Journal: Cell death & disease

Article Title: Inhibition of STAT5A promotes osteogenesis by DLX5 regulation.

doi: 10.1038/s41419-018-1184-7

Figure Lengend Snippet: Fig. 1 Suppressed osteoblastic differentiation of hBMSCs by overexpression of STAT5A. a Protein levels of STAT5A, STAT5B, DLX5, and RUNX2 during osteogenesis of hBMSCs. b Alizarin Red S staining demonstrating the effect of STAT5 overexpression on the osteogenesis at day 14. Scale bar, 60 μm. c Quantification of Alizarin Red S staining. d Western blot analysis of protein expression of osteogenic master genes, DLX5 and Runx2 after STAT5 overexpression. GAPDH was used as a control. e Quantification of DLX5 and RUNX2 protein levels. f Gene expression of DLX5 and expression of downstream genes of DLX5 after 4 days with STAT5 overexpression in hBMSCs using real-time PCR. All mRNA and protein levels were normalized with GAPDH. Each experiment was performed in triplicate (n = 3). All error bars indicate ± SEM. *P < 0.01; **P < 0.01; ***P < 0.001

Article Snippet: A STAT5 inhibitor which inhibits phosphorylation of STAT5 via binding to the SH2 domain of STAT5 was purchased from Santa Cruz Biotechnology (sc-355979, Dallas, TX, USA).

Techniques: Over Expression, Staining, Western Blot, Expressing, Control, Gene Expression, Real-time Polymerase Chain Reaction

Journal: Cell death & disease

Article Title: Inhibition of STAT5A promotes osteogenesis by DLX5 regulation.

doi: 10.1038/s41419-018-1184-7

Figure Lengend Snippet: Fig. 2 Induced osteoblastic differentiation through increased DLX5 expression by inhibition of STAT5A in hBMSCs. a Alizarin Red S staining to detect suppression of STAT5A and STAT5B on osteogenesis using targeted siRNA. Staining was performed on day 14 of osteogenesis. Scale bar, 30 μm. b Quantification of Alizarin Red S staining. c Western blot analysis of DLX5 protein expression after silencing of STAT5A in hBMSCs. d Quantification of DLX5 protein level compared with GAPDH expression level. e Alizarin Red S staining for the effect of STAT5 inhibitor during osteogenesis of hBMSCs. STAT5 inhibitor (sc-355797) was used at concentrations of 0, 2, 6, and 10 μM. Staining was performed at day 14 of osteogenesis. Scale bar, 30 μm. f Quantification of Alizarin Red S staining. g Analysis of DLX5 mRNA level after 10 μM STAT5 inhibitor treatment. mRNA level was checked on day 5 of osteogenesis. h Western blot analysis of DLX5 protein expression after STAT5 inhibitor treatment. Protein level was detected on day 5 of osteogenesis. Each experiment was performed in triplicate (n = 3). All error bars indicate ±SEM. *P < 0.01; **P < 0.01; ***P < 0.001

Article Snippet: A STAT5 inhibitor which inhibits phosphorylation of STAT5 via binding to the SH2 domain of STAT5 was purchased from Santa Cruz Biotechnology (sc-355979, Dallas, TX, USA).

Techniques: Expressing, Inhibition, Staining, Western Blot

Journal: bioRxiv

Article Title: The STAT5-IRF4-BATF pathway drives heightened epigenetic remodeling in naïve CD4 + T cell responses of older adults

doi: 10.1101/2021.08.27.457205

Figure Lengend Snippet: A . Heatmap representation of top 35 most variable TFs based on ChromVAR deviation scores of the ATAC-seq data as described in . Each column is the score median of the four young (left) and the four older (right) individuals. Stars indicate the representative TFs in B. B . ChromVAR deviation scores for representative TFs from are shown as mean+SEM. C . Phosphorylated STAT5 (pSTAT5), IRF4 and BATF were measured at indicated times after TCR stimulation. Western blots are shown for one pair of one young and one older individual representative of three experiments. D . Representative blots of pSTAT5 measured at 6 hours and IRF and BATF at 24 hours after low and higher intensity stimulation (top). Summary results for 4 to 6 young and older adults after median intensity stimulation shown as mean±SEM (bottom). Data were analyzed with unpaired t-test. **p < 0.01, ***p < 0.001. E . Naïve CD4 + T cells from older adults were activated for 48 hours with indicated αCD3 beads in the presence of DMSO or a STAT5 inhibitor. Western blots for pSTAT5, IRF4 and BATF are from 2 experiments representative of 5.

Article Snippet: Naïve CD4 + T cells from subjects 13-16 were stimulated with polystyrene beads (αCD3=1 μg/mL) for 48 h in presence of either PBS, 5 μg/mL of αCD25 and αCD122 Ab, DMSO or 50 μM of STAT5 inhibitor (Cayman chemical, #15784) in DMSO.

Techniques: Western Blot

Journal: bioRxiv

Article Title: The STAT5-IRF4-BATF pathway drives heightened epigenetic remodeling in naïve CD4 + T cell responses of older adults

doi: 10.1101/2021.08.27.457205

Figure Lengend Snippet: A . Four older individuals were activated in the presence of either DMSO control, STAT5 inhibitor, PBS control or IL-2R-blocking antibodies for 48 hours and then subjected to ATAC-seq. Data of the four treatments were subjected to PCA together with the 48 hours samples from the four young and four older individuals without any treatment described in and . Results for PC1 are shown. B . TF binding motif enrichment of PC1 loading sites in A. C . LogFC of differential peaks between STAT5 inhibition and DMSO treatment (p<0.05) are plotted against average peak size. Peaks that are also differentially accessible between young and older individuals are indicated in dark red. D . Accessibility tracks of representative genes. Magenta-shaded areas indicate peaks that are more open in older adults or with DMSO treatment; cyan indicates peaks more open in young adults or with STAT5 inhibition treatment. E . ChIP-qPCR with pSTAT5 antibody of 48-hour activated naïve CD4 + T cells treated with either PBS or IL-2R blocking antibodies. Data were analyzed with paired t-test. *p < 0.05. F . pSTAT5, IRF4 and BHLHE40 were measured by Western blot in 48-hour activated naïve CD4 + T cells for each treatment conditions. G . Naïve CD4 + T cells from 6 older adults were activated with 1 μg/mL αCD3-coated beads for five days together with DMSO or STAT5 inhibitor. TCF1 and BLIMP1 expression was measured by flow cytometry. The MFI ratio of TCF1 to BLIMP1 was plotted. Data were analyzed with paired t-test. *p < 0.05. H . Naïve CD4 + T cells from 4 young and 5 older adults were activated for five days. TCF1 and BLIMP1 protein expression was assayed as in . The MFI ratio of TCF1 to BLIMP1 is plotted. Data were analyzed with unpaired t-test. *p < 0.05.

Article Snippet: Naïve CD4 + T cells from subjects 13-16 were stimulated with polystyrene beads (αCD3=1 μg/mL) for 48 h in presence of either PBS, 5 μg/mL of αCD25 and αCD122 Ab, DMSO or 50 μM of STAT5 inhibitor (Cayman chemical, #15784) in DMSO.

Techniques: Control, Blocking Assay, Binding Assay, Inhibition, ChIP-qPCR, Western Blot, Expressing, Flow Cytometry

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

Journal: International Journal of Molecular Sciences

Article Title: Rationale for a Combination Therapy with the STAT5 Inhibitor AC-4-130 and the MCL1 Inhibitor S63845 in the Treatment of FLT3-Mutated or TET2-Mutated Acute Myeloid Leukemia

doi: 10.3390/ijms22158092

Figure Lengend Snippet: Dose-response curves of AML cell lines. AML cells were treated with the STAT5 inhibitor AC-4-130 ( A , B ) or the BCL2 inhibitor venetoclax ( C , D ) for 20 hours in the absence ( A , C ) or in the presence ( B , D ) of HS-5 stroma cells. Cell-viability data are average values of multiple repeat measurements per dosage. The standard deviation was 3–6%.

Article Snippet: The STAT5 inhibitor AC-4-130 was purchased at Aobious, Inc. Gloucester, MA, USA.

Techniques: Standard Deviation

Journal: International Journal of Molecular Sciences

Article Title: Rationale for a Combination Therapy with the STAT5 Inhibitor AC-4-130 and the MCL1 Inhibitor S63845 in the Treatment of FLT3-Mutated or TET2-Mutated Acute Myeloid Leukemia

doi: 10.3390/ijms22158092

Figure Lengend Snippet: Schematic representation of STAT5 signaling pathways in myeloid cells. STAT5 can be activated by FLT3-ITD and by cytokine receptor signaling via Janus tyrosine kinases (JAKs). FLT3-ITD is a constitutively active growth factor receptor signaling via PI3K-AKT , via RAS-MEK-ERK , and via STAT5 , leading to cell growth and proliferation via p53 inhibition and MCL1 induction. Hematopoietic cytokine receptor signaling is largely mediated by JAKs and their downstream transcription factors, the STATs . Mutations in the tumor suppressors IDH1/2 and TET2 may be functionally equivalent . The tumor suppressor FoxP3 may inhibit AKT and its downstream target MCL-1 [ , ]. Oncogenic functions are indicated in red, tumor suppressor functions in green, and chemical inhibitors in pink.

Article Snippet: The STAT5 inhibitor AC-4-130 was purchased at Aobious, Inc. Gloucester, MA, USA.

Techniques: Protein-Protein interactions, Inhibition