Journal: Aging Cell
Article Title: The Proinflammatory Secretome of Senescent Cells Can Be Controlled by a HIF2A ‐Dependent Upregulation and a FURIN ‐Dependent Cleavage of the ANGPTL4 Secreted Factor
doi: 10.1111/acel.70307
Figure Lengend Snippet: Knockdown of ANGPTL4 inhibits the ability of the SASP to induce activation of human neutrophils, a mark of inflammation. (A, B) Supernatants of MRC5/RAF:ER cells transfected with control (siCTRL), ANGPTL4 (siANGPTL4) or RELA (siRELA) siRNA and treated (+) or not (−) with 4‐OHT to induce senescence (OIS) were applied to primary human neutrophils. (A) Analysis of CD63 neutrophils degranulation marker. A representative flow cytometry profile is shown. Histograms on the right panel are the quantification of MFI. Mean ± SEM of n = 3 independent experiments. Paired one‐way ANOVA test results are shown. (B) Analysis of STAT5 pathway activation according to phosphorylation of STAT5. The left panel displays a representative flow cytometry profile. Histograms on the right panel are the quantification of MFI (Mean Fluorescence Intensity). Mean ± SEM of n = 3 independent experiments. Paired one‐way ANOVA test results are shown.
Article Snippet: Phosphorylation of STAT5, using pSTAT5 antibody (STAT5(PY694), BD), was evaluated using a panel of antibodies to identify neutrophils [CD45+ (HI30, BD), CD66b + (G10F5, BD), CD15high (HI98, BD), CD14‐ (M5E2, Biolegend), Siglec8‐ (REA1045, Miltenyi)].
Techniques: Knockdown, Activation Assay, Transfection, Control, Marker, Flow Cytometry, Phospho-proteomics, Fluorescence