Journal: Redox Report : Communications in Free Radical Research

Article Title: The novel thioredoxin reductase inhibitor butaselen suppresses lung cancer by inducing oxidative stress

doi: 10.1080/13510002.2025.2588086

Figure Lengend Snippet: BS affects NF-κB, p38/JNK, and PI3K-Akt signaling pathways in lung cancer. (a–d) A549 and H1299 cells were treated with BS (0, 5, 10 μM) for 24 h, followed by western blot. Error bars are means ± std. * P < 0.05, ** P < 0.01, *** P < 0.001, compared with the control group by One-way ANOVA (n = 3).

Article Snippet: The primary antibodies used for immunoblotting analysis are as follows: TrxR1 (Proteintech, 11117-1-AP), Trx1 (Proteintech, 14999-1-AP), HBP1 (Proteintech, 11746-1-AP), DNMT1 (Proteintech, 24206-1-AP), Bcl-2 (Proteintech, 12789-1-AP), Bax (Proteintech, 50599-2-Ig), β-actin (Bioss, bs-0061R), Flag (Sigma-Aldrich, F1804), HA (Covance, MMS-101P), p53 (Santa, sc-126), p21 (MBL, K0081-3), p27 (MBL, K0082), γ-H2AX (CST, 9718), NF-κB (Abcam, ab32536), p-NF-κB(CST, 3033), p38 (Santa, sc-7972), p-p38 (Santa, sc-101759), JNK (MCE, HY- P80728 ), p-JNK (Immunoway, YP0157), Akt (Santa, sc-5298), phospho-Akt (CST, 4060).

Techniques: Protein-Protein interactions, Western Blot, Control

Journal: Redox Report : Communications in Free Radical Research

Article Title: The novel thioredoxin reductase inhibitor butaselen suppresses lung cancer by inducing oxidative stress

doi: 10.1080/13510002.2025.2588086

Figure Lengend Snippet: Schematic model of lung cancer inhibition by BS. The TrxR/Trx inhibitor butaselen (BS) can inhibit lung cancer by inducing ROS-dependent apoptosis. The inactivation of the NF-κB and PI3K-Akt signaling pathways, along with the activation of the p38/JNK signaling pathway, contributes to the anti-cancer effects of BS on lung cancer. Although p53 itself is not activated by BS, the HBP1/DNMT1/p21/γ-H2AX/Bcl-2/Bax signaling pathway is activated by BS and contributes to its tumor-inhibitory role. Further mechanistic studies revealed HBP1 as a novel target of the Trx system. The Trx system inversely associates with HBP1 in lung cancer and regulates HBP1 expression at the post-translational level. Under normal conditions, TrxR1 catalyzes the reduction of Trx1 by utilizing NADPH. In its reduced form, Trx1 interacts with HBP1, promoting the ubiquitination of HBP1, which leads to its proteasomal degradation and maintains a low level of HBP1 within cancer cells. Treatment with butaselen inhibits the activity of TrxR1 in lung cancer cells, resulting in the oxidation of Trx1 and the subsequent excessive generation of ROS. HBP1 is activated after being released by the oxidized Trx1 and escaping proteasomal degradation. The activated HBP1 inhibits the expression of DNMT1 and elevates Bax. The decreased DNMT1 further results in the demethylation of the whole genome as well as the promoters of p21 and HOXA9. Ultimately, the upregulation of p21 and γ-H2AX, along with the downregulation of DNMT1 and Bcl-2/Bax, contributes to the apoptosis of lung cancer cells induced by BS. Taken together, the TrxR/Trx inhibitor butaselen inhibits lung cancer by promoting ROS-induced apoptosis through the NF-κB, PI3K-Akt, p38/JNK, and HBP1/DNMT1 signaling pathways.

Article Snippet: The primary antibodies used for immunoblotting analysis are as follows: TrxR1 (Proteintech, 11117-1-AP), Trx1 (Proteintech, 14999-1-AP), HBP1 (Proteintech, 11746-1-AP), DNMT1 (Proteintech, 24206-1-AP), Bcl-2 (Proteintech, 12789-1-AP), Bax (Proteintech, 50599-2-Ig), β-actin (Bioss, bs-0061R), Flag (Sigma-Aldrich, F1804), HA (Covance, MMS-101P), p53 (Santa, sc-126), p21 (MBL, K0081-3), p27 (MBL, K0082), γ-H2AX (CST, 9718), NF-κB (Abcam, ab32536), p-NF-κB(CST, 3033), p38 (Santa, sc-7972), p-p38 (Santa, sc-101759), JNK (MCE, HY- P80728 ), p-JNK (Immunoway, YP0157), Akt (Santa, sc-5298), phospho-Akt (CST, 4060).

Techniques: Inhibition, Protein-Protein interactions, Activation Assay, Expressing, Ubiquitin Proteomics, Activity Assay

Journal: Evidence-based Complementary and Alternative Medicine : eCAM

Article Title: Research on Roles of Mongolian Medical Warm Acupuncture in Inhibiting p38 MAPK Activation and Apoptosis of Nucleus Pulposus Cells

doi: 10.1155/2018/6571320

Figure Lengend Snippet: Phosphorylation level of p38 MAPK in the nucleus pulposus cells and effect of acupuncture on substrate metabolism of the rat intervertebral disc. (a) The phosphorylation level of p38 MAPK decreased following administration of analgesic decoction. (b/c) The expression of the Type II collagen, aggrecan, Sox-9, and TIMP-1 in the nucleus pulposus cells in the model group significantly decreased when compared with that in the control group. The expression of MMP-3 increased significantly when compared with that in the control group. # (##) indicates P<0.05 (P<0.01) when compared with the control group; ∗ ( ∗∗ ) indicates P<0.05 (P<0.01) when compared with the 25 mM glucose group.

Article Snippet: The protein sample was transferred to the nitrocellulose membrane, blocked, and incubated with the p38 MAPK antibody (dilution rate 1 : 1 000) (Miltenyi Biotec, Bergisch-Gladbach, Germany) and the GAPDH antibody (1 : 2 000) (Wuhan Boster Biological Engineering Co., Ltd., Wuhan, China), respectively, for 2 h at room temperature, incubated with the appropriate secondary antibody (Wuhan Boster Biological Engineering Co., Ltd., Wuhan, China) for 1 h following membrane washing, luminized, and developed with the chemiluminescence method.

Techniques: Phospho-proteomics, Expressing, Control

Journal: Frontiers in Pharmacology

Article Title: Computational and In Vitro Analysis of Plumbagin’s Molecular Mechanism for the Treatment of Hepatocellular Carcinoma

doi: 10.3389/fphar.2021.594833

Figure Lengend Snippet: (A) Protein expression levels of LC3-I, LC3-II, p38 MAPK, and p-p38 MAPK in SMMC-7721 cells measured by Western blot after treatment with 5 μM PL, 10 μM 3-MA autophagy inhibitor, 10 μM rapamycin autophagy agonist, 10 μM SB202190, and 10 μM SB203580 p38 MAPK inhibitor. (B) LC3-I, LC3-II, p38 MAPK, and p-p38 MAPK were used as internal reference total proteins for gray value analysis. * p < 0.05, ** p < 0.01 *** p < 0.001, compared with the PL group. # p < 0.05, ## p < 0.01 ### p < 0.001 vs. the control group. (C) Protein expression levels of LC3-I, LC3-II, p38 MAPK, and p-p38 MAPK in BEL-7404 cells measured by Western blot after treatment with 5 μM PL, 10 μM 3-MA autophagy inhibitor, 10 μM rapamycin autophagy agonist, 10 μM SB202190, and 10 μM SB203580 p38 MAPK inhibitor. (D) LC3-I, LC3-II, p38 MAPK, and p-p38 MAPK were used as internal reference total proteins for gray value analysis. * p < 0.05, ** p < 0.01 *** p < 0.001, compared with the PL group. # p < 0.05, ## p < 0.01 ### p < 0.001 vs. the control group.

Article Snippet: Plumbagin (PL) was purchased from Sigma-Aldrich (St. Louis, MO, United States) with purity ≥98%; the Acridine Orange (AO)/Ethidium bromide (EB) Double Stain Kit was from Solable Technology (Beijing, China); N-acetyl-l-cysteine, SB203580, and SB202190 were from Sigma-Aldrich (St. Louis, MO, United States); SC-79, MEK2206, 3-MA, and Z-VAD-FMK were from Selleck (Texas, United States); the BCA Protein Assay Kit, ROS Assay Kit, Annexin V-FITC Apoptosis Detection Kit and Cell lysis buffer for Western were all obtained from Beyotime Biotechnology (Shanghai, China); antibodies against Akt, phospho-Akt, mTOR, phospho-mTOR, p38 MAPK, phospho-p38 MAPK, PI3K, phospho-PI3K, LC3B, cleave-RP, and cleave-caspase 3 were from Cell Signaling Technology, Inc. (Boston, MA, United States); and β-p38 MAPK was purchased from Boster Technology (Wuhan, China).

Techniques: Expressing, Western Blot, Control

Journal: PLoS Pathogens

Article Title: Sustained Activation of Akt Elicits Mitochondrial Dysfunction to Block Plasmodium falciparum Infection in the Mosquito Host

doi: 10.1371/journal.ppat.1003180

Figure Lengend Snippet: Midguts from 3–5 day old female HM myrAkt and NTG An. stephensi were dissected at 0.5 h post blood feeding and processed for western blot. Data are represented as the average fold change ± SEM of phospho-protein levels for pERK, p-p38, and p-JNK quantified by densitometry and normalized first to GAPDH to control for protein loading differences and then to phospho-protein levels in NTG controls. Data collected from 6–8 separate cohorts of A. stephensi were analyzed by Student's t-test (alpha = 0.05). P values are noted on the graph.

Article Snippet: Anti-phospho-p38 MAPK (T180/Y182) was purchased from Cayman Chemical (Ann Arbor, MI) and anti-phospho-JNK 1/2 (pT183/pY185) was purchased from Invitrogen/Life Technologies (Grand Island, NY).

Techniques: Western Blot, Control

Journal: Scientific Reports

Article Title: PiHOG1 , a stress regulator MAP kinase from the root endophyte fungus Piriformospora indica , confers salinity stress tolerance in rice plants

doi: 10.1038/srep36765

Figure Lengend Snippet: ( A ) CLUSTALW Analysis: The PiHOG1 gene encodes a member of the stress-activated MAPK family. Amino acid sequences of homologous proteins to the P. indica PiHOG1, namely Coprinopsis cinerea Sty1 protein (XP_001829398.2), Heterobasidion annosum HOG1 (AEK12774.1), Cryptococcus neoformans MAP kinase (XP_569949.1), Magnaporthe oryzae MAP kinase (XP_003714838.1), Fusarium proliferatum HOG1-like protein (ABO46009.1), Metarhizium acridum stress-activated MAP kinase (EFY85878.1), Epichloe festucae stress-activated MAP kinase (ABW75775.1), Neurospora crassa osmosensitivity protein (XP_962163.2), Schizosaccharomyces pombe Sty1 MAP kinase (NP_592843.1) and S. cerevisiae HOG1 (U53878) were aligned with the CLUSTALW software. The serine/threonine protein kinase catalytic domain is shaded by gray (25%) in which the conserved TGY phosphorylation motif is distinguished by green shade. The C-terminal common docking (CD) motif is shown in rectangle shape in which the conserved hydrophobic amino acids tyrosine (Y) and histidine (H) are underlined and conserved acidic aspartic acids (D) are dark yellow shaded. PBS2 binding domain-2 is shaded in yellow color. [ * , perfectly conserved residues, : , very similar residues, • , similar residues]. ( B ) Phylogenetic tree with branch lengths: The tree was constructed by using different stress-activated MAP kinase/HOG1/P38 amino acid sequences. Member of different groups were marked with different shape i.e. Δ: insects, □: mammals, ◊: fungi and ○: plants. PiHOG1 protein is marked with filled shape to display its position.

Article Snippet: Blot was probed with 1:5000 dilutions of polyclonal Anti-phospho-p38 MAPK (pThr180/Tyr182; Signalway Antibody, USA) for 16 hours at 4 °C.

Techniques: Software, Phospho-proteomics, Binding Assay, Construct