stat5 inhibitor sh-4-54  (ApexBio)

Journal: Infection and Immunity

Article Title: GM-CSF engages multiple signaling pathways to enhance pro-inflammatory cytokine responses in human monocytes during Legionella infection

doi: 10.1128/iai.00565-24

Figure Lengend Snippet: GM-CSF-dependent JAK2-STAT5 signaling is required to enhance inflammatory cytokine expression during Legionella infection. ( A ) THP-1 human monocytes were pretreated with PBS or rGM-CSF for 1 h. Cells were harvested at 6 h after infection to perform immunoblot analysis for phospho-STAT5, total STAT5, or β-actin as loading control. Lanes from one membrane have been cropped to depict the appropriate conditions, as indicated by the dashed lines. No changes were made to the original image during the editing. (B to C) THP-1 monocytes were pretreated with vehicle control, ( B ) the JAK2 inhibitor NVP-BSK805, or ( C ) the STAT5 inhibitor SH-4-54 for 1 h. Cells were then treated with PBS or rGM-CSF for 30–60 min and left uninfected or infected with L.p . Cells were harvested at 6 h post-infection (hpi) to measure IL1A , IL1B , and IL6 transcript levels by qPCR. Data represent the mean ± SEM of triplicate wells from at least two ( C ) or three ( B ) independent experiments. Data were analyzed by two-way ANOVA with Šidák’s multiple comparisons test; ****, P < 0.0001; ***, P < 0.001; **, P < 0.01; *, P < 0.05; ns, not significant.

Article Snippet: For inhibitor treatments, cells were pretreated 1 h before GM-CSF stimulation with either 250 nM IKK inhibitor (BMS-345541; Selleck Chemicals S8044), 2 μM JAK2 inhibitor (NVP-BSK805; Selleck Chemicals S2686), 5 μM STAT5 inhibitor (SH-4-54; Selleck Chemicals S7337), 10 μM PI-3K inhibitor (Ly294002; Selleck Chemicals S1105), 5 μM Akt inhibitor (MK-2206; Selleck Chemicals S1078), or 100 nM rapamycin (AY-22989; Selleck Chemicals S1039).

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

Journal: Infection and Immunity

Article Title: GM-CSF engages multiple signaling pathways to enhance pro-inflammatory cytokine responses in human monocytes during Legionella infection

doi: 10.1128/iai.00565-24

Figure Lengend Snippet: Multistep regulation of GM-CSF-enhanced cytokine gene expression in Legionella -infected human monocytes. Graphical model depicting the findings of this study. GM-CSF-enhanced cytokine production requires an initial bacterial PAMP that activates TLR-dependent NF-κB signaling in Legionella -infected human monocytes. Cytokine gene expression is then enhanced by GM-CSF-dependent JAK2/STAT5 signaling. PI-3K/Akt/mTORC1 signaling, glycolysis, and amino acid metabolism are required for upregulation of cytokine responses by GM-CSF. Dashed lines represent putative links between signaling pathways that need to be further dissected in future studies. Figure created in https://BioRender.com .

Article Snippet: For inhibitor treatments, cells were pretreated 1 h before GM-CSF stimulation with either 250 nM IKK inhibitor (BMS-345541; Selleck Chemicals S8044), 2 μM JAK2 inhibitor (NVP-BSK805; Selleck Chemicals S2686), 5 μM STAT5 inhibitor (SH-4-54; Selleck Chemicals S7337), 10 μM PI-3K inhibitor (Ly294002; Selleck Chemicals S1105), 5 μM Akt inhibitor (MK-2206; Selleck Chemicals S1078), or 100 nM rapamycin (AY-22989; Selleck Chemicals S1039).

Techniques: Gene Expression, Infection, Protein-Protein interactions

Journal: EMBO Molecular Medicine

Article Title: Disrupting TSLP–TSLP receptor interactions via putative small molecule inhibitors yields a novel and efficient treatment option for atopic diseases

doi: 10.1038/s44321-024-00085-3

Figure Lengend Snippet: ( A , B ) TSLP-activated primary CD4 + T cells were treated with different concentrations of the TSLP inhibitors BP75 and BP79 for 36 h, respectively. IL-13 and IL-4 secretion was quantified by ELISA ( n = 6), mean ± SEM. IC 50 values [μM] and slope were determined using GraphPad Prism software. ( C – E ) Cytotoxicity of BP79 in primary human keratinocytes ( n = 4), fibroblasts ( n = 5), and CD4 + T cells ( n = 5) was determined by MTT assay. ( F ) Inhibition of TSLP-mediated T-cell hyperproliferation. Primary CD4 + T cells were treated with TSLP inhibitors (or DMSO) and stimulated with TSLP for 1, 2 and 5 days. Cell proliferation was determined using a cell counter; n = 3. ( G ) Primary human CD4 + T cells and ( H ) keratinocytes were treated with BP79 and stimulated with TSLP. BP79 inhibited STAT6 and STAT3 activation in primary CD4 + T cells and primary keratinocytes, respectively ( n = 3). The band intensities were quantified using ImageJ ( n = 3). Phospho-Stat6 and Phospho-Stat3 expressions were normalized to Stat6 and Stat3 expressions, respectively. ( I ) Primary human myeloid dendritic cells were treated with BP79 and stimulated with TSLP for 24 h. OX-40L expression was detected using flow cytometry. ( J ) Primary human myeloid dendritic cells from three donors were treated with BP79 and stimulated with TSLP for 24 h. CCL17 expression was detected using ELISA ( n = 3). ( K ) Inhibition of dendritic cell-mediated Th2 cell differentiation. IL-13 secretion was determined using ELISA ( n = 3). Data information: All data are presented as mean ± SEM. n represents the number of biological replicates. Statistical analysis was performed using Student’s t test, * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001. .

Article Snippet: JAK2 inhibitor (AZD1480), Stat3/Stat5 inhibitor (SH-4-54) and Stat6 inhibitor (AS1517499) were purchased from Cayman Chemicals.

Techniques: Enzyme-linked Immunosorbent Assay, Software, MTT Assay, Inhibition, Activation Assay, Expressing, Flow Cytometry, Cell Differentiation

Journal: Cancer Biology & Therapy

Article Title: Low UBE4B expression increases sensitivity of chemoresistant neuroblastoma cells to EGFR and STAT5 inhibition

doi: 10.1080/15384047.2019.1647049

Figure Lengend Snippet:

Article Snippet: The STAT5 inhibitor SH-4-54 (ApexBio Technology, B4789) was purchased as a 10 mM stock solution in DMSO.

Techniques: MTT Assay, Proliferation Assay, Ubiquitin Proteomics, Conjugation Assay, Saline, Concentration Assay, Protein Array