Journal: The FASEB Journal

Article Title: ALOX15‐Derived Oxylipins Attenuate Macrophage Inflammatory Signaling Via a Gα q –PLC–PKC Pathway

doi: 10.1096/fj.202600272R

Figure Lengend Snippet: Effect of 12‐HETE on LPS‐induced inflammatory signaling pathways in RAW264.7 cells. RAW264.7 cells were stimulated with lipopolysaccharide (LPS; 1 ng/mL) for 30 min. Cells were pretreated with 12‐HETE (0.1–1 μmol/L) for 1 h prior to LPS stimulation. Phosphorylated p65 (B), p38 (D), ERK (E), Akt (F), and JNK (G), and IκBα protein levels (C) were analyzed by immunoblotting. Representative immunoblot images are shown (A). Because the number of samples exceeded the capacity of a single gel, samples were run on separate gels. Composite images are indicated by clearly demarcated dividing lines. Control samples were included on each gel to allow normalization across gels. Data are presented as mean ± SEM. Statistical significance was assessed using the Tukey–Kramer post hoc test. Groups not sharing a common letter differ significantly ( p < 0.05).

Article Snippet: To examine the inflammatory signaling pathways involved in LPS‐mediated cytokine production, signaling inhibitors targeting p38 MAPK (SB239063; MedChemExpress, Monmouth Junction NJ, USA), NF‐κB (APDC; Merck KGaA), MEK1/2 (U0126; MedChemExpress), phosphatidyl inositol‐3 kinase (PI3K; Wortmannin; Merck KGaA), and c‐Jun N‐terminal kinase (JNK; SP600125; Merck KGaA) were administered 30 min prior to LPS treatment.

Techniques: Protein-Protein interactions, Western Blot, Control

Journal: The FASEB Journal

Article Title: ALOX15‐Derived Oxylipins Attenuate Macrophage Inflammatory Signaling Via a Gα q –PLC–PKC Pathway

doi: 10.1096/fj.202600272R

Figure Lengend Snippet: Effect of 13‐HODE on LPS‐induced inflammatory signaling pathways in RAW264.7 cells. RAW264.7 cells were stimulated with lipopolysaccharide (LPS; 1 ng/mL) for 30 min. Cells were pretreated with 13‐HODE (0.1–1 μmol/L) for 1 h prior to LPS stimulation. Phosphorylated p65 (B), p38 (D), ERK (E), Akt (F), and JNK (G), and IκBα protein levels (C) were analyzed by immunoblotting. Representative immunoblot images are shown (A). Because the number of samples exceeded the capacity of a single gel, samples were run on separate gels. Composite images are indicated by clearly demarcated dividing lines. Control samples were included on each gel to allow normalization across gels. Data are presented as mean ± SEM. Statistical significance was assessed using the Tukey–Kramer post hoc test. Groups not sharing a common letter differ significantly ( p < 0.05).

Article Snippet: To examine the inflammatory signaling pathways involved in LPS‐mediated cytokine production, signaling inhibitors targeting p38 MAPK (SB239063; MedChemExpress, Monmouth Junction NJ, USA), NF‐κB (APDC; Merck KGaA), MEK1/2 (U0126; MedChemExpress), phosphatidyl inositol‐3 kinase (PI3K; Wortmannin; Merck KGaA), and c‐Jun N‐terminal kinase (JNK; SP600125; Merck KGaA) were administered 30 min prior to LPS treatment.

Techniques: Protein-Protein interactions, Western Blot, Control

Journal: JBMR Plus

Article Title: Osteocytes regulate osteoprotegerin expression via the p38-MAPK-CREB pathway in rheumatoid arthritis

doi: 10.1093/jbmrpl/ziag023

Figure Lengend Snippet: TNF-α-induced changes in Wnt/β-catenin-related transcripts and activation of the p38-MAPK-CREB signaling pathway in osteocytes. (A) Dot plots showing relative mRNA levels of Ctnnb1 (β-catenin), Lef1 , and Axin2 in MLO-Y4 cells treated with or without TNF-α ( n = 3). (B) Representative immunoblotting images showing phosphorylated p38 MAPK (p-p38), total p38, phosphorylated CREB (p-CREB), total CREB, phosphorylated JNK (p-JNK), total JNK, phosphorylated ERK (p-ERK), total ERK, and β-actin in MLO-Y4 cells following TNF-α treatment. Data in (A) are shown as individual values with the mean ± SD and were analyzed by the two-sided Student’s t -test. (C) Representative immunoblot of OPG in TNF-α-stimulated MLO-Y4 cells treated with a p38 inhibitor or a CREB inhibitor. Cells were stimulated with TNF-α in the presence or absence of SB203580 (p38 inhibitor) or 666-15 (CREB inhibitor), and total cell lysates were subjected to immunoblotting for OPG. β-actin serves as the loading control. Numeric values shown beneath each band represent the relative expression levels of OPG/β-actin for the corresponding lane. (D) ChIP-PCR demonstrating enrichment of the OPG promoter in anti-CREB immunoprecipitates from MLO-Y4 cells with or without TNF-α stimulation, with densitometric quantification of the ChIP-PCR bands; input DNA served as the internal control, normal IgG as the negative control, and anti-histone H3 as the positive control.

Article Snippet: For inhibitor experiments, the p38 MAPK inhibitor SB203580 (Selleck Chemicals) was used at a concentration of 30 μM, and the CREB inhibitor 666-15 (Selleck Chemicals) was used at a concentration of 0.1 μM.

Techniques: Activation Assay, Western Blot, Control, Expressing, Negative Control, Positive Control

Journal: Translational Oncology

Article Title: NF-κB activation as a pro-survival signal from pharmacological inhibition of pyruvate dehydrogenase kinase 1 in non-small-cell lung carcinoma cell models

doi: 10.1016/j.tranon.2026.102681

Figure Lengend Snippet: 64 activated NF-κB pathway through P38-MAPK. NCI-H1975 and NCI-H1650 cells were treated for 24 h with varying concentrations of 64 either alone ( a – d ) or in combination with SB203580 ( e ). a . The top 17 KEGG pathways of these two cell lines were displayed. b . The heatmap displayed individual genes in the top regulated pathways, namely NF-κB and MAPK. c . qPCR assay was performed to measure the expression levels of individual genes highlighted in the heatmap. d . Western blotting was conducted to examine the levels of P65 in the cytoplasm and nuclear. e . Western blotting was conducted to examine the levels of P65 in the cytoplasm and nuclear under treatments with 64 (5µM) and the absence/ presence of SB203580 (20µM). The data was obtained from three independent experiments with n = 3 and were presented as mean ± SD. The statistical significance between compared groups was indicated by * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: P38 MAPK inhibitor SB203580 (S1076) and NF-κB inhibitor JSH-23 (S7351) were purchased from SelleckChem (Houston, TX, USA).

Techniques: Expressing, Western Blot

Journal: The Journal of Biological Chemistry

Article Title: Starvation of leukemic cells enhances DNA damage-induced apoptosis in vitro via ROS/p38 MAPK and prevents leukemia progression in fasting xenograft mice

doi: 10.1016/j.jbc.2026.111143

Figure Lengend Snippet: Starvation-induced killing of ALL cells involves ROS-mediated activation of p38 MAPK. A , REH cells (0.8 × 10 6 cells/ml) were cultured in CM or DM in the presence or absence of forskolin (Forsk, 70 μM) for 45 min prior to IR, 7 Gy. The cells were harvested 1 hour after IR, and total cell lysates were subjected to immunoblot analyses with antibodies against p38 MAPK (p38), phosphorylated p38 MAPK (pp38). Antibodies against vinculin were used as a control of equal loading. Left panel ( upper ) shows one representative Western blot of six independent experiments. Right panel shows a quantified overview of pp38 signal intensity relative to the vinculin signal. The data represent the mean ± SEM, n = 6, ∗ P < 0.05 (paired t-test). Left panel (lower) shows a representative Western blot from one of one experiment using antibodies against p38 and vinculin. B , REH cells (0.8 × 10 6 cells/ml) were cultured in DM in the presence or absence of 40 μM of the p38 MAPK inhibitor (SB 202190) for 30 min, followed by incubation with or without forskolin (Forsk, 70 μM) for 45 min prior to IR, 7 Gy. The cells were harvested 1 hour after IR, and total cell lysates were subjected to immunoblot analyses with antibodies against p38, pp38 and vinculin. Left panel (upper) shows one representative Western blot of four independent experiments. Right panel shows a quantification of the pp38 signal intensity relative to vinculin signal. The data represent the mean ± SEM, n = 3, ∗ P < 0.05 (paired t-test). Left panel ( lower ) shows a representative Western blot from one of one experiment using antibodies against p38 and vinculin. C , REH cells (0.8 × 10 6 cells/ml) were cultured in DM in the presence or absence of 10 mM NAC for 30 min, followed by incubation with or without forskolin (Forsk, 70 μM) for 45 min prior to IR, 7 Gy. The cells were harvested 1 hour after IR, and total cell lysates were subjected to immunoblot analyses with antibodies against pp38 and vinculin. Left panel shows one representative Western blot of three independent experiments. Right panel shows quantification of the pp38 signal intensity relative to the vinculin signal. The data represent the mean ± SEM, n = 4, ∗ P < 0.05 (paired t-test). In all panels, ‘control’ represents cells cultured in the relevant medium alone. CM, complete medium; DM, depleted medium.

Article Snippet: SB 202190, the p38 MAPK inhibitor and the vinculin mouse monoclonal antibody (#V9131) were from Sigma-Aldrich, the p53 (DO-1, #SC-126) mouse monoclonal antibody and the β-actin mouse monoclonal antibody (C4 #sc-47778) were purchased from Santa Cruz Biotechnology.

Techniques: Activation Assay, Cell Culture, Western Blot, Control, Incubation

Journal: The Journal of Biological Chemistry

Article Title: Starvation of leukemic cells enhances DNA damage-induced apoptosis in vitro via ROS/p38 MAPK and prevents leukemia progression in fasting xenograft mice

doi: 10.1016/j.jbc.2026.111143

Figure Lengend Snippet: Apoptosis induced by forskolin and IR in starving ALL cells involves the activation of p38 MAPK. A and D , REH cells (0.4 × 106 cells/ml) were cultured in DM in the presence or absence of 10 μM of the p38 MAPK inhibitor SB 202190 for 30 min, followed by incubation with forskolin (Forsk, 70 μM) for 45 min prior to IR, 10 Gy. A , the percentages of dead cells were monitored by flow cytometry of cells stained with PI 2 hours after IR. The data represent the mean ± SEM, n = 4, ∗ P < 0.05 (paired t-test). B , the loss of mitochondrial membrane potential was assessed by flow cytometry of cells stained with TMRM 2 hours after IR and shown as percentage of TMRM-low cells, representing apoptotic cells. The data represent the mean ± SEM, n = 3, ∗ P < 0.05 (paired t-test). C , total ROS levels were analyzed by staining the cells with CellROX green 1 hour after IR, and the MFI was analyzed by flow cytometry. The data represent the mean ± SEM, n = 4, ∗ P < 0.05 (paired t-test). D , the cells were subjected to CYTO-ID staining for detection of autophagy 24 hours after IR, and the MFI was analyzed by flow cytometry. The data represent the mean ± SEM, n = 3, ∗ P < 0.05 (paired t-test). E , model explaining how DNA damage-induced killing of ALL cells is enhanced under starving culture conditions in the presence of cAMP signaling. Our model suggests that starvation promotes cAMP-mediated ( via forskolin, Forsk) enhancement of DNA damage-induced ( via IR) killing of ALL cells by enhancing the ROS levels. Increased ROS levels activate p38 MAPK, which subsequently leads to apoptosis. Additionally, ROS is involved in starvation-induced autophagy, which is prompted by cAMP signaling in the presence of a DNA damaging agent. Notably, activation of p38 MAPK appears to function both upstream and downstream of ROS. According to our model, the cells die with – but not due to the high levels of autophagy. In all panels, ‘control’ represents cells cultured in the relevant medium alone. DM, depleted medium; MFI, mean fluorescence intensity.

Article Snippet: SB 202190, the p38 MAPK inhibitor and the vinculin mouse monoclonal antibody (#V9131) were from Sigma-Aldrich, the p53 (DO-1, #SC-126) mouse monoclonal antibody and the β-actin mouse monoclonal antibody (C4 #sc-47778) were purchased from Santa Cruz Biotechnology.

Techniques: Activation Assay, Cell Culture, Incubation, Flow Cytometry, Staining, Membrane, Control, Fluorescence