferation

Article Title: FKBP5 Mediates Alveolar Fibroblast Necroptosis During Acute Respiratory Distress Syndrome

doi: 10.1111/cpr.70075

Figure Lengend Snippet: FKBP5 mediates necroptosis‐induced inflammatory signalling in human alveolar fibroblasts. (A, B) Immunofluorescence of phosphorylated RIPK1, RIPK3, and MLKL after TSZ stimulation 9 h (200×; scale bar, 50 μm). (C, D) Western blot of NF‐κB signalling, HMGB1, IL‐1β, and TNF‐α. (E, F) siRNA Fkbp5 effect on TSZ‐induced necroptosis confirmed by Western blot. (G, H) Western blot showing suppression of NF‐κB signalling by siRNA Fkbp5 . (I) ELISA quantification of TNF‐α, IL‐1β, IL‐6, CXCL1, and CXCL2 in fibroblast supernatants. All data are presented as means ± SD from three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001; ns, not significant.

Article Snippet: ELISA kits for TNF‐α (EK182, EK282), IL‐1β (EK101B, EK201B), IL‐6 (EK206), CXCL1 (EK196, EK296), and CXCL2 (EK1264, EK214) were sourced from Multi Sciences Biotech (Hangzhou, China).

Techniques: Immunofluorescence, Western Blot, Enzyme-linked Immunosorbent Assay