Journal: Current Oncology

Article Title: Overcoming of Microenvironment Protection on Primary Chronic Lymphocytic Leukemia Cells after Treatment with BTK and MDM2 Pharmacological Inhibitors

doi: 10.3390/curroncol28040223

Figure Lengend Snippet: Ibrutinib plus nutlin-3 combination mediates mitochondria-dependent apoptosis through inhibition of phospho-BAD. Patients’ derived B-CLL cells were grown in vitro with stroma or with CpG-ODN and CD40L and exposed to ibrutinib (10 µM), nutlin-3 (10 µM), ibrutinib+nutlin-3 (10 µM) for 24 h or left untreated as control. In ( A ), mitochondrial activity, evaluated using MitoTracker ® Red staining on B-CLL cells stimulated with CpG-ODN and CD40L, was analyzed by flow cytometry and expressed as fold of modulation with respect to the MFI value of the untreated B-CLL cultures set at 1. Results are reported as mean ± SD of five independent experiments. Statistical analysis was performed by ANOVA followed by Bonferroni post-hoc test. Asterisks indicate: *, p < 0.05; **, p < 0.01; ***, p < 0.001. In ( B ), Western blotting analyses of pro-caspase 9 and pro-caspase 3 protein levels are shown. In C, Western blotting analyses of BAD phosphorylation in ser136, total BAD, BAX, Akt phosphorylation in ser473 and total Akt protein levels are shown. In ( B , C ), results are shown for representative B-CLL patients and β-actin levels are shown as loading control. In ( D ), schematic representation of early molecular events that drive the mitochondria-dependent apoptosis following the combination treatment ibrutinib plus nutlin-3.

Article Snippet: The following antibodies were used for immunoblotting onto nitrocellulose filters: anti c-MYC, anti-phospho p53-Ser15, anti-phospho p53-Ser392, BAD, phospho BAD-Ser136 and p14ARF from Cell Signaling (Danvers, MA, USA); anti-p53, anti-MDM2, anti-p21, anti-PUMA, anti-procaspase 9 and anti-procaspase 3 from Santa Cruz Biotechnology (Santa Cruz, CA, USA); anti-BAX and anti-Akt from BD (BD Biosciences); phospho Akt-Ser473 from Merck Millipore (KGaA, Darmstadt, Germany) and anti β-actin, from Sigma-Aldrich (Merck KGaA).

Techniques: Inhibition, Derivative Assay, In Vitro, Control, Activity Assay, Staining, Flow Cytometry, Western Blot, Phospho-proteomics

Journal: Aging (Albany NY)

Article Title: Exploring the therapeutic mechanism of Yuebi decoction on nephrotic syndrome based on network pharmacology and experimental study

doi: 10.18632/aging.206116

Figure Lengend Snippet: Molecular docking scores of main active ingredients and targets.

Article Snippet: PPP3CA (YN2056), AKT1 (YN0514), STAT3 (YN0127), NFATC3 (YN0789), and TRPC6 (YN1223) antibodies were purchased from ImmunoWay Biotechnology Company (USA).

Techniques: Positive Control

Journal: Aging (Albany NY)

Article Title: Exploring the therapeutic mechanism of Yuebi decoction on nephrotic syndrome based on network pharmacology and experimental study

doi: 10.18632/aging.206116

Figure Lengend Snippet: Effects of YBD on the expressions of core targets in NS rats induced by PAN. ( A – F ) The relative protein expressions of PPP3CA, STAT3, NFATC3, TRPC6, and AKT1 were measured by Western blotting. As an internal control respectively, β-actin was used to calculate the quantification of protein. Data were expressed as mean ± SD, n = 6. Vertical bars represent the standard deviation. # p < 0.05 and ## p < 0.01 versus control group; * p < 0.05 and ** p < 0.01 versus model group.

Article Snippet: PPP3CA (YN2056), AKT1 (YN0514), STAT3 (YN0127), NFATC3 (YN0789), and TRPC6 (YN1223) antibodies were purchased from ImmunoWay Biotechnology Company (USA).

Techniques: Western Blot, Control, Standard Deviation

Journal: Molecules

Article Title: Anti-Metabolic Syndrome Effects of Fucoidan from Fucus vesiculosus via Reactive Oxygen Species-Mediated Regulation of JNK, Akt, and AMPK Signaling

doi: 10.3390/molecules24183319

Figure Lengend Snippet: Effects of fucoidan from Fucus vesiculosus (FvF) on relieving insulin resistance (IR) in HepG2 cells. Effects of sodium palmitate (PA) on cellular glucose consumption ( a ). Cells were treated with a concentration range of PA for 24 h. Effects of FvF on glucose consumption in IR cells ( b ). Cells were treated with Metf (2 mM) or FvF (100 μg/mL) in the presence of 100 μM PA for 24 h. ( c ). Reactive oxygen species (ROS) was detected by in situ dihydroethidium (DHE) staining (200×). C, control group; M, cells were treated with 100 μM PA for 24 h; Metf and FvF, cells were treated with metformin (2 mM) or FvF (100 μg/mL) in the presence of 100 μM PA for 24 h. Phosphorylation of c-Jun N-terminal kinase (pJNK) ( d ) and phosphorylation of protein kinase B (pAkt) ( e ) protein levels changed between different treatment groups. C, control group; M, cells treated with 100 μM PA for 24 h; Metf and FvF, cells treated with 100 μM PA for 24 h then incubated with metformin (2 mM) or FvF (100 μg/mL) for another 6 h. Data are expressed as the mean ± SEM. Differences were assessed by ANOVAs and statistical results are denoted as follows: * p < 0.05 versus the control group; # p < 0.05 versus the model group.

Article Snippet: After that, HepG2 cells were rinsed with PBS and lysed in ice-cold RIPA buffer containing protease inhibitor and phosphatase inhibitors (Roche) for 30 min. After denaturation with 5× loading buffer at 100 °C for 10 min, proteins were electrophoresed on 10% SDS-PAGE and transferred to a PVDF membrane (0.22 μm) that was subsequently blocked with 5% ( w / v ) non-fat milk in Tris-buffered saline containing 0.1% Tween-20 (TBST) for 2 h. The membranes were incubated with primary antibodies directed against JNK, pJNK (T183/T185), Akt, pAkt (S473), AMPK, pAMPK (T172), HMGCR, pHMGCR (S872) (Merck Millipore, Billerica, MA, USA), SREBP-1C (Abcam, HongKang, China), ACC, pACC (S79), and β-actin antibodies in 5% ( w / v ) BSA in TBST overnight at 4 °C.

Techniques: Concentration Assay, In Situ, Staining, Control, Phospho-proteomics, Incubation

Journal: International Journal of Molecular Sciences

Article Title: Protective Effect of Acteoside on Ovariectomy-Induced Bone Loss in Mice

doi: 10.3390/ijms20122974

Figure Lengend Snippet: Acteoside treatment on the expression levels of TRAF6 ( A ), RANKL ( B ), RANK ( C ), NFKBIA ( D ), PI3K ( E ), AKT ( F ), IKKβ ( G ), NFAT2 ( H ), and c-Fos ( I ) ( n = 3); β-actin was shown as the loading control, and quantitative data of every signal protein was descriptive as percentages of the value of control. Values were expressed as the mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001 relative to the control group.

Article Snippet: Co. Ltd., Nanjing, China; primary antibodies including TRAF6, RANKL, RANK, NFKBIA, PI3K, AKT, IKKβ, NFAT2, c-Fos, β-actin, and secondary antibodies of horseradish peroxidase-conjugated goat anti-rabbit IgG were offered by ZSGB-BIO, Beijing, China; all other reagents were of analytical purity.

Techniques: Expressing, Control

Journal: Molecular Medicine Reports

Article Title: Lithium prevents glucocorticoid‑induced chondrocyte autophagy: An in vitro study

doi: 10.3892/mmr.2023.13070

Figure Lengend Snippet: Results of western blot analysis. (A) Representative western blots of rat chondrocytes. The average relative expression of (B) LC3II/I, (C) p-AKT/AKT and (D) p-mTOR/mTOR from rat chondrocytes. (E) Representative western blots of human chondrocytes. The average relative expression of (F) LC3II/I, (G) p-AKT/AKT, and (H) p-mTOR/mTOR from human chondrocytes. *P<0.05, **P<0.01, ***P<0.001 and ****P<0.0001. Con, control group; DEX, dexamethasone group (200 µM); DEX + Li, dexamethasone (200 µM) combined with lithium chloride (10 mM) group; p-AKT, phosphorylated AKT; p-mTOR, phosphorylated mTOR; p-, phosphorylated.

Article Snippet: Subsequently, the membranes were incubated overnight at 4°C with primary antibodies against LC3B (rabbit; 1:2,000; cat. no. ab192890; Abcam), phosphorylated (p-)AKT (rabbit; 1:1,000; cat. no. 310021; Chengdu Zen Bioscience Co., Ltd.), AKT (rabbit; 1:1,000; cat. no. 382804; Chengdu Zen Bioscience Co., Ltd.), phosphorylated mTOR (mouse; 1:1,000; cat. no. sc-293133; Santa Cruz Biotechnology, Inc.), mTOR (mouse; 1:1,000; cat. no. sc-517464; Santa Cruz Biotechnology, Inc.), and β-tubulin (rabbit; 1:2,000; cat. no. AF7011; Affinity Biosciences).

Techniques: Western Blot, Expressing, Control