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: Journal of Biological Chemistry

Article Title: Developmental Control of Apoptosis by the Immunophilin Aryl Hydrocarbon Receptor-interacting Protein (AIP) Involves Mitochondrial Import of the Survivin Protein

doi: 10.1074/jbc.m110.210120

Figure Lengend Snippet: FIGURE 6. Tom20 regulation of mitochondrial import of survivin. A, ali- quots of GST, GST-Tom20, or GST-Tom70 recombinant proteins were mixed with Raji cell extracts, and bead-bound material was analyzed by Western blotting. Bottom panel, Coomassie Blue staining of recombinant GST fusion proteins. B, Raji mitochondrial extract was immunoprecipitated with an anti- body to survivin or IgG, and proteins in pellets or supernatants (unbound) were analyzed by Western blotting. C, recombinant wild-type survivin or the survivin 1–141 mutant and GST-Tom20 beads were mixed with increasing concentration of recombinant AIP, and bead-bound proteins were analyzed by Western blotting (top panel). Bottom panel, GST or GST-Tom20 was incu- bated with recombinant AIP in the presence of increasing concentrations of survivin (up to 0.4 M), and bound proteins were analyzed by Western blot- ting. D, independently established clones (#) of HeLa cells stably transfected with control (Ctrl) or Tom70- or Tom20-directed shRNA were analyzed by Western blotting. E, HeLa cells were transfected with control, non-targeting (Ctrl) or Tom20-directed siRNA and analyzed by Western blotting. TCE, total cell extracts. F, clones of HeLa cells with stable knockdown of Tom70 or Tom20 were treated with 0.5 M staurosporine for 24 h and analyzed for cell viability by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide. DataaremeanS.E.ofreplicates(n3).***,p0.0001.Twodifferentclones per conditions were tested with identical results in two independent experiments.

Article Snippet: Antibodies against AIP (Novus Biologicals), survivin (Novus Biologicals), Tom20 (Santa Cruz Biotechnology), Tom70 (Novus Biologicals), COX-IV (Clontech), Smac (Pro-Sci), -actin (Sigma), and Hsp90 (BD Biosciences) were used.

Techniques: Recombinant, Western Blot, Staining, Immunoprecipitation, Mutagenesis, Concentration Assay, Clone Assay, Stable Transfection, Transfection, Control, shRNA, Knockdown

Journal: Frontiers in Neurology

Article Title: Effect of electroacupuncture on hippocampal protein lactylation in a rat model of vascular dementia

doi: 10.3389/fneur.2025.1629474

Figure Lengend Snippet: KEGG pathway clustering analysis of different comparison groups, the PPI network of differentially lactylated proteins in the pathways of interest, and verification of Vdac3 protein lactylation modifications. (A) Heatmap of lactylation sites for 12 key proteins. Each row represents a differentially modification site; each column represents a sample. Red indicates a high-level modification, blue indicates a low-level modification, and grey indicates sites that cannot be quantified in the corresponding samples. (B) The horizontal axis depicts different comparison groups, whereas the vertical axis indicates the enriched KEGG pathways. The colored blocks represent the functional descriptions of differentially expressed modified proteins enriched in each comparison group. Blue signifies high enrichment significance, whereas blue-white signifies low enrichment significance. * p < 0.05, ** p < 0.01, and *** p < 0.001. (C) PPI network of differentially lactylated proteins involved in the 4-VO group and sham group in the five pathways of interest. (D) PPI network in the 4-VO group and 4-VO + EA group. The color of the outer circle indicates the corresponding KEGG pathway of the protein, whereas the shape within the circle denotes the upregulation or downregulation of the modification sites contained within the protein. A diamond shape signifies proteins with upregulated modification sites, a downward arrow indicates proteins with downregulated modification sites, and a circle signifies proteins with upregulated and downregulated modification sites. The red box indicates the Vdac3 protein. (E) The IP experiment verified the lactylation modification level of Vdac3. (F) Quantification was performed using ImageJ software, and the values are expressed as the mean with the corresponding standard error from three replicate experiments ( n = 3, compared with the sham group, ## p < 0.01; compared with the 4-VO group, ** p < 0.01). 4-VO, four-vessel occlusion; EA, electroacupuncture; KEGG, Kyoto Encyclopedia of Genes and Genomes; PPI, protein–protein interaction; IP, immunoprecipitation; Vdac3, mitochondrial voltage-dependent anion channel protein3.

Article Snippet: One milliliter of total protein extract was pre-cleared with 40 μL of protein A/G agarose beads by rotation at 4 °C for 1 h. Subsequently, the supernatant was incubated with 5 μL of Vdac3 antibody (A04802-2, BosterBio, China) overnight at 4 °C with rotation.

Techniques: Comparison, Modification, Functional Assay, Software, Immunoprecipitation

Journal: Frontiers in Neurology

Article Title: Effect of electroacupuncture on hippocampal protein lactylation in a rat model of vascular dementia

doi: 10.3389/fneur.2025.1629474

Figure Lengend Snippet: Analysis of lactylation site motifs in different groups and representative mass spectra of key proteins. (A) A heatmap of amino acids adjacent to lysine lactylation sites, with a green/red scale (−5 to 5) indicating the frequency of amino acid detection; red indicates high frequency; green indicates low frequency. The height ratio of amino acid abbreviation letters at specific positions reflects motif characteristics, with a % difference greater than 0 indicating a higher frequency of that amino acid at that position compared to the background, and less than 0 indicating a lower frequency. (B) Four conserved motif sites of lysine. (C) Sequence motif map of lactylation modification, 10 amino acids upstream and downstream of the modification site were selected, and the vertical axis indicates the difference between the proportion of the various amino acids at the same site in the experimental data set and the reference background, that is, percentage difference (%difference). The height ratio of amino acid abbreviation letters at specific positions represents the motif characteristics, and a %difference greater than 0 indicates that the frequency of the amino acid at this position is higher than the background. Values lower than 0 indicate that the frequency of the amino acid at this position is lower than the background. (D–F) The lactylation site of Vdac3 was identified by mass spectrometry. PPI, protein–protein interaction; 4-VO, four-vessel occlusion; EA, electroacupuncture.

Article Snippet: One milliliter of total protein extract was pre-cleared with 40 μL of protein A/G agarose beads by rotation at 4 °C for 1 h. Subsequently, the supernatant was incubated with 5 μL of Vdac3 antibody (A04802-2, BosterBio, China) overnight at 4 °C with rotation.

Techniques: Sequencing, Modification, Mass Spectrometry

Journal: Molecular nutrition & food research

Article Title: Lack of β, β-carotene -9’, 10’-oxygenase 2 leads to hepatic mitochondrial dysfunction and cellular oxidative stress in mice

doi: 10.1002/mnfr.201600576

Figure Lengend Snippet: Functional categorization of proteins differentially expressed in hepatic mitochondria of BCO2 knockout (KO) vs. 129S6 wild type (WT) mice identified by spectrum counting. Values are means, n=3 mice/group with 4 technical replicates/mouse.

Article Snippet: Antibodies against fatty acid synthase (FASN) (catalog # 10624-2-Ap), ATP synthase α subunit 1 (ATP5A1) (catalog # 14676-1-AP), citrate synthase (catalog # 16131-1-AP), hydroxyacyl-coenzyme A dehydrogenase (HADH) (catalog # 19828-1-AP), succinate dehydrogenase α (SDHA) (catalog # , 14865-1-AP), and nicotinamide nucleotide transhydrogenase (NNT) (catalog # 13442-2-AP) were purchased from ProteinTech Group (Chicago, IL, USA); Antibodies against lysosome-associated membrane protein 1 (LAMP1) (catalog # sc-20011), catalase (catalog # sc-50508), NNT (catalog # sc-163154), glutathione reductase (GSR) (catalog # sc-133245) were purchased from Santa Cruz Biotech (Dallas, TX, USA); peroxisome proliferator-activated receptor α (PPARα) antibody (catalog # 101710) was purchased from Cayman (Ann Arbor, MI, USA); Antibodies against β-actin (catalog # 4967), phosphor-Thr172-AMP-activated protein kinase α (pT172-AMPKα) (catalog # 2535), and AMPKα (catalog # 2603) were purchased from Cell Signaling Technology (Danvers, MA, USA); voltage-dependent anion-selective channel protein 1 (VDAC1) antibody (catalog # PA1780) was purchased from Boster Biosciences (Pleasanton, CA, USA).

Techniques: Functional Assay, Knock-Out, Membrane