Journal: PLoS ONE

Article Title: Mechanical Ventilation Enhances HMGB1 Expression in an LPS-Induced Lung Injury Model

doi: 10.1371/journal.pone.0074633

Figure Lengend Snippet: Sham and LPS-treated rats randomized to either spontaneous respiration or moderate tidal volume mechanical ventilation (10 ml/kg) for 4 h. The proteins from lung tissues were used for the detection of p38, ERK, JNK (A) or IκB-α and p65 subunit of NF-κB in the nucleus and cytoplasm (B). Human lung epithelial cells (A549) were pre-incubated with NF-κB inhibitor SN-50, p38 inhibitor SB203580, ERK1/2 inhibitor PD98059 or JNK inhibitor SP 600125 followed by cyclic stretch (CS) at 20% strain, 30 cycle/min for 4 hrs. Cells were then lysed and protein was extracted for Western blot analysis (C). Note remarkable inhibition of HMGB1 expression in animals with SN-50 or SB203580 treatment, but not PD98059 or SP600125. β-actin was used as the loading control. Representative blots of three experiments are shown. All data are expressed as mean±SD. *, p<0.05 vs control group; #, p<0.05 vs CS+LPS group.

Article Snippet: When necessary, cells were pre-incubated with SB203580 (p38 MAPK inhibitor), PD98059 [extracellular signal-regulated kinase (ERK) inhibitor], SP600125 [c-Jun N-terminal kinase (JNK) inhibitor] (Cell Signaling, Danvers, MA, USA), SN-50 (NF-κB inhibitor, Calbiochem, San Diego, CA, USA) or vehicle dimethyl sulfoxide (DMSO).

Techniques: Incubation, Western Blot, Inhibition, Expressing, Control

Journal: The Journal of Biological Chemistry

Article Title: Transcription of the Transforming Growth Factor ? Activating Integrin ?8 Subunit Is Regulated by SP3, AP-1, and the p38 Pathway *

doi: 10.1074/jbc.M110.113977

Figure Lengend Snippet: p38 regulates ITGB8 expression and αvβ8-mediated TGF-β activation. A, flow cytometry for integrin β8 on adult lung fibroblasts treated with MAPK inhibitors, PD98059 (ERK), SB202190 (p38), and SP600125 (JNK) (± S.E.). MFI, mean fluorescence intensity. B, quantitative RT-PCR results for ITGB8 expression in adult lung fibroblasts treated with SB202190, normalized to GAPDH and β-actin and relative to control (± S.E.). C, immunoblot for phosphorylated HSP 27 and dual-phosphorylated ATF-2 from nuclear extracts from adult lung fibroblasts treated ± SB202190. Immunoblot for the nuclear localized proteins, lamins A and C, was used as a loading control. D, TGF-β activation assays of adult lung fibroblasts treated with anti-β8 blocking antibodies or SB202190 (± S.E.). E, quantitative RT-PCR results for MAPK14 (p38α) and ITGB8 in adult lung fibroblasts transfected with plasmids expressing a p38α dominant-negative isoform (p38αDN) or the empty vector control, pcDNA (± S.E.). The measured transcript is labeled above each respective graph. F, TGF-β activation assays of adult lung fibroblasts transfected with plasmids expressing a p38α dominant-negative isoform (p38αDN) or the empty vector control, pcDNA. Percentage (%) of αvβ8-mediated TGF-β activation shown (± S.E.). * = p ≤ 0.05; ** = p ≤ 0.01; *** = p ≤ 0.001.

Article Snippet: The chemical inhibitors of the p38 (SB202190) and extracellular-signaling response kinase (ERK) (PD98059) MAP kinase pathways were obtained from Calbiochem (EMD4 Biosciences, Gibbstown, NJ), whereas the inhibitor of the Jun N-terminal kinase (JNK) pathway (SP600125) was obtained from A.G. Scientific, Inc. (San Diego, CA).

Techniques: Expressing, Activation Assay, Flow Cytometry, Fluorescence, Quantitative RT-PCR, Western Blot, Blocking Assay, Transfection, Dominant Negative Mutation, Plasmid Preparation, Labeling

Journal: PLoS ONE

Article Title: Intestinal Serotonin Transporter Inhibition by Toll-Like Receptor 2 Activation. A Feedback Modulation

doi: 10.1371/journal.pone.0169303

Figure Lengend Snippet: Cells were treated for 30 min or 1 day with 5 μg/ml Pam3CSK4 or 50 ng/ml Pam2CSK4, and/or the different modifiers. Uptake of 5-HT was measured after 6 min of incubation, and 5-HT concentration was 0.2 μM. The results were compared with untreated cells (control). (A) ERK pathway. Cells were treated with Pam3CSK4, Pam2CSK4, and/or 40 μM PD98059. Absolute control values were 10.73±0.33 and 10.54±0.11 pmol 5-HT/mg protein for 30 min and 1 day, respectively. (B) cAMP/PKA pathway. Cells were treated with Pam3CSK4, Pam2CSK4, and/or 1 μM KT 5720. Control absolute values were 9.00±0.16 and 8.61±0.29 pmol 5-HT/mg protein for 30 min and 1 day, respectively. (C) p38 MAPK pathway. Cells were treated with Pam3CSK4, Pam2CSK4, and/or 1 μM SB 220025. Absolute control values were 7.11±0.26 and 7.93±0.12 pmol 5-HT/mg protein for 30 min and 1 day, respectively. The results, in all cases, are expressed as the percentage of the uptake control and are the mean ± SEM of four independent experiments. ***P<0.001, **P<0.01, and *P<0.05 compared with the corresponding control value.

Article Snippet: The following drugs and substances were used (respective suppliers in parentheses): Serotonin (5-hydroxytryptamine, 5-HT), selective p38MAPK inhibitor SB 220025, selective PKA inhibitor KT 5720 and selective inhibitor of ERK pathway PD98059, (Sigma–Aldrich; St. Louis, MO, USA).

Techniques: Incubation, Concentration Assay

Journal: Inflammatory Bowel Diseases

Article Title: Characterization of epithelial IL-8 response to inflammatory bowel disease mucosal E. coli and its inhibition by mesalamine

doi: 10.1002/ibd.20296

Figure Lengend Snippet: The IL-8 response to mucosal E. coli is mediated via the Ras-ERK1/2 and p38 MAPK pathways. A: IL-8 release in response to all 33 mucosal E. coli isolates was markedly reduced in the presence of inhibitors of the ERK and p38 MAPK pathway (U0126 and SB203580, respectively). In contrast, inhibition of JNK activation with 10 μM SP600125 only caused a modest reduction in IL-8 response. All P < 0.001 (ANOVA). B: Mucosal E. coli isolates cause rapid activation of Ras within 1 minute in HT29 cells, returning to basal levels 30 minutes after infection. C: Densitometric analysis of immunoblots for active (GTP-bound) Ras. Ras activation was expressed relative to control ( n = 3 experiments; mean ± SEM). D: Subsequent phosphorylation of ERK1/2 (p44 and p42) occurred within 5 minutes of infection with E. coli . Peak activation was observed at 15 minutes and phosphorylation sustained for at least 60 minutes. E: Densitometric analysis of the p44/p42 immunoblots. ERK1/2 activity was expressed relative to control ( n = 3 experiments; mean ± SEM).

Article Snippet: SP600125 (an inhibitor of c-Jun N-terminal kinase [JNK]), SB203580 (a pyridinyl imidazole inhibitor of p38-MAPK pathway), and Bay11-7082 (an irreversible inhibitor of NF-κB activation, through inhibition of IκB-α phosphorylation) were all obtained from Calbiochem (Nottingham, UK).

Techniques: Inhibition, Activation Assay, Infection, Western Blot, Control, Phospho-proteomics, Activity Assay

Journal: International Journal of Molecular Sciences

Article Title: MAPK Pathway under Chronic Copper Excess in Green Macroalgae (Chlorophyta): Influence on Metal Exclusion/Extrusion Mechanisms and Photosynthesis

doi: 10.3390/ijms20184547

Figure Lengend Snippet: Intracellular copper accumulation in Ulva compressa under copper and/or exposure to mitogen activated protein kinase (MAPK) inhibitors. Treatments: T1—control only with seawater; T2—copper only exposure as 10 µM CuSO 4 (Cu); T3—copper + 5 µM MAPK Extracellular Signal Regulated Kinase (ERK) inhibitor PD98059 (Cu + ERKi); T4—copper + 5 µM MAPK c-Jun N -terminal Kinase (JNK) inhibitor SP600125 (Cu + JNKi); T5—copper + MAPK Cytokinin Specific Binding Protein (p38) inhibitor SB203580 (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9—Cu + ERKi + JNKi + p38i. Samples were analyzed after ( A ) 6 h, ( B ) 24 h, ( C ) 48 h, and ( D ) 6 days. Different letters within each graph represent significant differences at the 95% confidence interval ( p < 0.05). Plots are mean ± SE ( n = 3).

Article Snippet: There was a total of nine treatments, with three replicates each: T1—control with only seawater; T2—chronic copper exposure at 10 μM CuSO 4 (Sigma-Aldrich, St. Louis, MO, USA) (Cu); T3—Cu + 5 μM MAPK ERK inhibitor PD98059 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + ERKi); T4—Cu + 5 μM MAPK JNK inhibitor SP600125 (Tocris Bioscience) (Cu + JNKi); T5—Cu + MAPK p38 inhibitor SB203580 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9—Cu + ERKi + JNKi + p38i.

Techniques: Control, Binding Assay

Journal: International Journal of Molecular Sciences

Article Title: MAPK Pathway under Chronic Copper Excess in Green Macroalgae (Chlorophyta): Influence on Metal Exclusion/Extrusion Mechanisms and Photosynthesis

doi: 10.3390/ijms20184547

Figure Lengend Snippet: Maximum quantum yield of photosystem II (PSII) ( F v /F m ) in U. compressa under copper and/or exposure to MAPK inhibitors. Treatments: T1—control only with seawater; T2—copper only exposure as 10 µM CuSO 4 (Cu); T3—copper + 5 µM MAPK ERK inhibitor PD98059 (Cu + ERKi); T4—copper + 5 µM MAPK JNK inhibitor SP600125 (Cu + JNKi); T5—copper + MAPK p38 inhibitor SB203580 (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9—Cu + ERKi + JNKi + p38i. Samples were analyzed after ( A ) 6 h, ( B ) 24 h, ( C ) 48 h, and ( D ) 6 d of treatment. No significant differences were detected in any experimental times at the 95% confidence interval ( p > 0.05). Plots are mean ± SE ( n = 3).

Article Snippet: There was a total of nine treatments, with three replicates each: T1—control with only seawater; T2—chronic copper exposure at 10 μM CuSO 4 (Sigma-Aldrich, St. Louis, MO, USA) (Cu); T3—Cu + 5 μM MAPK ERK inhibitor PD98059 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + ERKi); T4—Cu + 5 μM MAPK JNK inhibitor SP600125 (Tocris Bioscience) (Cu + JNKi); T5—Cu + MAPK p38 inhibitor SB203580 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9—Cu + ERKi + JNKi + p38i.

Techniques: Control

Journal: International Journal of Molecular Sciences

Article Title: MAPK Pathway under Chronic Copper Excess in Green Macroalgae (Chlorophyta): Influence on Metal Exclusion/Extrusion Mechanisms and Photosynthesis

doi: 10.3390/ijms20184547

Figure Lengend Snippet: Photosynthetic efficiency ( αETR ) in U. compressa under copper and/or exposure to MAPK inhibitors. Treatments: T1—control only with seawater; T2—copper only exposure as 10 µM CuSO 4 (Cu); T3—copper + 5 µM MAPK ERK inhibitor PD98059 (Cu + ERKi); T4—copper + 5 µM MAPK JNK inhibitor SP600125 (Cu + JNKi); T5—copper + MAPK p38 inhibitor SB203580 (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9—Cu + ERKi + JNKi + p38i. Samples were analyzed after ( A ) 6 h, ( B ) 24 h, ( C ) 48 h, and ( D ) 6 d of treatment. Different letters within each graph represent significant differences at the 95% confidence interval ( p < 0.05). Plots are mean ± SE ( n = 3).

Article Snippet: There was a total of nine treatments, with three replicates each: T1—control with only seawater; T2—chronic copper exposure at 10 μM CuSO 4 (Sigma-Aldrich, St. Louis, MO, USA) (Cu); T3—Cu + 5 μM MAPK ERK inhibitor PD98059 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + ERKi); T4—Cu + 5 μM MAPK JNK inhibitor SP600125 (Tocris Bioscience) (Cu + JNKi); T5—Cu + MAPK p38 inhibitor SB203580 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9—Cu + ERKi + JNKi + p38i.

Techniques: Control

Journal: International Journal of Molecular Sciences

Article Title: MAPK Pathway under Chronic Copper Excess in Green Macroalgae (Chlorophyta): Influence on Metal Exclusion/Extrusion Mechanisms and Photosynthesis

doi: 10.3390/ijms20184547

Figure Lengend Snippet: Maximal electron transport rate (ETR max ) in U. compressa under copper and/or exposure to MAPK inhibitors. Treatments: T1—control only with seawater; T2—copper only exposure as 10 µM CuSO 4 (Cu); T3—copper + 5 µM MAPK ERK inhibitor PD98059 (Cu + ERKi); T4—copper + 5 µM MAPK JNK inhibitor SP600125 (Cu + JNKi); T5—copper + MAPK p38 inhibitor SB203580 (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9)—Cu + ERKi + JNKi + p38i. Samples were analyzed after ( A ) 6 h, ( B ) 24 h, ( C ) 48 h, and ( D ) 6 d of treatment. Different letters within each graph represent significant differences at the 95% confidence interval ( p < 0.05). Plots are mean ± SE ( n = 3).

Article Snippet: There was a total of nine treatments, with three replicates each: T1—control with only seawater; T2—chronic copper exposure at 10 μM CuSO 4 (Sigma-Aldrich, St. Louis, MO, USA) (Cu); T3—Cu + 5 μM MAPK ERK inhibitor PD98059 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + ERKi); T4—Cu + 5 μM MAPK JNK inhibitor SP600125 (Tocris Bioscience) (Cu + JNKi); T5—Cu + MAPK p38 inhibitor SB203580 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9—Cu + ERKi + JNKi + p38i.

Techniques: Control

Journal: International Journal of Molecular Sciences

Article Title: MAPK Pathway under Chronic Copper Excess in Green Macroalgae (Chlorophyta): Influence on Metal Exclusion/Extrusion Mechanisms and Photosynthesis

doi: 10.3390/ijms20184547

Figure Lengend Snippet: Saturation of the irradiance of ETR (EkETR) in U. compressa under copper and/or exposure to MAPK inhibitors. Treatments: T1—control only with seawater; T2—copper only exposure as 10 µM CuSO 4 (Cu); T3—copper + 5 µM MAPK ERK inhibitor PD98059 (Cu + ERKi); T4—copper + 5 µM MAPK JNK inhibitor SP600125 (Cu + JNKi); T5—copper + MAPK p38 inhibitor SB203580 (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9)—Cu + ERKi + JNKi + p38i. Samples were analyzed after ( A ) 6 h, ( B ) 24 h, ( C ) 48 h, and ( D ) 6 d of treatment. Different letters within each graph represent significant differences at the 95% confidence interval ( p < 0.05). Plots are mean ± SE ( n = 3).

Article Snippet: There was a total of nine treatments, with three replicates each: T1—control with only seawater; T2—chronic copper exposure at 10 μM CuSO 4 (Sigma-Aldrich, St. Louis, MO, USA) (Cu); T3—Cu + 5 μM MAPK ERK inhibitor PD98059 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + ERKi); T4—Cu + 5 μM MAPK JNK inhibitor SP600125 (Tocris Bioscience) (Cu + JNKi); T5—Cu + MAPK p38 inhibitor SB203580 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9—Cu + ERKi + JNKi + p38i.

Techniques: Control

Journal: International Journal of Molecular Sciences

Article Title: MAPK Pathway under Chronic Copper Excess in Green Macroalgae (Chlorophyta): Influence on Metal Exclusion/Extrusion Mechanisms and Photosynthesis

doi: 10.3390/ijms20184547

Figure Lengend Snippet: Maximal non-photochemical quenching ( NPQ max ) in U. compressa under copper and/or exposure to MAPK inhibitors. Treatments: T1—control only with seawater; T2—copper only exposure as 10 µM CuSO 4 (Cu); T3—copper + 5 µM MAPK Extracellular Signal Regulated Kinase (ERK) inhibitor PD98059 (Cu + ERKi); T4—copper + 5 µM MAPK c-Jun N -terminal Kinase (JNK) inhibitor SP600125 (Cu + JNKi); T5—copper + MAPK Cytokinin Specific Binding Protein (p38) inhibitor SB203580 (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9—Cu + ERKi + JNKi + p38i. Samples were analyzed after ( A ) 6 h, ( B ) 24 h, ( C ) 48 h, and ( D ) 6 d of treatment. Different letters within each graph represent significant differences at the 95% confidence interval ( p < 0.05). No significant differences were detected at experimental times of 6, 24, and 48 h ( p > 0.05). Plots are represented as mean ± SE ( n = 3).

Article Snippet: There was a total of nine treatments, with three replicates each: T1—control with only seawater; T2—chronic copper exposure at 10 μM CuSO 4 (Sigma-Aldrich, St. Louis, MO, USA) (Cu); T3—Cu + 5 μM MAPK ERK inhibitor PD98059 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + ERKi); T4—Cu + 5 μM MAPK JNK inhibitor SP600125 (Tocris Bioscience) (Cu + JNKi); T5—Cu + MAPK p38 inhibitor SB203580 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9—Cu + ERKi + JNKi + p38i.

Techniques: Control, Binding Assay

Journal: International Journal of Molecular Sciences

Article Title: MAPK Pathway under Chronic Copper Excess in Green Macroalgae (Chlorophyta): Influence on Metal Exclusion/Extrusion Mechanisms and Photosynthesis

doi: 10.3390/ijms20184547

Figure Lengend Snippet: Principal Component Ordination (PCO) analysis diagrams expressed in relation to ( a ) time: T1—6 h; T2—24 h; T3—48 h; and T4—6 d; and ( b ) treatments: t1—control only with seawater; t2—copper only exposure as 10 µM CuSO 4 (Cu); t3—copper + 5 µM MAPK ERK inhibitor PD98059 (Cu + ERKi); t4—copper + 5 µM MAPK JNK inhibitor SP600125 (Cu + JNKi); t5—copper + MAPK p38 inhibitor SB203580 (Cu + p38i); t6—Cu + ERKi + JNKi; t7—Cu + ERKi + p38i; t8—Cu + p38i + JNKi; and t9—Cu + ERKi + JNKi + p38i. Vector overlays (Spearman’s rank correlation) indicate the relationship between the PCO axes and the physiological variables, namely, intracellular copper accumulation (copper), photoinhibition ( F v /F m ), productivity (ETR max ), efficiency (α ETR ), and saturation of irradiance (EkETR), accompanied by higher non-photochemical quenching (NPQ max ).

Article Snippet: There was a total of nine treatments, with three replicates each: T1—control with only seawater; T2—chronic copper exposure at 10 μM CuSO 4 (Sigma-Aldrich, St. Louis, MO, USA) (Cu); T3—Cu + 5 μM MAPK ERK inhibitor PD98059 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + ERKi); T4—Cu + 5 μM MAPK JNK inhibitor SP600125 (Tocris Bioscience) (Cu + JNKi); T5—Cu + MAPK p38 inhibitor SB203580 (Tocris Bioscience, St. Louis, MO, USA)) (Cu + p38i); T6—Cu + ERKi + JNKi; T7—Cu + ERKi + p38i; T8—Cu + p38i + JNKi; and T9—Cu + ERKi + JNKi + p38i.

Techniques: Control, Plasmid Preparation

Journal: Experimental and Therapeutic Medicine

Article Title: Baicalein reduces endometriosis by suppressing the viability of human endometrial stromal cells through the nuclear factor-κB pathway in vitro

doi: 10.3892/etm.2017.4860

Figure Lengend Snippet: Effect of signal pathway inhibition on endometrial stromal cell viability.

Article Snippet: Approximately 10,000 endometrial stromal cells were incubated with or without WP1066 [signal transducer and activator of transcription 3 (STAT3) inhibitor; 10 μM], N'-(4-Oxo-4H-chromen-3-yl) methylene) nicotinohydrazide (STAT5 inhibitor; 10 μM), LY294002 (Akt signal pathway inhibitor; 10 μM), SP600125 [c-Jun N-terminal kinase (JNK) inhibitor; 10 μM], SB203580 [p38/mitogen-activated protein kinase (MAPK) inhibitor; 10 μM], U0126 [extracellular signal-regulated kinases 1/2 (ERK1/2) inhibitor; 10 μM], BAY11-7080 [nuclear factor (NF)-κB inhibitor; 10 μM] (all inhibitors were from Santa Cruz Biotechnology, Inc.) for 6 h at 37°C, then treated with or without baicalein (40 μM; Sigma-Aldrich; Merck KGaA) for 48 h at 37°C.

Techniques: Inhibition, Control