Journal: Journal of Cancer

Article Title: Lasalocid inhibits melanoma by down-regulating FOXM1 through PI3K/AKT and JNK/P38 MAPK pathways

doi: 10.7150/jca.101798

Figure Lengend Snippet: Lasalocid inhibits the proliferation of melanoma cells by activating JNK/P38 MAPK pathway and down-regulating FOXM1. (A,B) The protein expression levels of JNK, P-JNK, P38 and P-P38 in melanoma cells treated with lasalocid for 24 h were detected by Western blot. (C,D) The expression of P38, P-P38 and FOXM1 proteins and the proliferation ability of melanoma cells after inhibiting the P38 MAPK pathway by SB202190 (20 μM, 24 h). (E,F) The expression of JNK, P-JNK and FOXM1 proteins and the proliferation ability of melanoma cells after inhibiting JNK MAPK pathway by SP600125 (20 μM, 24 h). (Data are mean±SD * P <0.05, ** P <0.01, *** P <0.001, **** P <0.0001.

Article Snippet: Lasalocid, SP600125, SB202190, and IGF1 proteins were purchased from MedChemExpress (MCE, USA).

Techniques: Expressing, Western Blot

Journal: Stem Cell Reports

Article Title: Chemically defined and growth factor-free system for highly efficient endoderm induction of human pluripotent stem cells

doi: 10.1016/j.stemcr.2024.11.012

Figure Lengend Snippet: Development of a fully synthetic system for highly efficient DE induction from hPSCs (A) Immunofluorescence analysis of DE markers SOX17 and FOXA2 on hESCs differentiated in E5 or B27 supplement-containing culture medium from 3 days as illustrated by the upper schematic ( n = 6 biologically independent experiments). CH, CHIR99021. Scale bars, 50 μm. (B) Percentage of SOX17 + /FOXA2 + DE, cell yields, and survival rate of cells under the indicted conditions at D3 ( n = 6 biologically independent experiments). A minus mark demonstrates withdrawal of the indicated component. I, insulin; T, transferrin; S, sodium selenite; V, vitamin C. (C and D) Representative (C) and quantitative (D) immunofluorescence analysis of SOX17 and FOXA2 on hESCs differentiated on the indicated matrix substrates for 3 days ( n = 6 biologically independent experiments). Scale bars, 50 μm. (E and F) Workflow (E) and high-throughput small screening results (F) for DE inducers in conjunction with 3C. Activin-A and DMSO serve as positive or negative controls, respectively. Chemical structure of the leading hit LDN-193189 is shown. (G) Quantitative reverse-transcription PCR (RT-qPCR) analyses of the expression of key DE transcripts ( n = 9 biologically independent experiments). (H) Immunofluorescence analysis of DE markers SOX17, FOXA2, GATA4, and GATA6 with quantification of the SOX17 + /FOXA2 + and GATA4 + /GATA6 + rations on D3 DE induced by 4C illustrated by the upper schematic ( n = 6 biologically independent experiments). Scale bars, 50 μm. (I) Flow cytometric analyses of the DE marker protein CXCR4 in 4C-DE at D3 ( n = 6 biologically independent experiments). Data are represented as mean ± SE. ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; n.s., no significant.

Article Snippet: 3 μM CHIR99021 (TargetMol, T2301) and 100 ng mL-1 AA (R&D, 338-AC) were added from days 0–1 and days 0–3, respectively.

Techniques: Immunofluorescence, High Throughput Screening Assay, Reverse Transcription, Quantitative RT-PCR, Expressing, Marker

Journal: Stem Cell Reports

Article Title: Chemically defined and growth factor-free system for highly efficient endoderm induction of human pluripotent stem cells

doi: 10.1016/j.stemcr.2024.11.012

Figure Lengend Snippet: Formulation and price of components of medium for AA- or 4C-based method

Article Snippet: 3 μM CHIR99021 (TargetMol, T2301) and 100 ng mL-1 AA (R&D, 338-AC) were added from days 0–1 and days 0–3, respectively.

Techniques: Formulation, Concentration Assay

Journal: EMBO Reports

Article Title: Trichoplein binds PCM 1 and controls endothelial cell function by regulating autophagy

doi: 10.15252/embr.201948192

Figure Lengend Snippet: A Representative images for p62 of control and TCHP knock‐down HUVECs treated with BAY11‐7082 (300 nM) or TYRPHOSTIN AG1288 (300 nM) or SB202190 (300 nM) or vehicle (DMSO) for 48 h. Scale bars, 50 μm. B, C (B) Expression of IL‐6, IL‐8 and IL‐1β (one‐way ANOVA; IL‐8: ** P = 0.008 vs. control DMSO; # P = 0.0198 (BAY) # P = 0.01 (AG) vs. shTCHP DMSO; IL‐6: ** P = 0.0023 vs. control DMSO; # P = 0.0063 (BAY) # P = 0.0108 (AG) vs. shTCHP DMSO; IL‐1β: ** P = 0.0011 vs. control DMSO; # P = 0.0004 (BAY) # P = 0.0015 (AG) vs. shTCHP DMSO) and (C), migration speed was measured (one‐way ANOVA; ** P = 0.0003 vs. control DMSO; # P = 0.0451 vs. shTCHP DMSO). C Western blot analysis for anti‐phosphor‐NF‐κB (S536), total NF‐κB, in TCHP knock‐down and control cells treated with BAY or vehicle. Below panel: quantification (one‐way ANOVA; ** P = 0.0014 vs. control DMSO; # P = 0.0293 vs. shTCHP DMSO). D p62 expression in TCHP knock‐down and control cells treated with BAY or vehicle (one‐way ANOVA; ** P = 0.0004 vs. control DMSO; # P = 0.0243 vs. shTCHP DMSO). E p62 expression in TCHP knock‐down and control cells treated with siRELA or control (one‐way ANOVA; ** P = 0.0023 vs. control siRNA; # P = 0.0127 vs. shTCHP siRELA). F ChIP‐qPCR analysis confirms the of NF‐κB p65 enrichment to IκBα and p62 promoter in TCHP knock‐down cells (unpaired t ‐test; ** P = 0.0057 and ** P = 0.0037 vs. control, respectively). Data information: Statistical analyses were performed on at least three independent experiments. Data are represented as mean ± SD.

Article Snippet: Reagents and doses were as follows : LysoTracker (Thermo Fisher), bafilomycin A1 (Sigma, 200 nM), torin‐1 (CST, 10 μM), BAY11‐7082 (Abcam, 300 nM), TYRPHOSTIN AG1288 (Abcam, 300 nM), SB202190 (Tocris, 300 nM), cycloheximide (Sigma, 2 μM) and MG132 (Sigma, 2 μM).

Techniques: Control, Knockdown, Expressing, Migration, Western Blot, ChIP-qPCR

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Transforming Growth Factor-β–Induced Expression of the Apolipoprotein E Gene Requires c-Jun N-Terminal Kinase, p38 Kinase, and Casein Kinase 2

doi: 10.1161/01.atv.0000220383.19192.55

Figure Lengend Snippet: Figure 1. The TGF-–induced expression of apoE is prevented by inhibitors of the JNK and p38 kinase pathways. THP-1 cells or primary cultures of human monocytes (A and B, respectively) were incubated in the absence or the presence of TGF-1 (30 ng/mL) without or with the indicated inhibitors (12 hours for experiments with curcumin and 24 hours for SB202190, SB203580, or SP600125). The concentration of the inhibitors for the experiment in A are shown on the figure, and those for B were at 10 mol/L. An equal amount of protein or RNA was then subjected to Western blot analysis and RT-PCR, respec- tively (A and B, respectively). The results shown are representa- tive of 2 independent experiments.

Article Snippet: Curcumin, Apigenin, SB202190, and SB203580 were from Merck Biosciences (Calbiochem), SP600125 was from Affiniti, cytokines from Peprotech, and the in vitro JNK kit from Cell Signaling Technology.

Techniques: Expressing, Incubation, Concentration Assay, Western Blot, Reverse Transcription Polymerase Chain Reaction

Journal: Arteriosclerosis, Thrombosis, and Vascular Biology

Article Title: Transforming Growth Factor-β–Induced Expression of the Apolipoprotein E Gene Requires c-Jun N-Terminal Kinase, p38 Kinase, and Casein Kinase 2

doi: 10.1161/01.atv.0000220383.19192.55

Figure Lengend Snippet: Figure 4. The action of JNK and p38 kinase converges on c-Jun expression and AP-1 binding. THP-1 monocytes were incubated for 24 hours in the absence or the presence of TGF-1 (TGF-; 30 ng/mL) alone or with SB202190 or SP600125 (both at 50 mol/L) as indicated. Cellular proteins were then subjected to either Western blot analysis with anti- sera against c-Jun or -actin as indicated (A) or EMSA using a consensus AP-1 recognition sequence (B). Competition EMSAs were performed with a 500-fold molar excess of AP-1 or NF-1 binding sites as shown. B and F show the position of the DNA– protein complexes and free probe, respectively. The results shown are representative of 3 independent experiments.

Article Snippet: Curcumin, Apigenin, SB202190, and SB203580 were from Merck Biosciences (Calbiochem), SP600125 was from Affiniti, cytokines from Peprotech, and the in vitro JNK kit from Cell Signaling Technology.

Techniques: Expressing, Binding Assay, Incubation, Western Blot, Sequencing

Journal: Cells

Article Title: Mycobacterium tuberculosis Fatty Acyl-CoA Synthetase fad D33 Promotes Bacillus Calmette-Guérin Survival in Hostile Extracellular and Intracellular Microenvironments in the Host.

doi: 10.3390/cells12222610

Figure Lengend Snippet: Figure 5. Expression of p38, ERK1/2, and JNK. THP-1 cells were infected with B1033, B1033C, and wild-type BCG (Ctrl-BCG) strains at an MOI of 10. (a) Western blot analysis of p38, phosphorylated p38 (p-p38), ERK1/2, phosphorylated ERK1/2 (p-ERK), JNK, and phosphorylated JNK (p-JNK) expression 12 and 24 h PI. (b–d) Ratios of the levels of phosphorylated to unphosphorylated protein for p38 (b), ERK1/2 (c), and JNK (d). *** p < 0.001.

Article Snippet: PMA-differentiated THP-1 cells were seeded in 12-well plates at a density of 1 × 106 cells per well and cultured for 12 h. Then, the cells were infected with wildtype, B1033, or B1033C BCG strain at an MOI of 10 for 2 h. For inhibition assay, PMAdifferentiated THP-1 cells were pretreated with a p38 inhibitor (SB202190, 10 μM, MedChemExpress, Monmouth Junction, NJ, USA) for 2 h and infected with wild-type, B1033, Cells 2023, 12, 2610 5 of 14 or B1033C BCG strain.

Techniques: Expressing, Infection, Western Blot

Journal: Cells

Article Title: Mycobacterium tuberculosis Fatty Acyl-CoA Synthetase fad D33 Promotes Bacillus Calmette-Guérin Survival in Hostile Extracellular and Intracellular Microenvironments in the Host.

doi: 10.3390/cells12222610

Figure Lengend Snippet: Figure 7. IL-1β, IL-6, and TNF-α expression in THP-1 cells after p38 inhibition. PMA-differentiated THP-1 macrophages were treated with a p38 inhibitor (SB202190, 10 µM) or dimethyl sulfoxide (DMSO) for 2 h and then infected with wild-type (BCG), B1033, and B1033C strains at an MOI of 10. IL-1β (a), IL-6 (b), and TNF-α (c) expressions were detected via ELISA 72 h PI. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: PMA-differentiated THP-1 cells were seeded in 12-well plates at a density of 1 × 106 cells per well and cultured for 12 h. Then, the cells were infected with wildtype, B1033, or B1033C BCG strain at an MOI of 10 for 2 h. For inhibition assay, PMAdifferentiated THP-1 cells were pretreated with a p38 inhibitor (SB202190, 10 μM, MedChemExpress, Monmouth Junction, NJ, USA) for 2 h and infected with wild-type, B1033, Cells 2023, 12, 2610 5 of 14 or B1033C BCG strain.

Techniques: Expressing, Inhibition, Infection, Enzyme-linked Immunosorbent Assay

Journal: Drug Design, Development and Therapy

Article Title: Induction of apoptosis and autophagy via sirtuin1- and PI3K/Akt/mTOR-mediated pathways by plumbagin in human prostate cancer cells

doi: 10.2147/DDDT.S75976

Figure Lengend Snippet: Crosstalk between apoptosis and autophagy in PC-3 and DU145 cells. Notes: Cells were treated with PLB or in combination with 10 μM SB202190 or 10 μM WM for 24 hours. Apoptosis and autophagy were determined using a flow cytometer. The ratio of apoptosis over autophagy and sum of apoptosis plus autophagy were calculated. Data are the mean ± SD of three independent experiments. * P <0.05; ** P <0.01; and *** P <0.001 by one-way ANOVA. Abbreviations: ANOVA, analysis of variance; PLB, plumbagin; SD, standard deviation; WM, wortmannin.

Article Snippet: 4′,6-Diamidino-2-phenylindole (DAPI), 5-(and 6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate (CM-H 2 DCFDA), SB202190 (4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)1 H -imidazole, a selective inhibitor of p38 mitogen-activated protein kinase [MAPK] used as an autophagy inducer), wortmannin (WM, a potent, irreversible, and selective PI3K inhibitor and a blocker of autophagosome formation), phenol red-free culture medium, and fetal bovine serum (FBS) were bought from Invitrogen Inc. (Carlsbad, CA, USA).

Techniques: Flow Cytometry, Standard Deviation

Journal: Experimental and Therapeutic Medicine

Article Title: Herbal melanin induces interleukin-1β secretion and production by human THP-1 monocytes via Toll-like receptor 2 and p38 MAPK activation

doi: 10.3892/etm.2021.10515

Figure Lengend Snippet: Effect of HM on the phosphorylation of p38 MAPK. Representative western blotting. (A) showing the effect of HM and LPS (10 µg/ml. on p38 MAPK phosphorylation in THP-1 cells after 1-, 6- and 24-h stimulation compared with the untreated cells. The bar graph. (B) shows the relative protein expression levels of p-p38 calculated as a ratio of total p38 expression (the loading control). The results are presented as the mean ± SD of three independent experiments. P-values signify a statistically significant difference compared with the control group. HM, herbal melanin; LPS, lipopolysaccharide; IL-1β, interleukin-1β; p-, phosphorylated.

Article Snippet: To investigate whether the HM-induced IL-1β production functions through the TLR and p38 MAPK pathway, the cells were treated with anti-TLR2 (cat. no. ab16894) and anti-TLR4 (cat. no. ab13867) antibodies (both Abcam), or the p38 MAPK pharmacological inhibitor SB202190 (cat no. sc-222294; Santa Cruz Biotechnology, Inc.) to neutralize the TLR receptors and block the p38 MAPK pathway.

Techniques: Phospho-proteomics, Western Blot, Expressing, Control

Journal: Experimental and Therapeutic Medicine

Article Title: Herbal melanin induces interleukin-1β secretion and production by human THP-1 monocytes via Toll-like receptor 2 and p38 MAPK activation

doi: 10.3892/etm.2021.10515

Figure Lengend Snippet: Impact of TLR2, TLR4 receptor neutralization and of the p38 MAPK pathway pharmacological inhibitor SB202190 on HM-induced p38 phosphorylation and IL-1β protein expression. Representative western blot analysis showing the impact of p38 MAPK pharmacological inhibitor SB202190 and TLR4 blockade using anti-TLR4 Ab on HM and LPS-induced. (A) p38 MAPK phosphorylation and (B) IL-1β expression in THP-1 cells after 1-h stimulation compared with the untreated cells, the control. (C) Representative western blotting showing the impact of TLR2 blockade using anti-TLR2 Ab on HM and LPS-induced p38 MAPK phosphorylation and IL-1β expression in the THP-1 cells after 1-h stimulation compared with the untreated cells, the control. The bar graphs show the relative protein expression levels of (A and C) p-p38 and (B and C) IL-1β calculated as a ratio of either total p38 expression or GAPDH (the loading controls). The results are presented as the mean ± SD of three separate experiments. * P<0.05, ** P<0.01, *** P<0.001 and **** P<0.0001 vs. control. TLR, Toll-like receptor; HM, herbal melanin; C, control; LPS, lipopolysaccharide; IL-1β, interleukin-1β; Ab, antibody; p-, phosphorylated.

Article Snippet: To investigate whether the HM-induced IL-1β production functions through the TLR and p38 MAPK pathway, the cells were treated with anti-TLR2 (cat. no. ab16894) and anti-TLR4 (cat. no. ab13867) antibodies (both Abcam), or the p38 MAPK pharmacological inhibitor SB202190 (cat no. sc-222294; Santa Cruz Biotechnology, Inc.) to neutralize the TLR receptors and block the p38 MAPK pathway.

Techniques: Neutralization, Phospho-proteomics, Expressing, Western Blot, Control

Journal: Microorganisms

Article Title: Spring Viremia of Carp Virus Infection Induces Carp IL-10 Expression, Both In Vitro and In Vivo

doi: 10.3390/microorganisms11112812

Figure Lengend Snippet: JAK-STAT, NF-κB and p38 signaling pathways played important roles in SVCV-induced carp IL-10 production. EPC cells were infected or uninfected with SVCV at MOI of 0.5, followed by treatment with JAK inhibitor CP-690550 ( A , B ), STAT3 inhibitor STA-21 ( C , D ), NF-κB inhibitor BAY11-7082 ( E , F ), p38 inhibitor SB202190 ( G , H ), JNK inhibitor SP600125 ( I , J ) or DMSO for 6 h, 12 h, 24 h and 48 h post-infection. The expression of carp IL-10 mRNA and protein were analyzed using RT-PCR and ELISA, respectively. Data are presented as the mean ± SD of three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001 and, ns indicates no significant difference p > 0.05 compared with DMSO-treated plus SVCV-infected cells.

Article Snippet: JAK inhibitor (CP-690550), STAT3 inhibitor (STA-21), NF-κB inhibitor (BAY11-7082) and MAPK inhibitors (SP600125 and SB202190) were purchased from TargetMol (Shanghai, China).

Techniques: Infection, Expressing, Reverse Transcription Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

Journal: Microorganisms

Article Title: Spring Viremia of Carp Virus Infection Induces Carp IL-10 Expression, Both In Vitro and In Vivo

doi: 10.3390/microorganisms11112812

Figure Lengend Snippet: ( A – F ): Gene expressions of JAK1, JAK2, JAK3, TYK2, STAT5 and STAT6 were analyzed using RT-PCR in EPC cells. EPC cells were infected or uninfected with SVCV at MOI of 0.5, followed by treatment with CP-690550, STA-21, BAY11-7082, SB202190, SP600125 and DMSO, respectively, at 6 h, 12 h, 24 h and 48 h post-infection. The gene expressions were analyzed using RT-PCR. Data are presented as the mean ± SD of three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001 compared with DMSO-treated plus SVCV-infected cells.

Article Snippet: JAK inhibitor (CP-690550), STAT3 inhibitor (STA-21), NF-κB inhibitor (BAY11-7082) and MAPK inhibitors (SP600125 and SB202190) were purchased from TargetMol (Shanghai, China).

Techniques: Reverse Transcription Polymerase Chain Reaction, Infection