Journal: Cancer Science

Article Title: System biology analysis reveals the role of voltage‐dependent anion channel in mitochondrial dysfunction during non‐alcoholic fatty liver disease progression into hepatocellular carcinoma

doi: 10.1111/cas.14651

Figure Lengend Snippet: VDAC1 is dysregulated in NAFLD‐driven HCC and associated with NAFLD. A, Volcano plot graph showing the differential expressed proteins in the quantitative analysis. The −log 10 ( P ‐value) was plotted against the log2 (ratio Tumor/Normal). The upregulated proteins in HCC tissues were marked with red dots, and the downregulated proteins in HCC tissues with green dots. B, Pathways analysis of significantly altered proteins. The canonical pathways associated with the differential proteins were tested alongside the P ‐values calculated using right‐tailed Fisher exact test. The top 10 pathways were listed. C, Transcriptome‐wide association study suggested that VDAC1 was associated with NAFLD. The correlation between differentially expressed protein and the phenotype of body fat mass percentage (Record ID 12 919) was shown. The red plots indicated the positive correlation, the green plots indicated the negative correlation, and the dark plots indicated no statistical significance. A significant correlation with the VDAC1 ( R = 0.469, P = .008) was shown in the red triangle. D, Overexpression of VDAC1 in tumor tissues was validated with western blot. C represented the HCC tissues, and N represented the adjacent normal tissues. RPLP0 is used as internal reference. E, Heat map showed that VDAC1 was associated with a serial of fatty liver disease phenotypes in both gene and protein expression level. The VDAC1 transcript had significant correlations with the body fat mass ( R = 0.472, P = .008), body weight ( R = 0.361, P = .049), liver mass ( R = 0.358, P = .021), white adipose mass ( R = 0.391, P = .035), cholesterol in plasma ( R = 0.525, P = .0003), Glucose AUC in OGTT ( R = 0.331, P = .034) and fasting glucose level ( R = 0.298, P = .058). Red represents positive correlations, and blue represents negative correlations. Transcript and peptide 1‐20 represent the gene and peptides expression level, respectively. A, B, D, obtained from the proteomic analysis of human samples; C, E were obtained from the transcriptomic and proteomic analysis of BXD family samples

Article Snippet: The SiRNA (vdac1‐homo‐815, 19601) and plasmid (HG17755‐NF, Sino Biological) were transfected for knockdown and overexpression of VDAC1, respectively; 0.25 mmol/L DIDS (32029, MCE) was used to inhibit VDAC1.

Techniques: Over Expression, Western Blot, Expressing, Clinical Proteomics

Journal: Cancer Science

Article Title: System biology analysis reveals the role of voltage‐dependent anion channel in mitochondrial dysfunction during non‐alcoholic fatty liver disease progression into hepatocellular carcinoma

doi: 10.1111/cas.14651

Figure Lengend Snippet: Identification of VDAC1 expression correlated with HCC in a human clinical cohort. A, Vdac1 upregulated in HCC samples compared with the normal samples was validated in the Cancer Genome Atlas Liver Hepatocellular Carcinoma (TCGA‐LIHC) ( n = 371), and Normal ( n = 50) cohorts. B, Vdac1 is upregulated with HCC grade increase and was identified in TCGA‐LIHC Grade 1 ( n = 54), Grade 2 ( n = 173), Grade 3 ( n = 118), Grade 4 ( n = 12), and Normal ( n = 50) cohorts. C, High‐level VDAC1 expression correlates with poor survival outcome in HCC and LIHC (High, n = 91; Low, n = 91) cohorts based on survival analysis

Article Snippet: The SiRNA (vdac1‐homo‐815, 19601) and plasmid (HG17755‐NF, Sino Biological) were transfected for knockdown and overexpression of VDAC1, respectively; 0.25 mmol/L DIDS (32029, MCE) was used to inhibit VDAC1.

Techniques: Expressing

Journal: Cancer Science

Article Title: System biology analysis reveals the role of voltage‐dependent anion channel in mitochondrial dysfunction during non‐alcoholic fatty liver disease progression into hepatocellular carcinoma

doi: 10.1111/cas.14651

Figure Lengend Snippet: System genetics analysis of Vdac1 in BXD mice strains. A, Variable expression levels of Vdac1 in the liver tissue. Expression data for 39 BXD strains and their parental strains (B6 and D2). The x‐axis denotes the strain, while the y‐axis denotes mean expression given as log 2 . Each bar shows mean expression values ± standard error of the mean (SEM). B, Interval mapping of the Vdac1 gene in the liver. The upper x‐axis shows the chromosome, the lower x‐axis shows the location in megabases, the y‐axis provides the likelihood ratio statistics score (LRS). Blue lines indicate the LRS values at a given position, with a significant LRS at 18.35 and suggestive LRS at 11.14. Interval mapping identified a significant cis ‐eQTL on chromosome 11 at 52.01 Mb and 4 suggestive trans ‐eQTLs on chromosomes 2, 5, 14, and X

Article Snippet: The SiRNA (vdac1‐homo‐815, 19601) and plasmid (HG17755‐NF, Sino Biological) were transfected for knockdown and overexpression of VDAC1, respectively; 0.25 mmol/L DIDS (32029, MCE) was used to inhibit VDAC1.

Techniques: Expressing

Journal: Cancer Science

Article Title: System biology analysis reveals the role of voltage‐dependent anion channel in mitochondrial dysfunction during non‐alcoholic fatty liver disease progression into hepatocellular carcinoma

doi: 10.1111/cas.14651

Figure Lengend Snippet: List of 4 suggested trans ‐eQTLs for Vdac1

Article Snippet: The SiRNA (vdac1‐homo‐815, 19601) and plasmid (HG17755‐NF, Sino Biological) were transfected for knockdown and overexpression of VDAC1, respectively; 0.25 mmol/L DIDS (32029, MCE) was used to inhibit VDAC1.

Techniques: Marker

Journal: Cancer Science

Article Title: System biology analysis reveals the role of voltage‐dependent anion channel in mitochondrial dysfunction during non‐alcoholic fatty liver disease progression into hepatocellular carcinoma

doi: 10.1111/cas.14651

Figure Lengend Snippet: VDAC1 is associated with mitochondria dysfunction. A, Vdac1 co‐expression network is involved in mitochondrial dysfunction based on the mice liver transcriptome data. The Vdac1 co‐expression genes were analyzed by IPA. The top canonical pathways associated with the network genes were tested alongside the P ‐values calculated using a right‐tailed Fisher exact test. Mitochondrial dysfunction was listed as the top pathway. B, Vdac1 shows correlation with 20 mitochondrial genes. The red plots indicate the significant positive correlation, and the dark plots indicate no statistical significance. C, Heatmap showing the correlation between VDAC1 and different species of CL based on the BXD family database. VDAC1 transcript had a negative relationship with the predominant CL species, CL (LLLL) and its precursor MLCL (LLL). VDAC1 had a positive relation with the nascent CL; and the VDAC1 correlated peptides showed the same trend. Red represents positive correlations, and blue represents negative correlations. D, Correlation between Vdac1 and cardiolipin synthetase gene Ptpmt1 and Tafazzin in a BXD mice cohort. Vdac1 has a positive correlation with Ptpmt1 ( R = 0.452, P = .003); and negative correlation with Tafazzin ( R =−0.334, P = .033) among 41 BXD strains. The blue line represents the correlation with Ptpmt1 , and green with Tafazzin

Article Snippet: The SiRNA (vdac1‐homo‐815, 19601) and plasmid (HG17755‐NF, Sino Biological) were transfected for knockdown and overexpression of VDAC1, respectively; 0.25 mmol/L DIDS (32029, MCE) was used to inhibit VDAC1.

Techniques: Expressing

Journal: Cancer Science

Article Title: System biology analysis reveals the role of voltage‐dependent anion channel in mitochondrial dysfunction during non‐alcoholic fatty liver disease progression into hepatocellular carcinoma

doi: 10.1111/cas.14651

Figure Lengend Snippet: Mitochondrial function analysis with or without VDAC1 expression in Hep3B and HepG2. A, Western blot analysis of VDAC1 level in HepG2 cells after transfection of VDAC1 siRNA and VDAC1 expression plasmid. Compared with the negative control, the expression level of VDAC1 reduced in the knockdown group, and increased in overexpression group. RPLPO was the internal reference protein. B, Determination of cardiolipin in Hep3B and HepG2 cells. Cardiolipin was measured using the fluorometric probe assay kit (ab241036). The left panel shows the cardiolipin content in Hep3B. Compared with the NC group, the cardiolipin contents both in knockdown group and inhibitor group were significantly higher ( P < .01), and the cardiolipin content in overexpression group was lower ( P < .05). The right panel showed the cardiolipin content in HepG2. Compared with the NC group, the cardiolipin contents both in knockdown group and inhibitor group were significantly higher ( P < .01), and the cardiolipin content in overexpression group was lower ( P < .05). C, Detection of cardiolipin levels in HepG2 and Hep3B by NAO staining. Compared with negative control, the fluorescence intensity increased in the knockdown group and inhibitor group, and decreased in the overexpression group both in Hep3B and HepG2. The fluorescence intensity of the 2 cell lines was observed under a microscope at ×10 and ×40 magnification, respectively. D, Detection of mitochondrial respiratory function in Hep3B and HepG2 cells. High‐resolution respirometry performed in an Oroboros Oxygraph‐2k. The different color curves showed the mitochondrial oxygen consumption of the different groups. In Hep3B cells, the OCR of knockdown group and inhibitor group were higher than the NC group, and the OCR of overexpression group and inhibitor group was lower than the NC group. Also, the same trend was shown in HepG2 cells. E, Relative VDAC1 expression was detected by IHC between NAFLD‐driven HCC and adjacent non‐tumor liver tissues (×40). N represented the VDAC1 in adjacent non‐tumor liver tissues. T represented the VDAC1 in NAFLD‐driven HCC tissues. (NC represented the negative control group; VDAC1 siRNA represented VDAC1 knockdown group; VDAC1 represented the overexpression group and DIDS represented the inhibitor group)

Article Snippet: The SiRNA (vdac1‐homo‐815, 19601) and plasmid (HG17755‐NF, Sino Biological) were transfected for knockdown and overexpression of VDAC1, respectively; 0.25 mmol/L DIDS (32029, MCE) was used to inhibit VDAC1.

Techniques: Expressing, Western Blot, Transfection, Plasmid Preparation, Negative Control, Knockdown, Over Expression, Staining, Fluorescence, Microscopy

Journal: Cancer Science

Article Title: System biology analysis reveals the role of voltage‐dependent anion channel in mitochondrial dysfunction during non‐alcoholic fatty liver disease progression into hepatocellular carcinoma

doi: 10.1111/cas.14651

Figure Lengend Snippet: VDAC1‐centralized correlation network. A, Correlation network of VDAC1‐centralized transcript‐lipids phenotypes based on the BXD mice family database. The network was constructed including VDAC1 expression, NAFLD phenotypes and representative CL species. Red lines show positive correlations, and blue lines show negative correlations. The network illustrates the VDAC1 associated with NAFLD phenotypes through correlating with CL profiling. B, Schematic for VDAC1 involved in NAFLD‐related HCC. VADC1 is upregulated in NAFLD, and dysregulation of VDAC1 causes a CL acyl chain composition shift, which led the NAFLD‐driven HCC tumorigenesis through mitochondrial dysfunction

Article Snippet: The SiRNA (vdac1‐homo‐815, 19601) and plasmid (HG17755‐NF, Sino Biological) were transfected for knockdown and overexpression of VDAC1, respectively; 0.25 mmol/L DIDS (32029, MCE) was used to inhibit VDAC1.

Techniques: Construct, Expressing

Journal: Cell Reports Medicine

Article Title: Urolithin A improves muscle strength, exercise performance, and biomarkers of mitochondrial health in a randomized trial in middle-aged adults

doi: 10.1016/j.xcrm.2022.100633

Figure Lengend Snippet: Urolithin A confers a proteomic signature of improved mitochondrial metabolism and mitophagy in human skeletal muscle (A) Venn diagram summarizing pathway enrichment analysis results from proteomics data in vastus lateralis skeletal muscle. Data represent upregulated pathways with an adjusted p value <0.1 in subjects treated with placebo or with UA at 500 and 1,000 mg for 4 months compared with baseline. (B and C) Dot plots showing top enriched pathways (WikiPathways 2019 Human), ranked by protein ratio, in the UA 500- (B) and UA 1,000 mg (C) groups from (A). Dot color and size indicate adjusted p value and protein count, respectively. Significant pathways for the placebo group were filtered out to identify treatment-specific pathways. (D) Western blot analysis of protein lysates from vastus lateralis skeletal-muscle biopsies in subjects treated as above. For each subject, both baseline (Pre) and end-of-the-treatment (Post) samples were run, and membranes were probed for phospho-Parkin, total Parkin, and the mitochondrial proteins ATP5A (belonging to the OXPHOS complex V), UQCRC2 (complex IV), SDHB (complex II), and NDUFB8 (complex I). Tubulin and VDAC were included as markers of total and mitochondrial protein abundance, respectively. Dashed line separates samples from individual subjects. (n = 6 Pre and Post, biologically independent samples). (E) Quantification of phospho-Parkin over Tubulin protein intensity from western blots (WBs) in (D) (n = 6). Two-sided, paired t-test. (F) Quantification of NDUFB8 (left) and SDHB (right) protein intensity, normalized over VDAC from WBs in (D) (n = 6). ∗p < 0.05; ∗∗p < 0.01; two-sided, paired t test.

Article Snippet: The following primary antibodies were incubated overnight diluted in blocking buffer: UBE2N (SantaCruz, sc-376470, 1:3000), Tubulin (Proteintech, 10004185, 1:3000), phospho-Parkin S65 (Cell Signaling, #36866, 1:1000), Parkin (Cell Signaling, #4211, 1:1000), OXPHOS Antibody Cocktail (Abcam, ab110413, 1:2000), VDAC (Proteintech, 10866-1-AP, 1:1000).

Techniques: Western Blot, Quantitative Proteomics

Journal: Cell Reports Medicine

Article Title: Urolithin A improves muscle strength, exercise performance, and biomarkers of mitochondrial health in a randomized trial in middle-aged adults

doi: 10.1016/j.xcrm.2022.100633

Figure Lengend Snippet:

Article Snippet: The following primary antibodies were incubated overnight diluted in blocking buffer: UBE2N (SantaCruz, sc-376470, 1:3000), Tubulin (Proteintech, 10004185, 1:3000), phospho-Parkin S65 (Cell Signaling, #36866, 1:1000), Parkin (Cell Signaling, #4211, 1:1000), OXPHOS Antibody Cocktail (Abcam, ab110413, 1:2000), VDAC (Proteintech, 10866-1-AP, 1:1000).

Techniques: Recombinant, RNA Extraction, Software, Protease Inhibitor

Journal: Aging Cell

Article Title: The ultrastructural and proteomic analysis of mitochondria‐associated endoplasmic reticulum membrane in the midbrain of a Parkinson's disease mouse model

doi: 10.1111/acel.14436

Figure Lengend Snippet: Initial proteomic parsing of midbrain MAM fractions in controls and MPTP‐treated mice. (a) Subcellular fractions extracted from midbrain tissues were validated by specific organelle protein markers. Calnexin, Calreticulin, and Sigma‐1R were considered as consensus proteins for ER and MAM. VDAC1 and Cox IV were applied for mitochondria and MAM markers, and GAPDH was to prove cytosolic fractions. (b) Venn graph of identified proteins in each group ( n = 5 per group) and consensus MAM proteins. (c, d) Sample distribution plots by PCA (c) and PLS‐DA analysis (d) in MAM proteomics between controls and MPTP‐treated mice.

Article Snippet: Primary antibodies used in this study included that anti‐tyrosine hydroxylase (TH) rabbit mAb (1:2000, Cat. 58844, Cell Signaling Technology), anti‐glial fibrillary acidic portein (GFAP) rabbit mAb (1:2000, Cat. 80788, Cell Signaling Technology), anti‐glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) rabbit mAb (1:2000, Cat. 5174, Cell Signaling Technology), anti‐cytochrome‐c‐oxidase subunit 4 (Cox IV) rabbit Ab (1:2000, Cat. 5844, Cell Signaling Technology), anti‐sigma‐1 receptor (Sig‐1R) rabbit mAb (1:2000, Cat. 61994, Cell Signaling Technology), anti‐phosphatase and tensin homolog (PTEN) rabbit mAb (1:2000, Cat. 9188, Cell Signaling Technology), anti‐calreticulin rabbit mAb (1:2000, Cat. 92516, Abcam), anti‐calnexin (CNX) rabbit pAb (1:2000, Cat. ADI‐SPA‐860, Enzo Life Sciences), anti‐voltage dependent anion channel 1 (VDAC1) rabbit pAb (1:2000, Cat. 55259‐1‐AP, Proteintech), anti‐leucine‐rich repeat kinase 2 (LRRK2) rabbit mAb (1:1000, Cat. R380823, Zen‐bioscience), anti‐excitatory amino acid transporter 2 (EAAT2) rabbit mAb (1:1000, Cat. R381853, Zen‐bioscience), anti‐tropomyosin‐1 (TPM1) rabbit mAb (1:1000, Cat. R383145, Zen‐bioscience), and anti‐coatomer protein complex subunit zeta 1 (COPZ1) rabbit pAb (1:1000, Cat. 824631, Zen‐bioscience).

Techniques:

Journal: BMC Molecular and Cell Biology

Article Title: Alterations in the chondrocyte surfaceome in response to pro-inflammatory cytokines

doi: 10.1186/s12860-020-00288-9

Figure Lengend Snippet: Western blotting and densitometric analysis of LRP-1, thrombospondin, VDAC1, VDAC2 and annexin A1 in cell lysates of chondrocytes under inflammatory (IL-1β + TNF-α, both 10 ng/mL) versus control conditions. a Non-significant decrease in LRP-1 expression upon cytokine exposure. b Increase in thrombospondin expression upon cytokine exposure. c Similar VDAC1 expression. d Similar VDAC2 expression. e Annexin A1 expression showed a trend to decrease upon cytokine exposure. f Beta-actin was measured on each western blot separately as a loading control to correct for the exact amount of protein per lane. Measurements from three horses (three biological replicates) were combined to provide final values for each group (mean ± SD)

Article Snippet: VDAC1 (C-terminal) , Aviva Systems Biology , ARP35122 (polyclonal) , 1:250 , IRDye 680RD Goat , LI-COR Biosciences , 926–68,071 , 1:5000.

Techniques: Western Blot, Control, Expressing

Journal: BMC Molecular and Cell Biology

Article Title: Alterations in the chondrocyte surfaceome in response to pro-inflammatory cytokines

doi: 10.1186/s12860-020-00288-9

Figure Lengend Snippet: Western blotting and densitometric analysis of VDAC1 and VDAC2 monomers and dimers in cell lysates of chondrocytes upon cytokine exposure (IL-1β + TNF-α, both 10 ng/mL) versus control conditions. a VDAC1 monomer expression was significantly decreased upon cytokine exposure. VDAC1 dimer expression was unchanged. b Similar VDAC2 monomer and dimer expression. Measurements from three horses (three biological replicates) were combined to provide final values for each group (mean ± SD)

Article Snippet: VDAC1 (C-terminal) , Aviva Systems Biology , ARP35122 (polyclonal) , 1:250 , IRDye 680RD Goat , LI-COR Biosciences , 926–68,071 , 1:5000.

Techniques: Western Blot, Control, Expressing

Journal: BMC Molecular and Cell Biology

Article Title: Alterations in the chondrocyte surfaceome in response to pro-inflammatory cytokines

doi: 10.1186/s12860-020-00288-9

Figure Lengend Snippet: Detailed specification of primary and secondary antibodies employed for western blotting

Article Snippet: VDAC1 (C-terminal) , Aviva Systems Biology , ARP35122 (polyclonal) , 1:250 , IRDye 680RD Goat , LI-COR Biosciences , 926–68,071 , 1:5000.

Techniques: Western Blot

Journal: bioRxiv

Article Title: SIRT3 deficiency decreases oxidative-metabolism capacity but increases lifespan under caloric restriction

doi: 10.1101/2022.05.09.491205

Figure Lengend Snippet: A) Immunoblot of pan-acetylation, normalized to VDAC, for liver mitochondrial enrichment from 25-month-old treatment groups. Data were analyzed by two-way ANOVA followed by multiple comparisons test. p value reported for each comparison is corrected by Tukey’s test. Results are plotted as mean ± SEM. *: p≤0.05. B) Percentage of significantly changed acetyl-lysine residues that show increased stoichiometry due to Sirt3 -/- , calculated by (the number of acetyl-lysine sites showing increased stoichiometry) / (the number of significantly changed acetyl-lysine sites, p<0.05) x100%. C) Heat map of significantly changed lysine sites (p<0.1) that are response to loss of SIRT3. Plotted sites are significantly changed (p<0.1) in either Sirt3 -/- CD vs. WTCD or Sirt3 -/- CR vs. WTCR comparison. Values are colored based on relative acetylation stoichiometry, normalized to the median value of each sites in all four groups, scaling ranging from -0.8 to 0.8 (x100%). D) Functional cluster analysis of KEGG pathways (DAVID 6.8). Significantly enriched (-log10(p value) >1.5) pathways are indicated, with Sirt3 -/- CD vs. WTCD in orange and Sirt3 -/- CR vs. WTCR in blue. E) Acetylation sites in FAO and BACC metabolism, TCA cycle, and ETC that displayed larger than 5% stoichiometry (p<0.1) for Sirt3 -/- CD vs. WTCD (orange colored) and Sirt3 -/- CR vs. WTCR comparison (blue colored).

Article Snippet: Western blot primary antibodies used include: SIRT3 (#5490, CST, 1:1000), Acetylated-Lysine (#9681, CST, 1:1000), VDAC (75-204, NeuroMab, 1:1250).

Techniques: Western Blot, Comparison, Functional Assay

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: 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: