Journal: Frontiers in Immunology

Article Title: Platelet-rich plasma-derived microRNA let-7a-5p alleviates knee osteoarthritis by regulating macrophage polarization and improving inflammatory microenvironment

doi: 10.3389/fimmu.2026.1756467

Figure Lengend Snippet: Validation of the targeting relationship between let-7a-5p and MAPK8. (A) Schematic representation of the predicted complementary binding site between let-7a-5p and the 3′-UTR of MAPK8. (B) Relative mRNA expression level of MAPK8 in macrophages overexpressing let-7a-5p detected by RT-qPCR. (C) WB analysis of MAPK8 protein expression in macrophages overexpressing let-7a-5p. (D) Inhibitory effect of let-7a-5p on MAPK8 expression was assessed via a dual luciferase reporter assay. ( **P < 0.01, ***P < 0.001).

Article Snippet: Subsequently, the tissue sections and cells were incubated overnight at 4°C with primary antibodies against iNOS (Proteintech 22226-1-AP), CD206 (Proteintech 18704-1-AP) and MAPK8 (Proteintech 66210-1-Ig).

Techniques: Biomarker Discovery, Binding Assay, Expressing, Quantitative RT-PCR, Luciferase, Reporter Assay

Journal: Frontiers in Immunology

Article Title: Platelet-rich plasma-derived microRNA let-7a-5p alleviates knee osteoarthritis by regulating macrophage polarization and improving inflammatory microenvironment

doi: 10.3389/fimmu.2026.1756467

Figure Lengend Snippet: Effect of PRP on expression of let-7a-5p and MAPK8. (A) Relative mRNA expression levels of let-7a-5p in knee joint sections of each group detected by RT-qPCR. (B) iNOS and CD206 co-immunolabeld with MAPK8 and counter-stained with DAPI in synovial tissues (Scale bar: 50 μm). (C, D) Quantification of iNOS and CD206 expression co-localized with MAPK8. ( **P < 0.01).

Article Snippet: Subsequently, the tissue sections and cells were incubated overnight at 4°C with primary antibodies against iNOS (Proteintech 22226-1-AP), CD206 (Proteintech 18704-1-AP) and MAPK8 (Proteintech 66210-1-Ig).

Techniques: Expressing, Quantitative RT-PCR, Staining

Journal: Frontiers in Immunology

Article Title: Platelet-rich plasma-derived microRNA let-7a-5p alleviates knee osteoarthritis by regulating macrophage polarization and improving inflammatory microenvironment

doi: 10.3389/fimmu.2026.1756467

Figure Lengend Snippet: The let-7a-5p/MAPK8 axis regulates macrophage polarization and inflammatory cytokine release in vitro . (A) IF staining showing expression of iNOS and CD206 in macrophages (Scale bar: 50 μm). (B) Quantification of iNOS-positive cell rate in transfected macrophages. (C) Quantification of CD206-positive cell rate in transfected macrophages. (D-G) Relative mRNA expression levels of pro-inflammatory cytokine (IL-1β and TNF-α) and anti-inflammatory cytokine (IL-4 and IL-10) in transfected macrophages detected by RT-qPCR. ( *P < 0.05, **P < 0.01, ns no significance).

Article Snippet: Subsequently, the tissue sections and cells were incubated overnight at 4°C with primary antibodies against iNOS (Proteintech 22226-1-AP), CD206 (Proteintech 18704-1-AP) and MAPK8 (Proteintech 66210-1-Ig).

Techniques: In Vitro, Staining, Expressing, Transfection, Quantitative RT-PCR

Journal: FEBS letters

Article Title: JNK1 inhibits transcriptional and pro-apoptotic activity of TAp63γ.

doi: 10.1016/j.febslet.2015.10.028

Figure Lengend Snippet: Fig. 1. JNK1 directly binds to TAp63c at its transactivation domain. (A), Hela cells transfected with Flag-JNK1 were lysed and subjected to GST pull-down assay with GST alone or GST-TAD. 10 lg of total protein from the same cell lysates were directly loaded as an input control. The adsorbates were analyzed by immunoblotting analysis (IB) using anti-Flag (top panel). Comparable amounts of GST and GST-TAD fusion proteins were shown by staining the membrane with Coomassie blue R-250 (bottom panel). (B and C) H1299 cells were co-transfected with HA-TAp63c and Flag-JNK1. Whole-cell lysates were subjected to immunoprecipitation (IP) with normal mouse IgG, anti-HA or anti-Flag. IP products were subjected to IB analysis with indicated primary antibodies and trueblot secondary antibody.

Article Snippet: JNK1 siRNA or scrambled control (Santa Cruz) was transfected with Lipofectamin 2000 to knock down endogenous JNK1.

Techniques: Transfection, Pull Down Assay, Control, Western Blot, Staining, Membrane, Immunoprecipitation

Journal: FEBS letters

Article Title: JNK1 inhibits transcriptional and pro-apoptotic activity of TAp63γ.

doi: 10.1016/j.febslet.2015.10.028

Figure Lengend Snippet: Fig. 2. Overexpression of JNK1 inhibits TAp63c transactivity. (A) H1299 cells were transfected with a mixture of Bax-Luc and TK-Renilla plus HA-TAp63c or (and) Flag-JNK1 plasmid; (B) H1299 cells were transfected with a mixture of Bax-Luc and TK-Renilla plus HA-TAp63c and increasing amounts of Flag-JNK1 plasmid. Firefly and Renilla luciferase activities were measured. The Bax-Luc activity was normalized to Renilla activity and presented as percentage of Bax-Luc expression with standard deviation (n = 3). Bax-Luc expression in cells transfected with TAp63c alone with Bax-Luc/TK-Renilla mixture was set as 100%. IB analysis was performed to detect the expression of TAp63c and JNK1 using anti-HA and anti-Flag respectively.

Article Snippet: JNK1 siRNA or scrambled control (Santa Cruz) was transfected with Lipofectamin 2000 to knock down endogenous JNK1.

Techniques: Over Expression, Transfection, Plasmid Preparation, Luciferase, Activity Assay, Expressing, Standard Deviation

Journal: FEBS letters

Article Title: JNK1 inhibits transcriptional and pro-apoptotic activity of TAp63γ.

doi: 10.1016/j.febslet.2015.10.028

Figure Lengend Snippet: Fig. 3. Ser12 in TAp63c is critical for JNK1-mediated trans-inhibition. H1299 cells were transfected with Bax-Luc/TK-Renilla mixture plus HA-TAp63c (wild-type or S12A mutant) or (and) Flag-JNK1 plasmid. Firefly and Renilla luciferase activities were measured. The Bax-Luc activity was normalized to Renilla activity and presented as percentage of Bax-Luc expression with standard deviation (n = 3). Bax-Luc expression in cells transfected with wild-type TAp63c alone with Bax-Luc/ TK-Renilla mixture was set as 100 percent. IB analysis was performed to detect the expression of TAp63c and JNK1 using anti-HA and anti-Flag respectively.

Article Snippet: JNK1 siRNA or scrambled control (Santa Cruz) was transfected with Lipofectamin 2000 to knock down endogenous JNK1.

Techniques: Inhibition, Transfection, Mutagenesis, Plasmid Preparation, Luciferase, Activity Assay, Expressing, Standard Deviation

Journal: FEBS letters

Article Title: JNK1 inhibits transcriptional and pro-apoptotic activity of TAp63γ.

doi: 10.1016/j.febslet.2015.10.028

Figure Lengend Snippet: Fig. 4. Inhibition or knockdown of JNK1 increases TAp63c transactivity. (A) H1299 cells were transfected with Bax-Luc/TK-Renilla mixture with or without HA-TAp63c, and treated with JNK inhibitor SP600125 (JNKin); (B) H1299 cells were transfected with Bax-Luc/TK-Renilla mixture plus HA-TAp63c or (and) siJNK1. Firefly and Renilla luciferase activities were measured. The Bax-Luc activity was normalized to Renilla activity and presented as percentage of Bax-Luc expression with standard deviation (n = 3). Bax-Luc expression in cells transfected with TAp63c alone with Bax-Luc/TK-Renilla mixture was set as 100%. IB analysis was performed to detect the expression of TAp63c, total or phosphorylated (p-JNK1) JNK1 using anti-HA, antibody specific to total or T183-phosphorylated JNK1, respectively.

Article Snippet: JNK1 siRNA or scrambled control (Santa Cruz) was transfected with Lipofectamin 2000 to knock down endogenous JNK1.

Techniques: Inhibition, Knockdown, Transfection, Luciferase, Activity Assay, Expressing, Standard Deviation

Journal: FEBS letters

Article Title: JNK1 inhibits transcriptional and pro-apoptotic activity of TAp63γ.

doi: 10.1016/j.febslet.2015.10.028

Figure Lengend Snippet: Fig. 5. JNK1 inhibits TAp63c-mediated cell apoptosis. H1299 cells were transfected with HA-TAp63c and (or) Flag-JNK1. 48 h post transfection, cells were trypsinized and subjected to IB (A) or flow cytometry analysis (B) of apoptosis. Cleaved PARP1 (CL-PARP), JNK1 and TAp63c were detected with antibody to N-terminal cleaved PARP1, Flag, or HA respectively.

Article Snippet: JNK1 siRNA or scrambled control (Santa Cruz) was transfected with Lipofectamin 2000 to knock down endogenous JNK1.

Techniques: Transfection, Cytometry

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