Journal: Molecules

Article Title: Cell Death Effects Induced by Sulforaphane and Allyl Isothiocyanate on P-Glycoprotein Positive and Negative Variants in L1210 Cells

doi: 10.3390/molecules25092093

Figure Lengend Snippet: Expression of proteins that are active in apoptosis (Bcl–2, Bax, and the members of NF-κB pathways: p50, P52 and p65) in S, R and T cells after 24 h of incubation in medium containing given concentrations of SFN or AITC. Panels A and B : Detection of proteins by Western blotting. GAPDH was used as an internal control. Data are representative of three independent measurements. Panels C and D : Densitometric quantification of protein bands from Western blots. Quantities are expressed as the mean ± S.E.M. of three independent measurements of data relative to the GAPDH signal. Data differ from values obtained in cells that were not incubated in the absence of either SFN or AITC at the levels: # – p < 0.05; * – p < 0.01.

Article Snippet: The primary antibodies against LC3B (L7543, Sigma-Aldrich, Saint Louis, MO, USA) diluted 1:300, Bax (N-20), Bcl2 (N-19), NFκ-B p52 (K-27), NFκ-B p50 (H-119), NFκ-B p65 (C-20), all from Santa Cruz Biotechnology, Dallas, TX, USA, diluted 1:100 and GAPDH (MAB374, EMD Millipore Chemicals, Billerica, MA, USA), were added to the blocking buffer containing 5% defatted milk overnight at 4 °C.

Techniques: Expressing, Incubation, Western Blot, Control

Journal: Journal of dairy science

Article Title: Sodium butyrate reduces bovine mammary epithelial cell inflammatory responses induced by exogenous lipopolysaccharide, by inactivating NF-κB signaling.

doi: 10.3168/jds.2020-18189

Figure Lengend Snippet: Figure 5. Effects of sodium butyrate (SB) on NF-κB signaling pathway. Bovine mammary epithelial cells (MAC-T) were treated with 1 µg/mL LPS for 6 h; with 0.5 mM SB for 18 h; or with 0.5 mM SB for 18 h, followed by a challenge with 1 µg/mL LPS for an additional 6 h. (A) Western blot analysis of p-IκB α, IκB α, p-p65, and p65. (B) Relative protein expression levels of p-IκB α to IκB α. (C) Relative protein expression levels of p-p65 to p65. Data from the control group were used to normalize the data of each treatment group. Comparisons among groups were calculated using a one-way ANOVA with subsequent Bonferroni correction. Data presented are mean ± SEM. Different lowercase letters in bar charts indicate significant differences (P < 0.05).

Article Snippet: Membranes were then incubated with antibodies against phosphorylatd-IκB α (p-IκB α; sc-271980, Santa Cruz Biotechnology; 1:5,000), IκB α (sc-166588, Santa Cruz Biotechnology; 1:1,000), NFκB p65 (sc-8008, Santa Cruz Biotechnology; 1:1,000), phosphorylatd-NF-κB p65 (p-NF-κB p65; sc-166748, Santa Cruz Biotechnology; 1:1,000), acetyl-H3 (8173, Cell Signaling Technology, Danvers, MA; 1:1,000), histone H3 (4499, Cell Signaling Technology; 1:1,000), and GAPDH (ab8245, Abcam, Cambridge, MA; 1:5,000) overnight at 4°C, followed by incubation with horseradish peroxidase-conjugated anti-mouse or anti-rabbit antibody (Boster, Wuhan, China) for 45 min at room temperature.

Techniques: Western Blot, Expressing, Control

Journal: Frontiers in Nutrition

Article Title: Cannabis oil modulates liver alterations and endocannabinoid system changes in a female rat model of diet-induced MASLD

doi: 10.3389/fnut.2026.1770150

Figure Lengend Snippet: Liver NrF2 and NFκB p65 expression in female rats fed a reference diet (RD), sucrose-rich diet (SRD) or SRD with cannabis oil (SRD + Ca). (A) Representative photomicrographs of immunocytochemical staining NrF2 in the liver sections of rats. Decreased levels of nuclear and cytoplasmic positive markers are observed in the SRD group. Scale bar 50 μm. (B) Quantitative immunohistochemical analysis of liver NrF2 expression expressed as integrated optical density (IOD). (C) Representative photomicrographs of immunocytochemical staining NFκB p65 in the liver sections of rats. Increased levels of nuclear and cytoplasmic positive markers are observed in the SRD group. Scale bar 50 μm. (D) Quantitative immunohistochemical analysis of NFκB p65 expression in the liver. Data are expressed as mean ± SEM ( n = 6). Statistical differences were evaluated by one-way ANOVA followed by the Newman–Keuls post hoc test (* P < 0.05).

Article Snippet: The samples were incubated in a humid chamber first with a specific primary antibody for 4-HNE (mouse monoclonal antibody; Catalog # MAB3249; R&D Systems), NrF2 (mouse monoclonal antibody; sc-365949; Santa Cruz Biotechnology), NF-κB p65 (mouse monoclonal antibody; sc-8008; Santa Cruz Biotechnology) and TGF-β1 (mouse monoclonal antibody; sc-52893; Santa Cruz Biotechnology) (for 14–16 h at 4°C) and then with biotin-conjugated secondary antibody (anti-mouse, 1:100 dilution, Sigma) for 30 min at room temperature.

Techniques: Expressing, Cannabis, Staining, Immunohistochemical staining

Journal: Journal of cell science

Article Title: The laminin-binding integrins regulate nuclear factor κB-dependent epithelial cell polarity and inflammation.

doi: 10.1242/jcs.259161

Figure Lengend Snippet: Fig. 7. α3−/−α6−/−collecting duct cells exhibit increased NF-κB activation. (A) α3f/fα6f/f and α3−/−α6−/−collecting duct cells were grown to 80% confluence. Then, 24 h later, conditioned medium from these cells was collected and subjected to cytokine multiplex analysis. A subset of cytokines with significant differences between α3−/−α6−/−and α3f/fα6f/f collecting duct cells (P<0.01) are shown. (B-D) α3f/fα6f/f and α3−/−α6−/−collecting duct cells grown to 80% confluence were lysed and subjected to separation of nuclear and cytosolic fractions. Western blot analysis of the fractions (20 µg total protein/lane) showed increased expression of nuclear p65 and cytosolic phospho-p65S536 (p-p65S536) in α3−/−α6−/−compared to α3f/fα6f/f collecting duct cells. Lamin A/C was used as a nuclear marker; β-actin was used as a loading control (B). The expression of p65 (C) or p-p65S536 (D) was normalized to β-actin and quantified as a ratio of α3f/fα6f/f cells. (E) α3f/fα6f/f and α3−/−α6−/−collecting duct cells were grown to 80% confluence. Then, 24 h later, NF-kB activity in these cells was detected using a Nano-Glo Luciferase Assay and quantified as a ratio of α3f/fα6f/f collecting duct cells. (F) α3f/fα6f/f and α3−/−α6−/−collecting duct cells grown to 80% confluence were treated with solvent (3% Tween 80 in regular growth medium) or 10 µM BMS-345541 in solvent for 1 h. NF-kB activity was detected using a Nano-Glo Luciferase Assay and quantified as a ratio of α3f/fα6f/f collecting cells. (G,H) Immunostaining for the collecting duct marker DBA (green) and phospho-p65S276

Article Snippet: In some experiments, the collecting duct cells were treated with IκB kinase inhibitor BMS-34554 (100 μM in water/3% Tween 80) or the vehicle alone for 1 h. To silence NFκB p65, collecting duct cells were transfected with non-silencing siRNA (20 nM, transfection control) or p65-silencing siRNA [NFκB p65 siRNA (m), 29411, Santa Cruz Biotechnology] using Lipofectamine RNAiMAX according to manufacturer’s instructions.

Techniques: Activation Assay, Multiplex Assay, Western Blot, Expressing, Marker, Control, Activity Assay, Luciferase, Solvent, Immunostaining

Journal: Journal of cell science

Article Title: The laminin-binding integrins regulate nuclear factor κB-dependent epithelial cell polarity and inflammation.

doi: 10.1242/jcs.259161

Figure Lengend Snippet: Fig. 8. The mesenchymal phenotype of α3−/−α6−/−collecting duct cells is mediated by NF-κB-dependent Snail and Slug induction. (A-C) α3f/fα6f/f and α3−/−α6−/−collecting duct cells grown to 80% confluence were lysed and subjected to separation of nuclear and cytosolic fractions. Western blot analysis of the fractions (20 µg total protein/lane) showed increased nuclear Snail and Slug in α3−/−α6−/−collecting duct cells compared to α3f/f/α6f/f collecting duct cells. Lamin A/C was used as a nuclear marker; β-actin was used as a loading control (A). The expression of Snail (B) or Slug (C) was normalized to β-actin and quantified as a ratio of α3f/fα6f/f cells. **P<0.01 between α3−/−α6−/−and α3f/fα6f/f collecting duct cells. (D-H) α3−/−α6−/−collecting duct cells grown to 80% confluence were transfected with non-silencing (n) or NF-κB p65-silencing (p65) siRNA for 48 h. Collecting duct cells were lysed and subjected to separation of nuclear and cytosolic fractions. Western blot analysis of the fractions (20 µg total protein/lane) showed decreased cytosolic and nuclear phospho-p65S536 (p-p65), total p65 and Slug, and decreased nuclear Snail in cells transfected with p65 siRNA compared to non-silencing siRNA. Lamin A/C was used as a nuclear marker; β-actin was used as a loading control (D). The expression of p-p65 (E), p65 (F), Snail (G) or Slug (H) was normalized to β-actin and quantified as a ratio of cells transfected with non-silencing siRNA. **P<0.01 between cells transfected with non-silencing and p65 siRNA. (I-M) α3f/f/α6f/f and α3−/−α6−/−collecting duct cells grown to 80% confluence were transfected with non-silencing or NF-κB p65-silencing (p65) siRNA, and lysed 48 h later. Total cell lysates (40 µg total protein/lane) were analyzed by western blotting for levels of p65, E-cadherin, N-cadherin and collagen I; β-actin was used as a loading control (I). The expression of p65 (J), E-cadherin (K), N-cadherin (L) or collagen I (M) proteins was normalized to actin and quantified as ratio of α3f/fα6f/f cells transfected with non-silencing siRNA. Data are mean±s.e.m. of at least three experiments. **P<0.01 between non-silencing and p65 siRNA-transfected α3f/f/α6f/f or α3−/−/α6−/−collecting duct cells. Statistical significance was determined using two-tailed, unpaired t-tests.

Article Snippet: In some experiments, the collecting duct cells were treated with IκB kinase inhibitor BMS-34554 (100 μM in water/3% Tween 80) or the vehicle alone for 1 h. To silence NFκB p65, collecting duct cells were transfected with non-silencing siRNA (20 nM, transfection control) or p65-silencing siRNA [NFκB p65 siRNA (m), 29411, Santa Cruz Biotechnology] using Lipofectamine RNAiMAX according to manufacturer’s instructions.

Techniques: Western Blot, Marker, Control, Expressing, Transfection, Two Tailed Test

Journal: Frontiers in Nutrition

Article Title: Astaxanthin alleviates altered hepatic lipid metabolism and oxidative stress in animals fed a high-sucrose diet

doi: 10.3389/fnut.2026.1781406

Figure Lengend Snippet: (A) Representative photomicrographs of p-NFκB p65 immunocytochemical staining in liver tissues (400 × magnification), and (B) quantitative immunohistochemical analysis of p-NFκB p65 expression in the liver of rats fed a reference diet (RD), RD plus ASTX (RD + ASTX), high-sucrose diet (HSD), and HSD plus ASTX (HSD + ASTX). Values (mean ± SEM, n = 6 ) are expressed as the in tegrated optical density (IOD). Values that do not share the same letter were significantly different ( p < 0.05) when one variable at a time was compared by two-way ANOVA. D, effect of Diet. I, effect of intervention. Int (D x I), interaction between diet x intervention. ns, not significant.

Article Snippet: The samples were incubated in a humid chamber first with a specific primary antibody for Nrf2 (mouse monoclonal antibody; sc-365949; Santa Cruz Biotechnology) and p-NFκB p65 (mouse monoclonal antibody; sc-136548; Santa Cruz Biotechnology) (for 14–16 h at 4 °C) and then with biotin-conjugated secondary antibody (antimouse, 1:100 dilution, Sigma) for 30 min at room temperature.

Techniques: Staining, Immunohistochemical staining, Expressing

Journal: Frontiers in Nutrition

Article Title: Astaxanthin alleviates altered hepatic lipid metabolism and oxidative stress in animals fed a high-sucrose diet

doi: 10.3389/fnut.2026.1781406

Figure Lengend Snippet: Mechanisms by which ASTX modulates liver lipid metabolism and oxidative stress in HSD-fed rats. ASTX supplementation mitigates hepatic injury by reducing steatosis and triglyceride accumulation, mainly through enhanced mitochondrial fatty acid β-oxidation (↑ CPT-1 activity) without affecting lipogenesis. In parallel, ASTX improves hepatic redox homeostasis by lowering ROS levels, increasing antioxidant enzyme activities (CAT, GST) and restoring GSH content. Increased Nrf2 and decreased p-NFκB p65 protein expression were also modulated by ASTX treatment. Solid lines indicate effects supported by the experimental data, whereas dashed lines represent suggested pathways not directly evaluated in this study.

Article Snippet: The samples were incubated in a humid chamber first with a specific primary antibody for Nrf2 (mouse monoclonal antibody; sc-365949; Santa Cruz Biotechnology) and p-NFκB p65 (mouse monoclonal antibody; sc-136548; Santa Cruz Biotechnology) (for 14–16 h at 4 °C) and then with biotin-conjugated secondary antibody (antimouse, 1:100 dilution, Sigma) for 30 min at room temperature.

Techniques: Activity Assay, Expressing