gata 1  (Proteintech)

Journal: bioRxiv

Article Title: Inhibition of NLRP1 Inflammasome Activation by Tyrosine Kinase Inhibitors Restores Erythropoiesis in Diamond-Blackfan Anemia Syndrome

doi: 10.1101/2025.02.20.639294

Figure Lengend Snippet: (A) Schematic representation of the experimental design. (B-D) K562 cells were pretreated with 0.1 μM nilotinib (B-D), 0.1–1 μM imatinib, 0.1–1 μM dasatinib (B), 20–1000 nM ponatinib, or 100–1000 nM bosutinib (C) for 24 h and subsequently differentiated with 50 μM hemin for another 24 h. (B, C) Hemoglobin accumulation and protein levels of NLRP1, phosphorylated P38 (pP38), GATA1, ZAKα, and ACTB were assessed by Western blot. Immunoblots are representative of three independent experiments. The asterisk (*) denotes the GATA1 protein band. (D) CASP1 activity was quantified using FLICA-CASP1 staining and analyzed by flow cytometry. Representative histograms of CASP1 activity at different differentiation times are shown.

Article Snippet: The primary antibodies used were human GATA1 (#3535, Cell Signaling), human NLRP1 (#AF6788, R&D Systems), human ZAKα (#A301-993A, Bethyl Laboratories), human phosphoP38 (#MA5-15177, ThermoFisher Scientific) and ACTB-HRP (#sc-47778, Santa Cruz Biotechnology).

Techniques: Western Blot, Activity Assay, Staining, Flow Cytometry

Journal: bioRxiv

Article Title: Inhibition of NLRP1 Inflammasome Activation by Tyrosine Kinase Inhibitors Restores Erythropoiesis in Diamond-Blackfan Anemia Syndrome

doi: 10.1101/2025.02.20.639294

Figure Lengend Snippet: Primary human CD34 + HSPCs from healthy donors were differentiated with EPO in the presence of 0.1 µM imatinib (A-D), or 10 (E-H) and 1 nM (I-K) dasatinib from 3 to 7 days of culture. Cells were stained with anti-CD235A-APC (Glycophorin A) and anti-CD71-FITC (Transferrin Receptor), and erythroid differentiation was then analyzed by flow cytometry. The transcript levels of GATA1-dependent genes (B), caspase-1 activity determined with FAM FLICA (D,G) and the differentiation score calculated as the ratio between CD235A + /CD71 + (intermediate erythroid progenitors) and CD235A - /CD71 + (early erythroid progenitors) (D, H, K) at 7 dpd are shown. Data are shown as the mean ± SEM. P values were calculated using one-way ANOVA and Tukey’s multiple range test (D, H, K) or a Student’s t -test (C, G). ns, non-significant; *p<0.05; **p<0.01; ***p<0.01 and ****p<0.0001.

Article Snippet: The primary antibodies used were human GATA1 (#3535, Cell Signaling), human NLRP1 (#AF6788, R&D Systems), human ZAKα (#A301-993A, Bethyl Laboratories), human phosphoP38 (#MA5-15177, ThermoFisher Scientific) and ACTB-HRP (#sc-47778, Santa Cruz Biotechnology).

Techniques: Staining, Flow Cytometry, Activity Assay