phospho p38 mapk  (Cell Signaling Technology Inc)

Journal: Hepatic Oncology

Article Title: RTA‑408 induces p38‑dependent apoptosis and suppresses cell viability in hepatocellular carcinoma cells

doi: 10.1080/20450923.2026.2659967

Figure Lengend Snippet: RTA-408 activates p38–NRF2 signaling and promotes LC3B accumulation in hepatocellular carcinoma cells. HepG2 and PP5 cells were treated with RTA-408 (0, 200, 400, or 600 nM) for 24 h, and protein expression was analyzed by western blotting. Representative immunoblots show phosphorylated p38 (p-p38), total p38, phosphorylated ERK1/2 (p-ERK), total ERK1/2, phosphorylated JNK (p-JNK), total JNK, NRF2, LC3B, p62, cleaved caspase-3 (c-cas3), and β-actin. Densitometric analyses of p-p38/p38, p-ERK/ERK, and p-JNK/JNK, as well as NRF2, LC3B, p62, and c-cas3 normalized to β-actin, are shown in the accompanying bar graphs. RTA-408 increased p38 phosphorylation and NRF2 expression in both cell lines and was accompanied by marked accumulation of LC3B and p62, while ERK and JNK responses differed between HepG2 and PP5 cells. Cleaved caspase-3 increased at higher concentrations of RTA-408. β-Actin was used as a loading control. Data are presented as mean ± SEM from three independent experiments ( n = 3). *P < 0.05 and **P < 0.01 versus control.

Article Snippet: RTA-408 (omaveloxolone) (MCE; 1474034-05-3; USA) and the p38 MAPK inhibitor, SB203580 (MCE; 152121-47-6; USA), were dissolved in dimethyl sulfoxide (DMSO) to prepare stock solutions (10 mM for RTA-408 and 10 mM for SB203580), which were stored at − 20 °C according to the manufacturer’s instructions.

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

Journal: Hepatic Oncology

Article Title: RTA‑408 induces p38‑dependent apoptosis and suppresses cell viability in hepatocellular carcinoma cells

doi: 10.1080/20450923.2026.2659967

Figure Lengend Snippet: p38 inhibition attenuates RTA-408-induced p38 signaling, LC3B/p62 accumulation, and apoptotic protein activation in hepatocellular carcinoma cells. HepG2 and PP5 cells were treated for 24 h with vehicle control (Con), RTA-408 (400 nM) alone, or RTA-408 (400 nM) plus SB203580 (10 μM). Protein expression was analyzed by western blotting. Representative immunoblots show phosphorylated p38 (p-p38), total p38, NRF2, LC3B, p62, cleaved caspase-3 (c-cas3), and β-actin. Densitometric analyses of p-p38/p38, as well as NRF2, LC3B, p62, and c-cas3 normalized to β-actin, are shown in the accompanying bar graphs. In both HepG2 and PP5 cells, RTA-408 increased p38 phosphorylation, NRF2 expression, LC3B accumulation, p62 accumulation, and cleaved caspase-3 levels, whereas co-treatment with SB203580 markedly attenuated these RTA-408-induced effects without altering total p38 expression. These findings indicate that p38 activity contributes to RTA-408-induced stress signaling, LC3B/p62-associated responses, and apoptotic protein activation in hepatocellular carcinoma cells. Data are presented as mean ± SEM from three independent experiments ( n = 3). *P < 0.05 and **P < 0.01 versus control; #P < 0.05 and ##P < 0.01 versus RTA-408 alone.

Article Snippet: RTA-408 (omaveloxolone) (MCE; 1474034-05-3; USA) and the p38 MAPK inhibitor, SB203580 (MCE; 152121-47-6; USA), were dissolved in dimethyl sulfoxide (DMSO) to prepare stock solutions (10 mM for RTA-408 and 10 mM for SB203580), which were stored at − 20 °C according to the manufacturer’s instructions.

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

Journal: Hepatic Oncology

Article Title: RTA‑408 induces p38‑dependent apoptosis and suppresses cell viability in hepatocellular carcinoma cells

doi: 10.1080/20450923.2026.2659967

Figure Lengend Snippet: Pharmacological inhibition of p38 MAPK partially restores cell viability and attenuates RTA-408-induced apoptosis in hepatocellular carcinoma cells. (A) HepG2 and PP5 cells were treated for 24 h with vehicle control (Con), RTA-408 (400 nM) alone, or RTA-408 (400 nM) plus SB203580 (10 μM), and cell viability was assessed by the MTT assay. Bar graphs show the percentage of viable cells relative to the control group. (B) Under the same treatment conditions, apoptosis was evaluated by Annexin V/7-AAD flow cytometry. Representative dot plots show the distribution of viable (Annexin V − /7-AAD − ), early apoptotic (Annexin V + /7-AAD − ), late apoptotic (Annexin V + /7-AAD + ), and necrotic (Annexin V − /7-AAD + ) cell populations. The accompanying bar graphs summarize total apoptosis (early + late apoptosis). In both cell lines, RTA-408 reduced cell viability and increased apoptosis, whereas co-treatment with SB203580 partially reversed the loss of viability and attenuated the pro-apoptotic effect of RTA-408. Data are presented as mean ± SEM from three independent experiments ( n = 3). Statistical significance was determined by one-way analysis of variance with appropriate post hoc testing. *P < 0.05, **P < 0.01, and ***P < 0.001 versus control; ##P < 0.01 versus RTA-408 alone.

Article Snippet: RTA-408 (omaveloxolone) (MCE; 1474034-05-3; USA) and the p38 MAPK inhibitor, SB203580 (MCE; 152121-47-6; USA), were dissolved in dimethyl sulfoxide (DMSO) to prepare stock solutions (10 mM for RTA-408 and 10 mM for SB203580), which were stored at − 20 °C according to the manufacturer’s instructions.

Techniques: Inhibition, Control, MTT Assay, Flow Cytometry

Journal: Poultry Science

Article Title: PKA and PKC signaling pathways mediate vitamin D₃-regulated intestinal phosphorus absorption in broiler chickens

doi: 10.1016/j.psj.2026.106762

Figure Lengend Snippet: Effects of dietary vitamin D 3 supplementation on jejunal phosphorylation levels of PKA, PKC, PI3K, p38MAPK, and ERK in broiler chickens (19 d) (Experiment 1). The control and vitamin D 3 diets contained 0 and 1000 IU/kg vitamin D 3 , respectively. (a) p-PKA/t-PKA; (b) p-PKC/t-PKC; (c) p-PI3K/t-PI3K; (d) p-p38MAPK/t-p38MAPK; (e) p-ERK/t-ERK. Protein abundance values are means ± SD of four replicates per treatment (1 broiler per replicate) (n = 4). Values with different letters differ between treatments ( P < 0.05).

Article Snippet: PVDF membranes were incubated with primary antibodies against NaPi-IIb (1:1000, A9460; ABclonal, Wuhan, China), phosphorylated PKA (p-PKA; 1:1000, 5661S), phosphorylated PI3K (p-PI3K; 1:1000, 4228), phosphorylated ERK (p-ERK; 1:1000, 9101S), phosphorylated p38MAPK (p-p38MAPK; 1:1000, 9216S), total PKA (t-PKA; 1:1000, 5842S) (Cell Signaling Technology, Boston, USA), total PI3K (t-PI3K; 1:10000, 60225-1-Ig), total ERK (t-ERK; 1:8000, 11257-1-AP-50), total p38MAPK(t-p38MAPK; 1:1000, 14064-1-AP), total PKC (t-PKC; 1:1000, 21991-1-AP), phosphorylated PKC (p-PKC; 1:5000, 29123-1-AP), and GAPDH (1:10000, 60004-1-Ig) (Proteintech, Wuhan, China).

Techniques: Phospho-proteomics, Control, Quantitative Proteomics

Journal: Poultry Science

Article Title: PKA and PKC signaling pathways mediate vitamin D₃-regulated intestinal phosphorus absorption in broiler chickens

doi: 10.1016/j.psj.2026.106762

Figure Lengend Snippet: Effects of the PKA inhibitor H-89 on jejunal phosphorylation levels of PKC, PI3K, p38MAPK, and ERK in broiler chickens (10–15 d) (Experiment 2). (a) p-PKC/t-PKC; (b) p-PI3K/t-PI3K; (c) p-p38MAPK/t-p38MAPK; (d) p-ERK/t-ERK. Protein abundance values are means ± SD of four replicates per treatment (1 broiler per replicate) (n = 4). Values with different letters differ between treatments ( P < 0.05).

Article Snippet: PVDF membranes were incubated with primary antibodies against NaPi-IIb (1:1000, A9460; ABclonal, Wuhan, China), phosphorylated PKA (p-PKA; 1:1000, 5661S), phosphorylated PI3K (p-PI3K; 1:1000, 4228), phosphorylated ERK (p-ERK; 1:1000, 9101S), phosphorylated p38MAPK (p-p38MAPK; 1:1000, 9216S), total PKA (t-PKA; 1:1000, 5842S) (Cell Signaling Technology, Boston, USA), total PI3K (t-PI3K; 1:10000, 60225-1-Ig), total ERK (t-ERK; 1:8000, 11257-1-AP-50), total p38MAPK(t-p38MAPK; 1:1000, 14064-1-AP), total PKC (t-PKC; 1:1000, 21991-1-AP), phosphorylated PKC (p-PKC; 1:5000, 29123-1-AP), and GAPDH (1:10000, 60004-1-Ig) (Proteintech, Wuhan, China).

Techniques: Phospho-proteomics, Quantitative Proteomics

Journal: Poultry Science

Article Title: PKA and PKC signaling pathways mediate vitamin D₃-regulated intestinal phosphorus absorption in broiler chickens

doi: 10.1016/j.psj.2026.106762

Figure Lengend Snippet: Effects of the PKC inhibitor staurosporine on duodenal phosphorylation levels of PKA, PI3K, p38MAPK, and ERK in broiler chickens (13 d) (Experiment 3). (a) p-PKA/t-PKA; (b) p-PI3K/t-PI3K; (c) p-p38MAPK/t-p38MAPK; (d) p-ERK/t-ERK. Protein abundance values are means ± SD of four replicates per treatment (1 broiler per replicate) (n = 4). Values with different letters differ between treatments ( P < 0.05).

Article Snippet: PVDF membranes were incubated with primary antibodies against NaPi-IIb (1:1000, A9460; ABclonal, Wuhan, China), phosphorylated PKA (p-PKA; 1:1000, 5661S), phosphorylated PI3K (p-PI3K; 1:1000, 4228), phosphorylated ERK (p-ERK; 1:1000, 9101S), phosphorylated p38MAPK (p-p38MAPK; 1:1000, 9216S), total PKA (t-PKA; 1:1000, 5842S) (Cell Signaling Technology, Boston, USA), total PI3K (t-PI3K; 1:10000, 60225-1-Ig), total ERK (t-ERK; 1:8000, 11257-1-AP-50), total p38MAPK(t-p38MAPK; 1:1000, 14064-1-AP), total PKC (t-PKC; 1:1000, 21991-1-AP), phosphorylated PKC (p-PKC; 1:5000, 29123-1-AP), and GAPDH (1:10000, 60004-1-Ig) (Proteintech, Wuhan, China).

Techniques: Phospho-proteomics, Quantitative Proteomics

Journal: Poultry Science

Article Title: PKA and PKC signaling pathways mediate vitamin D₃-regulated intestinal phosphorus absorption in broiler chickens

doi: 10.1016/j.psj.2026.106762

Figure Lengend Snippet: Effects of dietary vitamin D 3 supplementation on jejunal phosphorylation levels of PKA, PKC, PI3K, p38MAPK, and ERK in broiler chickens (19 d) (Experiment 1). The control and vitamin D 3 diets contained 0 and 1000 IU/kg vitamin D 3 , respectively. (a) p-PKA/t-PKA; (b) p-PKC/t-PKC; (c) p-PI3K/t-PI3K; (d) p-p38MAPK/t-p38MAPK; (e) p-ERK/t-ERK. Protein abundance values are means ± SD of four replicates per treatment (1 broiler per replicate) (n = 4). Values with different letters differ between treatments ( P < 0.05).

Article Snippet: PVDF membranes were incubated with primary antibodies against NaPi-IIb (1:1000, A9460; ABclonal, Wuhan, China), phosphorylated PKA (p-PKA; 1:1000, 5661S), phosphorylated PI3K (p-PI3K; 1:1000, 4228), phosphorylated ERK (p-ERK; 1:1000, 9101S), phosphorylated p38MAPK (p-p38MAPK; 1:1000, 9216S), total PKA (t-PKA; 1:1000, 5842S) (Cell Signaling Technology, Boston, USA), total PI3K (t-PI3K; 1:10000, 60225-1-Ig), total ERK (t-ERK; 1:8000, 11257-1-AP-50), total p38MAPK(t-p38MAPK; 1:1000, 14064-1-AP), total PKC (t-PKC; 1:1000, 21991-1-AP), phosphorylated PKC (p-PKC; 1:5000, 29123-1-AP), and GAPDH (1:10000, 60004-1-Ig) (Proteintech, Wuhan, China).

Techniques: Phospho-proteomics, Control, Quantitative Proteomics

Journal: Poultry Science

Article Title: PKA and PKC signaling pathways mediate vitamin D₃-regulated intestinal phosphorus absorption in broiler chickens

doi: 10.1016/j.psj.2026.106762

Figure Lengend Snippet: Effects of the PKA inhibitor H-89 on jejunal phosphorylation levels of PKC, PI3K, p38MAPK, and ERK in broiler chickens (10–15 d) (Experiment 2). (a) p-PKC/t-PKC; (b) p-PI3K/t-PI3K; (c) p-p38MAPK/t-p38MAPK; (d) p-ERK/t-ERK. Protein abundance values are means ± SD of four replicates per treatment (1 broiler per replicate) (n = 4). Values with different letters differ between treatments ( P < 0.05).

Article Snippet: PVDF membranes were incubated with primary antibodies against NaPi-IIb (1:1000, A9460; ABclonal, Wuhan, China), phosphorylated PKA (p-PKA; 1:1000, 5661S), phosphorylated PI3K (p-PI3K; 1:1000, 4228), phosphorylated ERK (p-ERK; 1:1000, 9101S), phosphorylated p38MAPK (p-p38MAPK; 1:1000, 9216S), total PKA (t-PKA; 1:1000, 5842S) (Cell Signaling Technology, Boston, USA), total PI3K (t-PI3K; 1:10000, 60225-1-Ig), total ERK (t-ERK; 1:8000, 11257-1-AP-50), total p38MAPK(t-p38MAPK; 1:1000, 14064-1-AP), total PKC (t-PKC; 1:1000, 21991-1-AP), phosphorylated PKC (p-PKC; 1:5000, 29123-1-AP), and GAPDH (1:10000, 60004-1-Ig) (Proteintech, Wuhan, China).

Techniques: Phospho-proteomics, Quantitative Proteomics

Journal: Poultry Science

Article Title: PKA and PKC signaling pathways mediate vitamin D₃-regulated intestinal phosphorus absorption in broiler chickens

doi: 10.1016/j.psj.2026.106762

Figure Lengend Snippet: Effects of the PKC inhibitor staurosporine on duodenal phosphorylation levels of PKA, PI3K, p38MAPK, and ERK in broiler chickens (13 d) (Experiment 3). (a) p-PKA/t-PKA; (b) p-PI3K/t-PI3K; (c) p-p38MAPK/t-p38MAPK; (d) p-ERK/t-ERK. Protein abundance values are means ± SD of four replicates per treatment (1 broiler per replicate) (n = 4). Values with different letters differ between treatments ( P < 0.05).

Article Snippet: PVDF membranes were incubated with primary antibodies against NaPi-IIb (1:1000, A9460; ABclonal, Wuhan, China), phosphorylated PKA (p-PKA; 1:1000, 5661S), phosphorylated PI3K (p-PI3K; 1:1000, 4228), phosphorylated ERK (p-ERK; 1:1000, 9101S), phosphorylated p38MAPK (p-p38MAPK; 1:1000, 9216S), total PKA (t-PKA; 1:1000, 5842S) (Cell Signaling Technology, Boston, USA), total PI3K (t-PI3K; 1:10000, 60225-1-Ig), total ERK (t-ERK; 1:8000, 11257-1-AP-50), total p38MAPK(t-p38MAPK; 1:1000, 14064-1-AP), total PKC (t-PKC; 1:1000, 21991-1-AP), phosphorylated PKC (p-PKC; 1:5000, 29123-1-AP), and GAPDH (1:10000, 60004-1-Ig) (Proteintech, Wuhan, China).

Techniques: Phospho-proteomics, Quantitative Proteomics

Journal: The Journal of Experimental Medicine

Article Title: MYCT1–IFITM2/3 interaction links endothelial endolysosomal trafficking to white adipose tissue expansion

doi: 10.1084/jem.20251497

Figure Lengend Snippet: MYCT1 limits endothelial mTORC1 signaling, related to Figs. 3 and 4. (A) Flow cytometry gating strategy (CD45 neg CD31 + ) for sorting of ECs from mesenteric fat for scRNA-seq. (B) Dot plot of markers for the indicated clusters. Color code: scaled average expression level in each cluster; the dot size denotes the percent of cells in each cluster expressing the given gene. (C) Number of differentially expressed genes (DEGs) between wild-type and Myct1 ecKO cell clusters. (D) Volcano plot of DEGs between the wild-type and Myct1 ecKO mice in the BEC cluster. Mat2a gene was selected for scRNA-seq validation. Mat2a, methionine adenosyltransferase 2A. (E) MYCT1 protein levels in human primary ECs. Western blot analysis for the indicated proteins. HPMECs, human pulmonary ECs; HUVECs, human umbilical vein ECs; HIECs, human intestinal ECs. (F and G) MYCT1 antibody and siRNAs validation for identification of endogenous human MYCT1 protein. Human primary ECs were transfected with two different MYCT1 targeting siRNAs. (F) Staining of ECs for MYCT1 (black) and DAPI (blue). Arrow, not transfected EC. Scale bar, 50 μm. (G) Western blot analysis showing MYCT1 migration profile and siRNA specificity. (H) MYCT1 knockdown increases phosphorylation of S6 but does not affect AKT and ERK1/2 phosphorylation status. Western blot analysis for the indicated proteins. (I) Quantification of p-S6 Ser240/244 levels normalized to total S6 (tot-S6). P = 0.037 (*). (J) Quantification of p-AKT Ser473 levels normalized to total AKT (tot-AKT). P > 0.05. (K) Quantification of p-AKT Thr308 levels normalized to total AKT (tot-AKT). P > 0.05. (L) Quantification of p-ERK1/2 Thr202/Tyr204 levels normalized to total ERK1/2 (tot-ERK1/2). P > 0.05. (M) Quantification of MYCT1 levels normalized to vinculin. P = 0.001 (*). (I–M) n = 5 independent experiments; paired t tests. (N) MYCT1 knockdown increases phosphorylation of p70/S6 kinase (p70/S6K), a key downstream effector of mTORC1 signaling, in response to amino acids. Western blot for the indicated proteins. (O) Quantification of data shown in N. n = 2 independent experiments; mean ± SD; two-way ANOVA with Tukey’s multiple comparison test, P = 0.0194 (*). (P) MYCT1 knockdown hyperactivates mTORC1 signaling in response to amino acids. 2 days after siRNA transfection, confluent ECs were serum- and growth factor–starved overnight, then starved in PBS for 1 h before 30-min stimulation with amino acids, glucose, growth factors, FBS, their combination, or PBS as control. Staining of ECs for p-S6 (gray), VE-cadherin (magenta), and DAPI (blue). Scale bar, 50 μm. Source data are available for this figure: .

Article Snippet: Rabbit monoclonal anti-phospho-p44/42 MAPK (Erk1/2) (Thr202/Tyr204) , Cell Signaling Technology , Cat# 4376, RRID:AB_331772.

Techniques: Flow Cytometry, Expressing, Biomarker Discovery, Western Blot, Transfection, Staining, Migration, Knockdown, Phospho-proteomics, Comparison, Control