p67phox  (Proteintech)

Journal: Translational Cancer Research

Article Title: Prognostic value of palmitoylation-regulated mechanisms in glioblastoma: integrated multi-omics analysis via least absolute shrinkage and selection operator (LASSO) regression and single-cell sequencing

doi: 10.21037/tcr-2025-1953

Figure Lengend Snippet: Expression of GABRB2, NCF2, and GRIN2A in GBM. (A-C) Differential expression of the three candidate genes between GBM and normal tissues. (D-F) Correlation analyses between the three candidate genes and palmitoylation-related genes. (G-I) IHC staining for the protein expression levels of GABRB2, NCF2, and GRIN2A in the human cerebral cortex and glioma derived from HPA. The original data are available from HPA v25 ( https://v25.proteinatlas.org/ ) with the following specific URLs: GABRB2 ( https://www.proteinatlas.org/ENSG00000145864-GABRB2/brain/cerebral+cortex ; https://www.proteinatlas.org/ENSG00000145864-GABRB2/cancer/glioma ), NCF2 ( https://www.proteinatlas.org/ENSG00000116701-NCF2/brain/cerebral+cortex ; https://www.proteinatlas.org/ENSG00000116701-NCF2/cancer/glioma ), GRIN2A ( https://www.proteinatlas.org/ENSG00000183454-GRIN2A/brain/cerebral+cortex ; https://www.proteinatlas.org/ENSG00000183454-GRIN2A/cancer/glioma ). Scale bar, 100 μm. *, P<0.05. GBM, glioblastoma multiforme; HPA, Human Protein Atlas; IHC, immunohistochemical; TPM, transcript per million.

Article Snippet: Following blocking with 5% non-fat milk in tris-buffered saline with Tween 20 (TBST) for 1 hour at room temperature, membranes were incubated overnight at 4 °C with the following primary antibodies: anti- GABRB2 (1:1,000, Affinity Biosciences, Changzhou, China; DF6671), anti- NCF2 (1:1,000, Proteintech, Rosemont, IL, USA; 15551-1-AP), anti- GRIN2A (1:1,000, Proteintech, 28525-1-AP), and anti- GAPDH (1:1,000, Proteintech, 10491-1-AP).

Techniques: Expressing, Quantitative Proteomics, Immunohistochemistry, Derivative Assay, Immunohistochemical staining

Journal: Translational Cancer Research

Article Title: Prognostic value of palmitoylation-regulated mechanisms in glioblastoma: integrated multi-omics analysis via least absolute shrinkage and selection operator (LASSO) regression and single-cell sequencing

doi: 10.21037/tcr-2025-1953

Figure Lengend Snippet: GABRB2 , NCF2 , and GRIN2A regulate the proliferation and migration of glioblastoma cells via colony formation assay and transwell migration assay. (A-F) The colony formation assay was performed in LN229 and U251 cells when GABRB2 or GRIN2A was overexpressed, or NCF2 was knocked down (n=3). Overexpression of GABRB2 , GRIN2A decreases colony formation ability in LN229 and U251 cells. Knockdown of NCF2 decreases colony formation ability in LN229 and U251 cells. (G-L) Transwell assay was performed to assess the migration ability of LN229 and U251 cells when GABRB2 or GRIN2A was overexpressed, or NCF2 was knocked down (n=3). Overexpression of GABRB2 , GRIN2A decreases cell migration in LN229 and U251 at 24 h. Knockdown of NCF2 reduces cell migration in LN229 and U251. Stained with crystal violet. Scale bar, 100 μm. **, P<0.01; ***, P<0.001; ****, P<0.0001. oe-NC, overexpression plasmid negative control; si-NC, small interfering RNA negative control.

Article Snippet: Following blocking with 5% non-fat milk in tris-buffered saline with Tween 20 (TBST) for 1 hour at room temperature, membranes were incubated overnight at 4 °C with the following primary antibodies: anti- GABRB2 (1:1,000, Affinity Biosciences, Changzhou, China; DF6671), anti- NCF2 (1:1,000, Proteintech, Rosemont, IL, USA; 15551-1-AP), anti- GRIN2A (1:1,000, Proteintech, 28525-1-AP), and anti- GAPDH (1:1,000, Proteintech, 10491-1-AP).

Techniques: Migration, Colony Assay, Transwell Migration Assay, Over Expression, Knockdown, Transwell Assay, Staining, Plasmid Preparation, Negative Control, Small Interfering RNA

Journal: Translational Cancer Research

Article Title: Prognostic value of palmitoylation-regulated mechanisms in glioblastoma: integrated multi-omics analysis via least absolute shrinkage and selection operator (LASSO) regression and single-cell sequencing

doi: 10.21037/tcr-2025-1953

Figure Lengend Snippet: Expression of GABRB2, NCF2, and GRIN2A in GBM. (A-C) Differential expression of the three candidate genes between GBM and normal tissues. (D-F) Correlation analyses between the three candidate genes and palmitoylation-related genes. (G-I) IHC staining for the protein expression levels of GABRB2, NCF2, and GRIN2A in the human cerebral cortex and glioma derived from HPA. The original data are available from HPA v25 ( https://v25.proteinatlas.org/ ) with the following specific URLs: GABRB2 ( https://www.proteinatlas.org/ENSG00000145864-GABRB2/brain/cerebral+cortex ; https://www.proteinatlas.org/ENSG00000145864-GABRB2/cancer/glioma ), NCF2 ( https://www.proteinatlas.org/ENSG00000116701-NCF2/brain/cerebral+cortex ; https://www.proteinatlas.org/ENSG00000116701-NCF2/cancer/glioma ), GRIN2A ( https://www.proteinatlas.org/ENSG00000183454-GRIN2A/brain/cerebral+cortex ; https://www.proteinatlas.org/ENSG00000183454-GRIN2A/cancer/glioma ). Scale bar, 100 μm. *, P<0.05. GBM, glioblastoma multiforme; HPA, Human Protein Atlas; IHC, immunohistochemical; TPM, transcript per million.

Article Snippet: Following blocking with 5% non-fat milk in tris-buffered saline with Tween 20 (TBST) for 1 hour at room temperature, membranes were incubated overnight at 4 °C with the following primary antibodies: anti- GABRB2 (1:1,000, Affinity Biosciences, Changzhou, China; DF6671), anti- NCF2 (1:1,000, Proteintech, Rosemont, IL, USA; 15551-1-AP), anti- GRIN2A (1:1,000, Proteintech, 28525-1-AP), and anti- GAPDH (1:1,000, Proteintech, 10491-1-AP).

Techniques: Expressing, Quantitative Proteomics, Immunohistochemistry, Derivative Assay, Immunohistochemical staining

Journal: Translational Cancer Research

Article Title: Prognostic value of palmitoylation-regulated mechanisms in glioblastoma: integrated multi-omics analysis via least absolute shrinkage and selection operator (LASSO) regression and single-cell sequencing

doi: 10.21037/tcr-2025-1953

Figure Lengend Snippet: GABRB2 , NCF2 , and GRIN2A regulate the proliferation and migration of glioblastoma cells via colony formation assay and transwell migration assay. (A-F) The colony formation assay was performed in LN229 and U251 cells when GABRB2 or GRIN2A was overexpressed, or NCF2 was knocked down (n=3). Overexpression of GABRB2 , GRIN2A decreases colony formation ability in LN229 and U251 cells. Knockdown of NCF2 decreases colony formation ability in LN229 and U251 cells. (G-L) Transwell assay was performed to assess the migration ability of LN229 and U251 cells when GABRB2 or GRIN2A was overexpressed, or NCF2 was knocked down (n=3). Overexpression of GABRB2 , GRIN2A decreases cell migration in LN229 and U251 at 24 h. Knockdown of NCF2 reduces cell migration in LN229 and U251. Stained with crystal violet. Scale bar, 100 μm. **, P<0.01; ***, P<0.001; ****, P<0.0001. oe-NC, overexpression plasmid negative control; si-NC, small interfering RNA negative control.

Article Snippet: Following blocking with 5% non-fat milk in tris-buffered saline with Tween 20 (TBST) for 1 hour at room temperature, membranes were incubated overnight at 4 °C with the following primary antibodies: anti- GABRB2 (1:1,000, Affinity Biosciences, Changzhou, China; DF6671), anti- NCF2 (1:1,000, Proteintech, Rosemont, IL, USA; 15551-1-AP), anti- GRIN2A (1:1,000, Proteintech, 28525-1-AP), and anti- GAPDH (1:1,000, Proteintech, 10491-1-AP).

Techniques: Migration, Colony Assay, Transwell Migration Assay, Over Expression, Knockdown, Transwell Assay, Staining, Plasmid Preparation, Negative Control, Small Interfering RNA

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 BAX (50599‐2‐lg), BCL2 (26593‐2‐AP), gp91 phox (19013‐1‐AP), p67 phox (15551‐1‐AP), SIRT1 (13161‐1‐AP), SIRT3 (10099‐1‐AP), SIRT6 (13572‐1‐AP), GAPDH (60004‐1‐lg), and β‐tubulin (10094‐1‐AP) were bought from ProteinTech (Chicago, 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 BAX (50599‐2‐lg), BCL2 (26593‐2‐AP), gp91 phox (19013‐1‐AP), p67 phox (15551‐1‐AP), SIRT1 (13161‐1‐AP), SIRT3 (10099‐1‐AP), SIRT6 (13572‐1‐AP), GAPDH (60004‐1‐lg), and β‐tubulin (10094‐1‐AP) were bought from ProteinTech (Chicago, 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 BAX (50599‐2‐lg), BCL2 (26593‐2‐AP), gp91 phox (19013‐1‐AP), p67 phox (15551‐1‐AP), SIRT1 (13161‐1‐AP), SIRT3 (10099‐1‐AP), SIRT6 (13572‐1‐AP), GAPDH (60004‐1‐lg), and β‐tubulin (10094‐1‐AP) were bought from ProteinTech (Chicago, USA).

Techniques: Expressing, Immunohistochemical staining

Journal: Redox Report : Communications in Free Radical Research

Article Title: Neutrophil extracellular traps drive osteoporosis via NCF2-dependent signaling: integrated transcriptomics with mechanistic validation

doi: 10.1080/13510002.2025.2534745

Figure Lengend Snippet: Integrative transcriptomic and experimental validation of NET formation in osteoporosis. Schematic overview of the study design. A rat model of postmenopausal osteoporosis (OVX) was established and subjected to transcriptomic analysis (RNA-seq) to identify differentially expressed genes (DEGs) enriched in neutrophil extracellular traps (NETs) formation pathways. Mechanistic insights into NETosis were explored, including activation of neutrophils, NOX complex assembly, and release of NETs containing NE, MPO, and CitH3. For in vitro validation, primary osteoblasts isolated from 7-day-old rats were treated with NET-conditioned medium derived from PMA-induced bone marrow neutrophils or HL-60–derived neutrophil-like cells. siRNA-mediated knockdown of NCF2 was performed to assess its role in NETs formation and subsequent osteogenic inhibition.

Article Snippet: The sections were incubated in a blocking solution to reduce non-specific binding, followed by overnight incubation at 4°C with primary antibodies: VDAC1 (Novus Biologicals, NBP3-15871), PLCG2 (Novus Biologicals, NBP1-86030), and NCF2 (Novus Biologicals, NBP3-16257).

Techniques: Biomarker Discovery, RNA Sequencing, Activation Assay, In Vitro, Isolation, Derivative Assay, Knockdown, Inhibition

Journal: Redox Report : Communications in Free Radical Research

Article Title: Neutrophil extracellular traps drive osteoporosis via NCF2-dependent signaling: integrated transcriptomics with mechanistic validation

doi: 10.1080/13510002.2025.2534745

Figure Lengend Snippet: SMR discovery and experimental validation of NETs-related biomarkers in osteoporosis. A, Forest plot summarizing SMR results for VDAC1, PLCG2 and NCF2 (β, SE, P, HEIDI P, and OR ± 95% CI). B, Representative IHC images of distal femur (brown = positive DAB; hematoxylin counter-stain) showing increased staining for the three proteins in OVX rats versus Sham; scale bar = 50 µm. C, Western blots of neutrophil lysates (Control vs PMA-induced NETs) probed for NCF2 (65 kDa), VDAC1 (31 kDa) and PLCG2 (148 kDa); GAPDH (36 kDa) serves as loading control. D–F, Quantification of IHC-positive area (% of marrow) for each protein (mean ± SD). G–I, Densitometric ratios of target proteins to GAPDH in western blots (mean ± SD). P < 0.05*, < 0.01 **, < 0.001 ***, < 0.0001 **** (two-tailed t-test).

Article Snippet: The sections were incubated in a blocking solution to reduce non-specific binding, followed by overnight incubation at 4°C with primary antibodies: VDAC1 (Novus Biologicals, NBP3-15871), PLCG2 (Novus Biologicals, NBP1-86030), and NCF2 (Novus Biologicals, NBP3-16257).

Techniques: Biomarker Discovery, Staining, Western Blot, Control, Two Tailed Test

Journal: Redox Report : Communications in Free Radical Research

Article Title: Neutrophil extracellular traps drive osteoporosis via NCF2-dependent signaling: integrated transcriptomics with mechanistic validation

doi: 10.1080/13510002.2025.2534745

Figure Lengend Snippet: Knockdown of NCF2 decreases NETS formation and reverses NETs-mediated suppression of osteoblast differentiation. (A) Representative Western blot confirming NCF2 knockdown efficiency in HL-60 neutrophil-like cells. (B, C) Quantification of NCF2 protein and mRNA expression levels (**** P < 0.0001). (D) Triple immunofluorescence staining (DAPI, NE, MPO, CitH3) illustrating reduced NETS formation upon NCF2 knockdown; scale bar = 50 µm. (E–G) Quantitative analysis of NE-, MPO-, and CitH3-positive areas (**** P < 0.0001). (H) Extracellular DNA concentrations in supernatants measured by UV spectrophotometry (**** P < 0.0001). (I, J) ALP staining at day 7 (scale bar = 200 µm) and Alizarin red S staining of mineralisation at day 21 (scale bar = 200 µm) after exposure to indicated conditioned media. (K–M) Western blot and quantitative densitometric analysis of ALP and OCN protein expression levels normalised to GAPDH (*** P < 0.001, **** P < 0.0001). Data represent mean ± SD of three independent experiments; statistical analysis was performed by one-way ANOVA with Tukey’s multiple comparisons test.

Article Snippet: The sections were incubated in a blocking solution to reduce non-specific binding, followed by overnight incubation at 4°C with primary antibodies: VDAC1 (Novus Biologicals, NBP3-15871), PLCG2 (Novus Biologicals, NBP1-86030), and NCF2 (Novus Biologicals, NBP3-16257).

Techniques: Knockdown, Western Blot, Expressing, Immunofluorescence, Staining, Spectrophotometry

Journal: Redox Report : Communications in Free Radical Research

Article Title: Neutrophil extracellular traps drive osteoporosis via NCF2-dependent signaling: integrated transcriptomics with mechanistic validation

doi: 10.1080/13510002.2025.2534745

Figure Lengend Snippet: Docking results of available proteins small molecules. (A) NCF2 docking Diltiazem. (B) VDAC1 docking Diltiazem. (C) PLCG2 docking Exemestane. (D) NCF2 docking Cimetidine. (E) VDAC1 docking Cimetidine. (F) NCF2 docking Desipramine. (G) NCF2 docking Profenamine. (H) VDAC1 docking Emodin. (I) VDAC1 docking Tolazoline. (J) VDAC1 docking Zuclopenthixol. (K) VDAC1 docking Ethynyl estradiol. (L) PLCG2 docking Simvastatin.

Article Snippet: The sections were incubated in a blocking solution to reduce non-specific binding, followed by overnight incubation at 4°C with primary antibodies: VDAC1 (Novus Biologicals, NBP3-15871), PLCG2 (Novus Biologicals, NBP1-86030), and NCF2 (Novus Biologicals, NBP3-16257).

Techniques: