Journal: Acta Pharmaceutica Sinica. B

Article Title: Macrophage P2Y 6 R activation aggravates psoriatic inflammation through IL-27-mediated Th1 responses

doi: 10.1016/j.apsb.2024.06.008

Figure Lengend Snippet: Macrophage P2Y 6 R mediated Th1 type psoriasis-like inflammation via p-JAK2/p-STAT1/T-bet signaling. (A) Flow diagram. (B, C) FACs and the statistical analysis of IFN γ + CD4 + T-cells ( n = 3). (D) IFN γ concentration after WT or P2Y 6 R −/− mice BMDM supernatant inducing Naïve CD4 + T-cells ( n = 4). (E) The mRNA expression of the Th1-associated cytokines, Ifnγ , Cxcl9 , Cxcl10, and Cxcl11 were detected by RT-qPCR ( n = 3). All mRNA levels were normalized to GAPDH. (F) Linear regression analysis indicated a positive correlation between P2ry6 and Stat1 in GSE181318 and GSE14905. (G) GSEA analysis of GSE50400 showed JNK/STAT signaling pathways were up-regulated in IMQ-induced mice. (H) Top 15 putative transcription factors based on ChEA3. (I, J) The protein expression levels of p-JAK2, p-STAT1, and T-bet in T cells. Normal protein levels were normalized to β -actin, the values for the phosphorylated forms of proteins were normalized to phosphorylation-independent levels of the same protein ( n = 4). (K) CD4 + Naïve T cells supernatant IFN γ concentration after WT or P2Y 6 R −/− mice BMDM supernatant inducing and being treated with or without RO8191 ( n = 4). (L) The proportion of IFN γ + CD4 + T cells of CD4 + Naïve T cells after WT or P2Y 6 R −/− mice BMDM supernatant inducing and being treated with or without RO8191 ( n = 3). (M, N) T-bet protein expression of CD4 + Naïve T cells after WT or P2Y 6 R −/− mice BMDM supernatant inducing and being treated with or without RO8191 ( n = 4). Data are expressed as mean ± SEM. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001.

Article Snippet: The following antibodies were used: PLC β (bs-6976R, Bioss, Beijing, China), PKC (ab181558, Abcam, Cambridge, UK), p-PKC (EP2730Y, Abcam), p38 (AF6456, Affinity, Jiangsu, China), p-p38 (AF4001, Affinity), ERK (bs-0022R, Bioss), p-ERK (bs-3016R, Bioss), p-JAK2 (A19629, Abclonal, Wuhan, China), JAK2 (AP0531, Abclonal), p-STAT1 (bs1657R, Bioss), STAT1 (10144-2-AP, Proteintech, Wuhan, China), p-JNK (bs1640R, Bioss), JNK (AF6318, Affinity), T-bet (14-5825-82, Abclonal), β -actin (T0022, Affinity).

Techniques: Concentration Assay, Expressing, Quantitative RT-PCR

Journal: medRxiv

Article Title: Polymorphism in IFNAR contributes to glucocorticoid response and outcome in ARDS and COVID-19

doi: 10.1101/2022.03.10.22272123

Figure Lengend Snippet: (A ) STAT1 expression in the lung after 4-day culture in the presence of IFN beta with or without hydrocortisone (HC). ( B) pSTAT1 expression in the same specimens as in A. ( C) Example photomicrographs showing higher STAT2 expression in a TT patient than in a CT patient and the effect of HC on its nuclear translocation. Most STAT2 remains in the cytoplasm of the CT patients, whereas nuclear expression is prominent in the TT patient. Indicated insets are shown in the bottom row. Arrows. ( D ) Combined results of all patients noting that two CT samples are excluded in the data as the patients were already under glucocorticoid treatment at the time of sample acquisition. Ns, not significant; *P<0.05; **P<0.01; and ***P<0.001

Article Snippet: The first stage antibodies were anti-alpha chain of the IFN alpha/beta receptor (St John’s Laboratory STJ112765) that was used 1:2000 and 1:5000 and anti-Stat1 (1:400, 9175S), anti-pStat1(1:100, 9167S) and anti-Stat2 (1:200, 72604S) all from Cell Signalling.

Techniques: Expressing, Translocation Assay

Journal: PLoS ONE

Article Title: Interferon-γ Produced by Microglia and the Neuropeptide PACAP Have Opposite Effects on the Viability of Neural Progenitor Cells

doi: 10.1371/journal.pone.0011091

Figure Lengend Snippet: NPCs were treated with 100 ng/ml IFNγ for different times and analyzed as described in . *p<0.05 and **p<0.001 comparing IFNγ treated vs. controls. ( A ) Immunoblot shows increased levels of phospho-STAT1 (p-Stat1) after 0.5 h. ( B ) Immunostaining showed the presence of p-Stat1 in the nucleus after 2 h treatment. Scale bar, 10 µm. ( C ) Cell viability was determined by the MTT assay as described in . Values are means ±SD, n = 4. ( D ) Dose response curve. Cells were treated for 48 h and cell viability determined. Values are means ±SD, n = 4. ( E ) Cells were treated for 48 h and the number of nestin positive NPCs was analyzed. Values are means ±SD, n = 4. ( F ) Neurospheres were dissociated and about 5000 cells per well were incubated for 3days in absence or presence of 100 ng/ml IFNγ. The number of secondary neurospheres formed was counted. Values are means ±SD, n = 4.

Article Snippet: Antibodies used were: cyclin D1 (1∶750) and Bax (1∶250) from Santa-Cruz (CA, USA), STAT1 (1∶400), p-STAT1 (1∶500); caspase-3 (1∶1000), cleaved caspase-3 (1∶1000); PARP (1∶2000), p53 (1∶1000), Bad (1∶1000), p-Bad (1∶500; phospho-Ser112), and Puma (1∶1000) all from Cell Signaling, Bcl2 (1∶400) and Bcl-xL (1; 1000) from BD Bioscience, IFNγR-2 (1∶3000; Abcam), p21 (1∶200; Millipore, MA, USA), p-vimentin (1∶2000; phospho-Ser55 Assay Designs, Ann Arbor, MI, USA), and β-actin (1∶2000, Sigma).

Techniques: Western Blot, Immunostaining, MTT Assay, Incubation

Journal: PLoS ONE

Article Title: Interferon-γ Produced by Microglia and the Neuropeptide PACAP Have Opposite Effects on the Viability of Neural Progenitor Cells

doi: 10.1371/journal.pone.0011091

Figure Lengend Snippet: NPCs were treated for 24 h with 100 ng/ml IFNγ alone or in conjunction with 100 ng/ml PACAP. ( A ) Immunoblot. Cells were treated for 2 h. PACAP did not influence levels of p-Stat1 induced by IFNγ. ( B ) RT-PCR. SOCS-1 and –3 levels are not influenced by PACAP. IFNγ upregulated SOCS1. ( C ) Immunoblots. IFNγ increased whilst PACAP decreased levels of PUMA. PACAP also decreased active caspase-3 (17 kDa band). Typical blot is shown and was repeated three times. ( D ) Immunoblots. PACAP increased levels of p-Bad. Typical blot is shown and was repeated three times. ( E ) Effects of the kinase inhibitors on cell viability measured by MTT. 1 µM Gö6976 reduced the beneficial effect of PACAP whereas 10 µM H89 had no effect. Values are means ±SD, n = 4. *** p<0.005 for IFNγ vs. controls, and *p<0.0 for IFNγ+PACAP vs. IFNγ, and for GÖ+ IFNγ+PACAP vs. IFNγ+PACAP. ( F ) BrdU labeling. PACAP increases cell proliferation significantly in control but not in IFNγ treated cells. Values are means±SD, n = 4. * p<0.05 for PACAP vs control cells, n = 3.

Article Snippet: Antibodies used were: cyclin D1 (1∶750) and Bax (1∶250) from Santa-Cruz (CA, USA), STAT1 (1∶400), p-STAT1 (1∶500); caspase-3 (1∶1000), cleaved caspase-3 (1∶1000); PARP (1∶2000), p53 (1∶1000), Bad (1∶1000), p-Bad (1∶500; phospho-Ser112), and Puma (1∶1000) all from Cell Signaling, Bcl2 (1∶400) and Bcl-xL (1; 1000) from BD Bioscience, IFNγR-2 (1∶3000; Abcam), p21 (1∶200; Millipore, MA, USA), p-vimentin (1∶2000; phospho-Ser55 Assay Designs, Ann Arbor, MI, USA), and β-actin (1∶2000, Sigma).

Techniques: Western Blot, Reverse Transcription Polymerase Chain Reaction, Labeling, Control

Journal: Cell death & disease

Article Title: PLA2G7 promotes immune evasion of bladder cancer through the JAK-STAT-PDL1 axis.

doi: 10.1038/s41419-025-07593-1

Figure Lengend Snippet: Fig. 5 PLA2G7 regulates the JAK–STAT pathway through the phosphorylations of STAT1 and STAT3. A Volcano plot of genes differentially expressed in sh-NC and sh-PLA2G7 cells following stimulation with IFN-γ. B KEGG enrichment analysis for significant down-regulated genes. C The correlation of PLA2G7 with dysfunction score in TCGA bladder cancer cohort. D The correlation of PLA2G7 with exhausted score in TCGA bladder cancer cohort. E The correlation of PLA2G7 with PD-L1 mRNA level in TCGA bladder cancer cohort. F The correlation of PLA2G7 with PD-L1 expression in bladder cancer tissues analyzed by RT-qPCR. G Western blot analysis showing PLA2G7, JAK2, STAT1, p-STAT1, STAT3, p-STAT3, IRF1 and PD-L1 levels in sh-NC and sh-PLA2G7 cells.

Article Snippet: The antibodies used in this study were as follows: PLA2G7 (Proteintech, 15526–1-AP), JAK2 (Abcam, ab108596), STAT3 (Abcam, ab68153), p-STAT3 (Abcam, ab76315), GAPDH (Proteintech, 60004-1-Ig), STAT1 (Proteintech, 10144-2-AP), p-STAT1 (Cell Signaling Technology, 9167), IRF1 (Proteintech, 11335-1-AP), PD-L1 (Proteintech, 66248-1-Ig), ETS1 (Proteintech, 66598-1-Ig), CD4 (Abcam, ab237722), and CD8 (Abcam, ab217344).

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: Cell death & disease

Article Title: PLA2G7 promotes immune evasion of bladder cancer through the JAK-STAT-PDL1 axis.

doi: 10.1038/s41419-025-07593-1

Figure Lengend Snippet: Fig. 6 PLA2G7 inhibitor darapladib suppresses bladder cancer immune evasion. A MTS assays of 5637 cell viability treated with placebo and darapladib. B RT-qPCR analysis of the mRNA levels of PD-L1 in bladder cancer cells treated with placebo and darapladib. C Western blot analysis showing p-STAT1, p-STAT3 and PD-L1 levels in bladder cancer cells treated with placebo and darapladib. D Tumor growth curves for immunocompetent BALB/C mice injected with bladder cancer cells treated with placebo and darapladib. E Tumor weights for immunocompetent BALB/C mice injected with bladder cancer cells treated with placebo and darapladib.

Article Snippet: The antibodies used in this study were as follows: PLA2G7 (Proteintech, 15526–1-AP), JAK2 (Abcam, ab108596), STAT3 (Abcam, ab68153), p-STAT3 (Abcam, ab76315), GAPDH (Proteintech, 60004-1-Ig), STAT1 (Proteintech, 10144-2-AP), p-STAT1 (Cell Signaling Technology, 9167), IRF1 (Proteintech, 11335-1-AP), PD-L1 (Proteintech, 66248-1-Ig), ETS1 (Proteintech, 66598-1-Ig), CD4 (Abcam, ab237722), and CD8 (Abcam, ab217344).

Techniques: Quantitative RT-PCR, Western Blot, Injection

Journal: Cell death & disease

Article Title: PLA2G7 promotes immune evasion of bladder cancer through the JAK-STAT-PDL1 axis.

doi: 10.1038/s41419-025-07593-1

Figure Lengend Snippet: Fig. 7 PLA2G7 overexpression promotes PD-L1 expression and tumor immune evasion. A MTS cell viability assay of OE-NC and OE-PLA2G7 cells. B Surface PD-L1 protein levels analyzed by FACS in OE-NC and OE-PLA2G7 cells. C Results from the T cell killing assays of OE-NC and OE- PLA2G7 cells. D Western blot analysis of PLA2G7, p-STAT1, p-STAT3 and PD-L1 levels in OE-NC and OE-PLA2G7 cells with or without STAT1 and STAT3 knockdown.

Article Snippet: The antibodies used in this study were as follows: PLA2G7 (Proteintech, 15526–1-AP), JAK2 (Abcam, ab108596), STAT3 (Abcam, ab68153), p-STAT3 (Abcam, ab76315), GAPDH (Proteintech, 60004-1-Ig), STAT1 (Proteintech, 10144-2-AP), p-STAT1 (Cell Signaling Technology, 9167), IRF1 (Proteintech, 11335-1-AP), PD-L1 (Proteintech, 66248-1-Ig), ETS1 (Proteintech, 66598-1-Ig), CD4 (Abcam, ab237722), and CD8 (Abcam, ab217344).

Techniques: Over Expression, Expressing, Viability Assay, Western Blot, Knockdown

Journal: ACS Omega

Article Title: Rhoifolin Attenuates Concanavalin A-Induced Autoimmune Hepatitis in Mice via JAKs/STATs Mediated Immune and Apoptotic Processes

doi: 10.1021/acsomega.4c07915

Figure Lengend Snippet: Molecular docking and dynamic simulations of ROF and JAKs/STATs signaling molecules (A) The binding modes of ROF toward JAK2, JAK3, STAT1, and STAT3. (B–F) The dynamic simulations of JAK2-ROF and JAK3-ROF complexes. The HBond (B), RMSD curves (C), RMSF values (D), Rg values (E), and SASA (F) were analyzed by GROMACS.

Article Snippet: The antibodies for cleaved-caspase-3, cleaved-caspase-9, Bax, Bcl-2, JAK2, p-JAK2, JAK3, p-JAK3, IL-6, STAT1, p-STAT1, STAT3, p-STAT3, and BNIP3 were diluted at a ratio of 1:1000, and β-actin (#BA2305, BOSTER) was diluted at a ratio of 1:5000 in the primary antibody dilution buffer.

Techniques: Binding Assay

Journal: ACS Omega

Article Title: Rhoifolin Attenuates Concanavalin A-Induced Autoimmune Hepatitis in Mice via JAKs/STATs Mediated Immune and Apoptotic Processes

doi: 10.1021/acsomega.4c07915

Figure Lengend Snippet: Effects of ROF on the JAK2/3-STAT1/3 signaling pathways in Con A-induced hepatitis (A) The phosphorylated levels of JAK2, STAT1, and STAT3 were measured in hepatic tissues using IHC staining ( n = 3; scale bar = 40 μm). (B) The IODs of each protein were detected by using ImageJ. (C) The JAK2/3-STAT1/3 signaling molecules were measured in hepatic tissues with Western blot analysis ( n = 3). The gray value analysis was standardized using the intensity of β-actin and is depicted in bar graphs. The results are presented as mean ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.001 vs the Con A group.

Article Snippet: The antibodies for cleaved-caspase-3, cleaved-caspase-9, Bax, Bcl-2, JAK2, p-JAK2, JAK3, p-JAK3, IL-6, STAT1, p-STAT1, STAT3, p-STAT3, and BNIP3 were diluted at a ratio of 1:1000, and β-actin (#BA2305, BOSTER) was diluted at a ratio of 1:5000 in the primary antibody dilution buffer.

Techniques: Protein-Protein interactions, Immunohistochemistry, Western Blot

Journal: ACS Omega

Article Title: Rhoifolin Attenuates Concanavalin A-Induced Autoimmune Hepatitis in Mice via JAKs/STATs Mediated Immune and Apoptotic Processes

doi: 10.1021/acsomega.4c07915

Figure Lengend Snippet: Effects of ROF on Th1/Th17 cells differentiation in vitro (A) The purity of isolated CD4 + T cells was measured by flow cytometry. (B) The viability of CD4 + T cells following 5–20 μM ROF treatments for 24 h was assessed utilizing the CCK-8 assay. (C, D) CD4 + T cells were pre-exposed for 1 h to ROF (20 μM), TOF (1 μM), or ROF (20 μM) + TOF (1 μM), followed by stimulation with Con A for 24 h. The levels of p-JAK2, p-JAK3, p-STAT1, and p-STAT3 were monitored by Western blotting and normalized to each protein’s total concentration in the graphs. (E) The generation of IFN-γ or IL-17 in the cellular supernatants of Th1 or Th17 subtypes were measured using ELISA. (F) The gene expression levels of T-bet or RORγt in differentiated T cells were evaluated by PCR. The values are presented as mean ± SEM ( n = 3). ** P < 0.01, *** P < 0.001 vs the Con A group. # P < 0.05, ## P < 0.01, ### P < 0.001 vs the ROF only group or TOF only group.

Article Snippet: The antibodies for cleaved-caspase-3, cleaved-caspase-9, Bax, Bcl-2, JAK2, p-JAK2, JAK3, p-JAK3, IL-6, STAT1, p-STAT1, STAT3, p-STAT3, and BNIP3 were diluted at a ratio of 1:1000, and β-actin (#BA2305, BOSTER) was diluted at a ratio of 1:5000 in the primary antibody dilution buffer.

Techniques: In Vitro, Isolation, Flow Cytometry, CCK-8 Assay, Western Blot, Concentration Assay, Enzyme-linked Immunosorbent Assay, Gene Expression

Journal: ACS Omega

Article Title: Rhoifolin Attenuates Concanavalin A-Induced Autoimmune Hepatitis in Mice via JAKs/STATs Mediated Immune and Apoptotic Processes

doi: 10.1021/acsomega.4c07915

Figure Lengend Snippet: Effects of ROF on primary hepatocytes apoptosis in vitro (A) Primary hepatocytes were pretreated with ROF (5, 10, 20 μM) for 6 h and treated with or without Con A (10 μg/mL) for 24 h, the cell viability of each group was analyzed by CCK-8 assay. (B) Flow cytometry analysis of apoptosis in primary hepatocytes. (C) Primary hepatocytes were pretreated with ROF (20 μM), TOF (1 μM), or ROF (20 μM) + TOF (1 μM) for 6 h, and subsequently triggered with Con A (10 μg/mL) for 24 h. The IL-6, p-JAK2, p-STAT1, p-STAT3, BNIP3, and Bax expression levels were evaluated by using Western blotting. (D) The quantification and standardization of the protein expression were performed using ImageJ. The results are presented as mean ± SEM of three independent experiments. ** P < 0.01, *** P < 0.001 vs the Con A group. # P < 0.05, ## P < 0.01 vs the ROF only group or TOF only group.

Article Snippet: The antibodies for cleaved-caspase-3, cleaved-caspase-9, Bax, Bcl-2, JAK2, p-JAK2, JAK3, p-JAK3, IL-6, STAT1, p-STAT1, STAT3, p-STAT3, and BNIP3 were diluted at a ratio of 1:1000, and β-actin (#BA2305, BOSTER) was diluted at a ratio of 1:5000 in the primary antibody dilution buffer.

Techniques: In Vitro, CCK-8 Assay, Flow Cytometry, Expressing, Western Blot

Journal: Cell reports

Article Title: Janus Kinase 1 Plays a Critical Role in Mammary Cancer Progression

doi: 10.1016/j.celrep.2018.10.063

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit polyclonal, pY-STAT1 , Origene , Cat# TA309955.

Techniques: Recombinant, Sample Prep, Software

Journal:

Article Title: Signaling of short- and long-term regulation of intestinal epithelial type 1 Na + /H + exchanger by interferon- γ

doi: 10.1038/sj.bjp.0706167

Figure Lengend Snippet: Abundance of total STAT1, phospho-STAT1 and phospho-p38 MAPK in Caco-2 cells treated with vehicle and IFN-γ (1000 U ml−1) or anisomycin (100 nM) for 1, 3 and 24 h.

Article Snippet: Total and phosphorylated STAT1 protein levels in Caco-2 cells exposed to IFN- γ (1000 U ml −1 ) and anisomycin (100 n M ) for 1, 3 and 24 h were analyzed using the PhosphoPlus® Stat1 (Tyr701) antibody kit (Cell Signaling TechnologyTM) according to the manufacturer's protocol.

Techniques:

Journal:

Article Title: Signaling of short- and long-term regulation of intestinal epithelial type 1 Na + /H + exchanger by interferon- γ

doi: 10.1038/sj.bjp.0706167

Figure Lengend Snippet: Effect of EGCG (20 μM), SB 203580 (10 μM) and PKC downregulation (PKC dr; overnight treatment with 100 nM PDBu) on the abundance of STAT1 and phospho-STAT1 in Caco-2 cells treated for 24 h in the absence (vehicle) and presence of IFN-γ (1000 U ml−1).

Article Snippet: Total and phosphorylated STAT1 protein levels in Caco-2 cells exposed to IFN- γ (1000 U ml −1 ) and anisomycin (100 n M ) for 1, 3 and 24 h were analyzed using the PhosphoPlus® Stat1 (Tyr701) antibody kit (Cell Signaling TechnologyTM) according to the manufacturer's protocol.

Techniques: