anti ncf1  (Santa Cruz Biotechnology)

Journal: Phytotherapy Research

Article Title: Lithospermic Acid Improves Doxorubicin‐Induced Cardiomyopathy Through Sirtuin‐3‐Mediated Deacetylation of p53

doi: 10.1002/ptr.70188

Figure Lengend Snippet: Lithospermic acid (LA) ameliorates doxorubicin (Dox)‐induced apoptosis and oxidative damage in human cardiomyocytes. (A) Schematic diagram illustrating the differentiation protocol for human embryonic stem cell (hESC)‐derived cardiomyocytes (hESC‐CMs) in chemically defined medium. (B) The expression of caspase 3, BAX, and BCL2 protein. (C) Mitochondrial membrane potential was measured using JC‐1 fluorescent probe. (D) The expression of gp91 phox , p67 phox , P‐p47 phox and p47 phox protein. (E) The reactive oxygen species (ROS) level determined by DCFH‐DA staining. (F) The mitochondrial superoxide formation examined by MitoSOX Red and Mito‐Tracker staining. (G) The expression of NFR2 in cytoplasm and nucleus of hESC‐CMs. H, The expression of HO‐1 and NQO‐1 protein. The experiments were conducted after 24‐h treatment with Dox (2 μM) in combination with vehicle (Veh) or LA (20 μM). n = 6 in B, D, H, n = 7 in C, E, F, n = 8 in G, n = 6 in H.

Article Snippet: Antibodies against p47 phox (AF5220) and P‐p47 phox (AF3167) were gained from Affinity Biosciences (Cincinnati, OH, USA).

Techniques: Derivative Assay, Expressing, Membrane, Staining

Journal: Phytotherapy Research

Article Title: Lithospermic Acid Improves Doxorubicin‐Induced Cardiomyopathy Through Sirtuin‐3‐Mediated Deacetylation of p53

doi: 10.1002/ptr.70188

Figure Lengend Snippet: Lithospermic acid (LA) improves doxorubicin (Dox)‐induced oxidative stress damage in mouse cardiomyocytes. (A and B) The reactive oxygen species (ROS) level determined by DCFH‐DA probe. (C and D) The mitochondrial superoxide formation examined by MitoSOX Red and Mito‐Tracker probe. (E) The levels of malondialdehyde (MDA). (F–H) The expression of gp91 phox , p67 phox , P‐p47 phox and p47 phox protein. (I and J) The expression of NFR2 in cytoplasm and nucleus of HL‐1 cells. (K and L) The expression of HO‐1 and NQO‐1 protein. (M) The expression of Ho‐1 and Nqo‐1 mRNA. The experiments were made after 24‐h treatment with Dox (1 μM) in combination with vehicle (Veh) or LA (20 μM). n = 8 in A‐D, n = 6 in E–H, n = 3 in I and J, n = 6 in K–M. Data represent the mean ± SEM. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001. Two‐way ANOVA was used.

Article Snippet: Antibodies against p47 phox (AF5220) and P‐p47 phox (AF3167) were gained from Affinity Biosciences (Cincinnati, OH, USA).

Techniques: Expressing

Journal: Phytotherapy Research

Article Title: Lithospermic Acid Improves Doxorubicin‐Induced Cardiomyopathy Through Sirtuin‐3‐Mediated Deacetylation of p53

doi: 10.1002/ptr.70188

Figure Lengend Snippet: Lithospermic acid (LA) prevents doxorubicin (Dox)‐induced cardiac apoptosis and oxidative damage in mice. (A and B) The expression of caspase 3, BAX, and BCL2 protein. (C) Representative images of immunohistochemical analysis of 4‐hydroxynonenal (4‐HNE, brown color) in the cardiac tissue. (D) The levels of malondialdehyde (MDA). (E and F) The expression of gp91 phox , p67 phox , P‐p47 phox and p47 phox protein. (G and H) The expression of HO‐1 and NQO‐1 protein. Measurements were made 4 weeks after the administration of LA (100 μL, 50 mg/kg per day, 30 days, i.g.) or Dox (100 μL, 5 mg/kg per times, 4 times, i.p.). n = 6 in all group. Data represent the mean ± SEM. * p < 0.05; ** p < 0.01; *** p < 0.001; **** p < 0.0001. Two‐way ANOVA was used.

Article Snippet: Antibodies against p47 phox (AF5220) and P‐p47 phox (AF3167) were gained from Affinity Biosciences (Cincinnati, OH, USA).

Techniques: Expressing, Immunohistochemical staining

Journal: Frontiers in Cellular Neuroscience

Article Title: Galectin-3 shapes microglial phenotype through endogenous and exogenous mechanisms

doi: 10.3389/fncel.2025.1729776

Figure Lengend Snippet: Analysis of microglial activation markers and inflammatory pathways under endogenous and exogenous galectin-3 modulation. (a) Flow cytometry analysis of BV2 WT and Gal3KO cells after Gal3 24 h pretreatment (preGal3) by CD11b and CD45 surface markers. 20,000 events were analyzed. Gating strategy is shown in . n = 3 per Control group and n = 4 per preGal3 group. (b) Quantification of median fluorescence intensity (MFI) of CD45 surface marker. (c) Quantification of MFI of CD11b surface marker. (d) Representative immunofluorescence images of NOX2 subunit p47phox. Scale bars, 20 μm. (e) Quantification of total p47phox content within BV2 cells. n = 4 per group. (f) TNFα concentration in cell medium measured by Mesoscale using a V-PLEX panel. TNFα was detected within the detection range consistently throughout all samples. n = 4 per group. (g) Representative Western blot images of BV2 WT and Gal3KO protein lysates. (h) Western blot quantification of TREM2, TLR4 and Clec7a expressed as the ratio divided by Actin. n = 3–4 per group. (i) Clec7a ELISA quantification. n = 5 per group. (j) Representative immunofluorescence images of phosphorylated Syk (pSyk). Scale bars, 20 μm. (k) Quantification of total pSyk content within BV2 cells. n = 4 per group. Data are shown as individual replicates with mean ±SEM (each experimental replicate corresponds to an independent culture performed from a different cell passage). In (b,c,e,f,h,i,k) , two-way ANOVA with Tukey’s multiple comparisons was performed and significant ANOVA differences are shown. p -values are expressed with 3 decimals.

Article Snippet: Primary antibodies used: TOM20 (1:500, rabbit, Proteintech, Cat: 11802-1-AP), NDUFB8 (1:250, mouse, Abcam, Cat: ab110242), p47phox (1:100, mouse, Santa Cruz Biotechnology, Cat: sc-17844), p22phox (1:100, rabbit, Abcam, Cat: ab75941), pSyk (1:250, rabbit, Cell Signaling, Cat: 2717S).

Techniques: Activation Assay, Flow Cytometry, Control, Fluorescence, Marker, Immunofluorescence, Concentration Assay, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Redox Biology

Article Title: PTH-driven modulation of platelet activity via the NOX2 pathway in postsurgical hypoparathyroidism

doi: 10.1016/j.redox.2025.103896

Figure Lengend Snippet: In-vivo evaluation of oxidative stress in HypoPT and healthy subjects . Serum levels of sNOX2-dp (A), H 2 O 2 (B) and 8-OHdG (C) in 24 HypoPT subjects and 40 healthy subjects (HS) at basal conditions. P-p47 phox /p47 phox expression in platelets from HS (N = 12) and HypoPT (N = 12) subjects at basal conditions and representative Western blot bands of p-P47 phox /P47 phox (D–E). Differences between groups were analyzed by the Mann-Whitney test (for non-normally distributed data) or t -test (for normally distributed data). ∗∗p < 0.01; ∗∗∗∗p < 0.001.

Article Snippet: Membranes were blocked for 1 h at room temperature with 5 % BSA in Tris-buffered saline containing 0.1 % Tween-20 (TBS-T) and then incubated overnight at 4 °C with a primary antibody against p47 phox (Cell Signaling Technology, Cat #4301S) and phospho-P47 phox (AMSBIO, Cat #A01586S370); followed by incubation with HRP-conjugated secondary anti-mouse (BioRad, Cat #1706516) or anti-rabbit (BioRad, Cat #1706515) antibodies.

Techniques: In Vivo, Expressing, Western Blot, MANN-WHITNEY

Journal: Redox Biology

Article Title: PTH-driven modulation of platelet activity via the NOX2 pathway in postsurgical hypoparathyroidism

doi: 10.1016/j.redox.2025.103896

Figure Lengend Snippet: In-vivo evaluation of oxidative stress in HypoPT and healthy subjects . Serum levels of sNOX2-dp (A), H 2 O 2 (B) and 8-OHdG (C) in 24 HypoPT subjects and 40 healthy subjects (HS) at basal conditions. P-p47 phox /p47 phox expression in platelets from HS (N = 12) and HypoPT (N = 12) subjects at basal conditions and representative Western blot bands of p-P47 phox /P47 phox (D–E). Differences between groups were analyzed by the Mann-Whitney test (for non-normally distributed data) or t -test (for normally distributed data). ∗∗p < 0.01; ∗∗∗∗p < 0.001.

Article Snippet: Membranes were incubated overnight at 4 °C with the following primary antibodies: PTH1R (Abcam, Cat #AB75150), PKC (Cell Signaling, Cat #46809S), phospho-PKC (Cell Signaling, Cat #9371S), p47 phox (Cell Signaling, Cat #4301S), phospho-p47 phox (AMSBIO, Cat #A01586S370), and vinculin (Cell Signaling, Cat #13901S) as a loading control.

Techniques: In Vivo, Expressing, Western Blot, MANN-WHITNEY