Journal: bioRxiv

Article Title: Cell Type–Dependent Uptake of Extracellular Vesicles Independent of Cellular Origin

doi: 10.64898/2026.05.19.726167

Figure Lengend Snippet: Uptake of SKBR-3-derived EVs by human epithelial cell lines. (A) Quantification of EV uptake following 2 h incubation of SKBR-3-derived mNG-labeled EVs with SKBR-3, HEK293T, Caco-2, PDAK, HepG2, and Huh7 cells at EV concentrations of 1 × 10 8 , 1 × 10 9 , and 1 × 10 10 particles. (B) Representative flow cytometry plots corresponding to panel A. (C) Quantification of EV uptake following 4 h incubation at the indicated concentrations. (D) Representative flow cytometry plots corresponding to panel C. Data are presented as mean ± SD (n = 3 independent experiments). Statistical analysis was performed using two-way ANOVA. Statistical significance is indicated as follows: **p < 0.01, ****p < 0.0001.

Article Snippet: Human embryonic kidney cells (HEK293T), SKBR-3 human breast cancer cells, pancreatic ductal adenocarcinoma (PDAK) cells, HepG2 human hepatic cancer cells, Huh7 human hepatocellular carcinoma cells, Caco-2 human colorectal adenocarcinoma cells, and the murine melanoma cell lines B16F10 and Yummer (all obtained from ATCC) were cultured in DMEM supplemented with 10% FBS and 1% antibiotic–antimycotic solution.

Techniques: Derivative Assay, Incubation, Labeling, Flow Cytometry

Journal: Genome biology

Article Title: Systematic interrogation of functional genes underlying cholesterol and lipid homeostasis.

doi: 10.1186/s13059-025-03531-8

Figure Lengend Snippet: Fig. 1 Synthetic lethal CRISPR screening to identify potential cholesterol regulators in HepG2 cells. A Immunoblot analysis of HMGCR in HepG2 cells undergoing CRISPR-mediated gene knockout with two independent sgRNAs (numbered as _1 and _2). Vector without specific sgRNA insert serves as a control. GAPDH serves as a loading control. B Immunoblot analysis of LDLR in HepG2 cells that have undergone CRISPR-mediated gene knockout with two independent sgRNAs. C The cell growth analysis of HepG2 cells after introducing indicated sgRNAs via lentiviral infection for 7 days. Cells were counted with a hemacytometer. Mean ± SD with n = 3. Ordinary one-way ANOVA with Tukey’s test, **p < 0.01, ***p < 0.001. D The relative cell viability was determined by CCK-8 assay for HepG2 cells expressing indicated sgRNAs and treated with indicated doses of lovastatin. Mean ± SD with n = 6. Ordinary one-way ANOVA with Dunnett’s test, *p < 0.05, **p < 0.01, ***p < 0.001. E The workflow of genome-scale synthetic lethal CRISPR screens (Screen 1) to identify negative GIs with HMGCR using its inhibitor lovastatin in HepG2 cells. F The scatter plot showing the β score of each gene and the correlation of both CRISPR screens (vehicle and lovastatin) in HepG2 cells. The genes in blue box are preferential targets as the synthetic lethal or negative GI hits. G The rank-ordered list of each gene in the CRISPR screens according to the strength of synthetic lethality measured by differential β scores between lovastatin and vehicle conditions. The top interesting gene hits are highlighted. H The top selected functional terms enriched among synthetic lethal hits of the CRISPR screens (Screen 1) as determined by the gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis

Article Snippet: Human hepatic cancer cell line HepG2, cervix cancer cell line HeLa, and HEK293FT cells were obtained from the American Type Culture Collection (ATCC).

Techniques: CRISPR, Western Blot, Gene Knockout, Plasmid Preparation, Control, Infection, CCK-8 Assay, Expressing, Functional Assay

Journal: Genome biology

Article Title: Systematic interrogation of functional genes underlying cholesterol and lipid homeostasis.

doi: 10.1186/s13059-025-03531-8

Figure Lengend Snippet: Fig. 3 Identification of potential cholesterol regulators at the transcriptional level. A Venn diagram showing the overlap of up- or down-regulated DEGs of HepG2 cells upon LDLR KO, HMGCR, or double KO (DKO) determined by RNA-seq analysis. B Heatmap showing the DEGs across different samples of HepG2 cells. C Volcano plot showing the DEGs in DKO cells compared to control HepG2 cells with several typical genes highlighted. D The top five enriched functional terms for either up-regulated or down-regulated DEGs in DKO cells compared to control HepG2 cells by GO and KEGG analysis. E Venn diagram showing the overlap of DEGs between sterol deprivation condition (sterol depleted vs. normal) and SREBF2 KO under sterol deprivation (SREBF2 KO vs. vector control) in HepG2 cells. F Heatmap showing the DEGs across indicated samples of HepG2 cells. G Volcano plot showing the DEGs between sterol deprivation and normal conditions of HepG2 cells with several representative genes highlighted. H Volcano plot showing the DEGs in SREBF2 KO cells compared to vector control HepG2 cells in sterol deprivation condition with several representative genes highlighted. I Venn diagram showing the overlap of DEGs for indicated comparison groups. J Heatmap showing the expression change of representative DEGs that were shared between at least three comparison groups of I

Article Snippet: Human hepatic cancer cell line HepG2, cervix cancer cell line HeLa, and HEK293FT cells were obtained from the American Type Culture Collection (ATCC).

Techniques: RNA Sequencing, Control, Functional Assay, Plasmid Preparation, Comparison, Expressing

Journal: Genome biology

Article Title: Systematic interrogation of functional genes underlying cholesterol and lipid homeostasis.

doi: 10.1186/s13059-025-03531-8

Figure Lengend Snippet: Fig. 4 Integrative multi-omics analysis to pinpoint key cholesterol regulators. A Heatmap showing the differentially expressed proteins in LDLR/HMGCR DKO cells compared to Vector control HepG2 cells determined by mass spectrometry-based proteomics profiling. The number of up- or down-regulated proteins is indicated along the heatmap. B Venn diagram showing the overlap of either up-regulated or down-regulated genes in LDLR/HMGCR DKO cells compared to Vector control HepG2 cells between RNA-seq and proteomics analysis. C Volcano plot showing the differentially expressed proteins in LDLR/HMGCR DKO cells compared to control HepG2 cells with several typical proteins highlighted. D The top ten enriched functional terms for up-regulated proteins in LDLR/HMGCR DKO cells compared to control HepG2 cells by GO and KEGG analysis. E The analytic scheme of human genes associated with lipid disorders or cardiovascular diseases. HDL: high-density lipoprotein; LDL: low-density lipoprotein; VLDL: very low-density lipoprotein; TC: total cholesterol; TG: triglycerides; CAD: coronary artery disease. F Venn diagram showing the overlap of potential cholesterol or lipid regulators between different angles including synthetic lethal CRISPR screens (Screen 1–3), DEGs in RNA-seq analysis (DKO vs. vector in HepG2 and HeLa cells, sterol depleted vs normal, and SREBF2 KO vs vector), differentially expressed proteins in proteomics analysis (DKO vs. vector in HepG2 cells), and the compiled list of human genes with variants related to lipid disorders or related diseases. The genes of indicated intersections are highlighted

Article Snippet: Human hepatic cancer cell line HepG2, cervix cancer cell line HeLa, and HEK293FT cells were obtained from the American Type Culture Collection (ATCC).

Techniques: Biomarker Discovery, Plasmid Preparation, Control, Mass Spectrometry, RNA Sequencing, Functional Assay, CRISPR

Journal: Genome biology

Article Title: Systematic interrogation of functional genes underlying cholesterol and lipid homeostasis.

doi: 10.1186/s13059-025-03531-8

Figure Lengend Snippet: Fig. 5 Functions of GGT7 during cholesterol and lipid homeostasis. A RNA expression analysis of indicated genes by RT-qPCR in HepG2 cells upon LDLR KO, HMGCR KO, or LDLR/HMGCR DKO. Mean ± SD with n = 3. Ordinary one-way ANOVA with Dunnett’s test, compared to Vector control, **p < 0.01, ***p < 0.001. B RNA expression analysis of indicated genes by RT-qPCR in HepG2 cells upon SREBF2 KO under sterol deprivation condition. Mean ± SD with n = 3. Unpaired two-sided t test, compared to vector control, **p < 0.01, ***p < 0.001. C Immunoblot analysis of GGT7 in HepG2 cells undergoing CRISPR-mediated gene knockout with two independent sgRNAs. GAPDH serves as a loading control. D Decreased total cholesterol levels of HepG2 cells upon GGT7 KO using two independent sgRNAs. Mean ± SD with n = 3. Unpaired two-sided t test, compared to vector control, *p < 0.05. E Decreased sterol and other lipids upon GGT7 KO in HepG2 cells determined by untargeted metabolomic profiling. n = 6 biological replicates for each group

Article Snippet: Human hepatic cancer cell line HepG2, cervix cancer cell line HeLa, and HEK293FT cells were obtained from the American Type Culture Collection (ATCC).

Techniques: RNA Expression, Quantitative RT-PCR, Plasmid Preparation, Control, Western Blot, CRISPR, Gene Knockout

Journal: Genome biology

Article Title: Systematic interrogation of functional genes underlying cholesterol and lipid homeostasis.

doi: 10.1186/s13059-025-03531-8

Figure Lengend Snippet: Fig. 6 Mechanistic insights of GGT7 in regulating cholesterol metabolism. A RNA expression analysis of indicated genes by RT-qPCR in HepG2 cells upon GGT7 KO. Mean ± SD with n = 3. Unpaired two-sided t test, compared to Vector control, *p < 0.05, **p < 0.01, ***p < 0.001. B RNA expression analysis of indicated genes by RT-qPCR in HepG2 cells upon GGT7 KO under normal or sterol deprivation conditions. Mean ± SD with n = 3. Unpaired two-sided t test, *p < 0.05, **p < 0.01, ***p < 0.001. C Volcano plot showing the DEGs in GGT7 KO cells compared to vector control HepG2 cells with several typical genes highlighted. The number of up- or down-regulated DEGs is indicated. D The top selected functional terms enriched for down-regulated DEGs in GGT7 KO cells compared to control HepG2 cells by GO and KEGG analysis. E Coomassie blue staining of SDS-PAGE gel with FLAG bead immunoprecipitated materials from vector control- or FLAG-GGT7-expressing HepG2 cells. The band position corresponding to GGT7 or MYH10 is indicated with an asterisk. F The top ten list of GGT7-interacting protein partners identified by mass spectrometry. G Immunoblot analysis of total cell lysis and immunoprecipitants (using IgG control, GGT7, or MYH10 antibody) for indicated proteins derived from HepG2 cells. H Immunoblot analysis of MYH10 in HepG2 cells that have undergone CRISPR-mediated gene knockout. GAPDH serves as a loading control. I Decreased total cholesterol levels of HepG2 cells upon MYH10 knockout. Mean ± SD with n = 3. Unpaired two-sided t test, compared to AAVS1 KO, *p < 0.05. Decreased Dil-LDL uptake by HepG2 cells upon J GGT7 or K MYH10 knockout. Mean ± SD with n = 3. Unpaired two-sided t test, compared to AAVS1 KO, **p < 0.01, ***p < 0.001

Article Snippet: Human hepatic cancer cell line HepG2, cervix cancer cell line HeLa, and HEK293FT cells were obtained from the American Type Culture Collection (ATCC).

Techniques: RNA Expression, Quantitative RT-PCR, Plasmid Preparation, Control, Functional Assay, Staining, SDS Page, Immunoprecipitation, Expressing, Mass Spectrometry, Western Blot, Lysis, Derivative Assay, CRISPR, Gene Knockout, Knock-Out

Journal: Genome biology

Article Title: Systematic interrogation of functional genes underlying cholesterol and lipid homeostasis.

doi: 10.1186/s13059-025-03531-8

Figure Lengend Snippet: Fig. 7 Impaired cholesterol and lipid homeostasis in Ggt7 knockout mice. A The construction strategy of whole-body Ggt7 KO mice using two sgRNAs targeting the flanks of exon 2 and exon 3 of Ggt7 in the mouse genome. B Immunoblot analysis of Ggt7 in different types of tissues derived from Ggt7+/+ and Ggt7−/− mice. β-actin serves as a loading control. C Body weight measurements for Ggt7+/+ and Ggt7−/− mice under normal diet (ND) (n = 12 and 10, respectively) or high-fat high cholesterol (HFHC) diet (n = 7 and 8, respectively). Unpaired two-sided t test, *p < 0.05, ns means not significant. D The serum total cholesterol levels (n = 12, 10, 7 and 8 for each group), E serum HDL-cholesterol (HDL-C) levels (n = 10, 10, 7 and 8), F serum LDL-cholesterol (LDL-C) levels (n = 11, 10, 7 and 8) and G serum triglycerides (TG) levels (n = 12, 10, 7 and 8) in Ggt7+/+ and Ggt7−/− mice fed with ND or HFHC diet. Unpaired two-sided t test, *p < 0.05, **p < 0.01, ns means not significant. H The total cholesterol (TC) levels (n = 12, 10, 7 and 7) or I TG levels (n = 12, 10, 7 and 8) in the livers of Ggt7+/+ or Ggt7−/− mice fed with ND or HFHC diet. Unpaired two-sided t test, ns means not significant. J The TC levels (n = 12, 10, 7, and 7) or K TG levels (n = 12, 10, 7, and 7) in the brain tissues of Ggt7+/+ or Ggt7−/− mice fed with ND or HFHC diet. Unpaired two-sided t test, **p < 0.01, ns means not significant. L Volcano plot showing the DEGs upon Ggt7 knockout in the mouse liver or brain tissues. The number of up- or down-regulated DEGs is indicated. M Venn diagram showing the overlap of DEGs between Ggt7 KO in mouse liver, Ggt7 KO in mouse brain, and GGT7 KO in HepG2 cells as determined by RNA-seq analysis. N The top five enriched functional terms for either up-regulated or down-regulated DEGs in the liver or brain tissues of Ggt7−/− mice vs. Ggt7+/+ mice

Article Snippet: Human hepatic cancer cell line HepG2, cervix cancer cell line HeLa, and HEK293FT cells were obtained from the American Type Culture Collection (ATCC).

Techniques: Knock-Out, Western Blot, Derivative Assay, Control, RNA Sequencing, Functional Assay

Journal: Scientific reports

Article Title: Ferulic acid restores mitochondrial dynamics and autophagy via AMPK signaling pathway in a palmitate-induced hepatocyte model of metabolic syndrome.

doi: 10.1038/s41598-024-66362-w

Figure Lengend Snippet: Figure 1. Ferulic acid (FA) reduced mitochondrial reactive oxygen species (ROS) production and increased ATP levels in the MetS human HepG2 hepatocyte model. (A) Representative fluorescent images of mitochondrial ROS accumulation in HepG2 cells treated with/without 0.3 mM PA-BSA, 1 μM antimycin (positive control, 1-h treatment), and 50, 100, and 200 μM of FA (24-h treatments); (B) Quantification of mitochondrial ROS levels; (C) Quantification of total cellular ROS (D) Quantification of cellular ATP levels. Data are expressed as mean ± standard deviation (SD) for three independent samples. Bars with no letters in common are significantly different (p < 0.05).

Article Snippet: Human hepatic cancer HepG2 cells were obtained from the American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Positive Control, Standard Deviation

Journal: Scientific reports

Article Title: Ferulic acid restores mitochondrial dynamics and autophagy via AMPK signaling pathway in a palmitate-induced hepatocyte model of metabolic syndrome.

doi: 10.1038/s41598-024-66362-w

Figure Lengend Snippet: Figure 2. FA activated AMPK signaling in the MetS human HepG2 hepatocyte model. (A) Representative images of p-AMPKα, AMPKα, p-mTOR, mTOR, p-ULK, and ULK protein expressions; (B) Relative protein expression level of p-AMPKα /AMPKα ratio; (C) Relative protein expression level of p-mTOR /mTOR ratio; and (D) Relative protein expression level of p-ULK1 /ULK1 ratio. Data are expressed as mean ± standard deviation (SD) for three independent samples. Bars with no letters in common are significantly different (p < 0.05).

Article Snippet: Human hepatic cancer HepG2 cells were obtained from the American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Expressing, Standard Deviation

Journal: Scientific reports

Article Title: Ferulic acid restores mitochondrial dynamics and autophagy via AMPK signaling pathway in a palmitate-induced hepatocyte model of metabolic syndrome.

doi: 10.1038/s41598-024-66362-w

Figure Lengend Snippet: Figure 3. FA improved autophagy response in the MetS human HepG2 hepatocyte model. (A) Representative confocal fluorescent microscopy image showing autophagic vacuoles (green, FITC filter), nucleus (blue, DAPI filter), and the merge of the two channels in HepG2 cells treated with/without 0.3 mM PA-BSA, FA or rapamycin for 24 h; (B) Quantitative analysis of relative autophagy level calculated as a ratio between the fluorescence intensity of green and blue channels; (C) Representative image of Tom20 protein expression; (D) Relative protein expression level of Tom20. Data are expressed as mean ± standard deviation (SD) for three independent samples. Bars with no letters in common are significantly different (p < 0.05).

Article Snippet: Human hepatic cancer HepG2 cells were obtained from the American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Microscopy, Fluorescence, Expressing, Standard Deviation

Journal: Scientific reports

Article Title: Ferulic acid restores mitochondrial dynamics and autophagy via AMPK signaling pathway in a palmitate-induced hepatocyte model of metabolic syndrome.

doi: 10.1038/s41598-024-66362-w

Figure Lengend Snippet: Figure 4. FA enhanced protein levels of mitochondrial biogenesis-related markers including SirT1 (A, B), PGC-1α (A, C), NRF1 (A, D), and TFAM (A, E) in MetS human HepG2 hepatocyte model. Data are expressed as mean ± standard deviation (SD) for three independent samples. Bars with no letters in common are significantly different (p < 0.05).

Article Snippet: Human hepatic cancer HepG2 cells were obtained from the American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Standard Deviation

Journal: Scientific reports

Article Title: Ferulic acid restores mitochondrial dynamics and autophagy via AMPK signaling pathway in a palmitate-induced hepatocyte model of metabolic syndrome.

doi: 10.1038/s41598-024-66362-w

Figure Lengend Snippet: Figure 5. FA treatments enhanced MMP in MetS human HepG2 hepatocyte model. (A) Representative confocal fluorescent microscopy images of HepG2 cells stained with JC-1 dye. JC-1 monomers yielded green fluorescence with emission of 530 nm, and JC-1 aggregates yielded red fluorescence with emission of 590 nm. The combination of the two displays was shown in the merge. HepG2 cells were treated with/without 0.3 mM PA-BSA, FA for 24 h. 100 μM of FCCP was administered to the cells for 4 h as the negative control. (B) Quantitative analysis of MMP calculated as the ratio of red fluorescence counts/ green fluorescence counts. Data are expressed as mean ± standard deviation (SD) for three independent samples. Bars with no letters in common are significantly different (p < 0.05).

Article Snippet: Human hepatic cancer HepG2 cells were obtained from the American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Microscopy, Staining, Fluorescence, Negative Control, Standard Deviation

Journal: Scientific reports

Article Title: Ferulic acid restores mitochondrial dynamics and autophagy via AMPK signaling pathway in a palmitate-induced hepatocyte model of metabolic syndrome.

doi: 10.1038/s41598-024-66362-w

Figure Lengend Snippet: Figure 7. FA promoted mitochondrial fusion by activating its related regulators including OPA1 (A, B), Mitofusin-1 (A, C), and Mitofusin-2 (A, D) in MetS human HepG2 hepatocyte model. Data are expressed as mean ± standard deviation (SD) for three independent samples. Bars with no letters in common are significantly different (p < 0.05).

Article Snippet: Human hepatic cancer HepG2 cells were obtained from the American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Standard Deviation

Journal: Scientific reports

Article Title: Ferulic acid restores mitochondrial dynamics and autophagy via AMPK signaling pathway in a palmitate-induced hepatocyte model of metabolic syndrome.

doi: 10.1038/s41598-024-66362-w

Figure Lengend Snippet: Figure 6. FA suppressed mitochondrial fission through downregulating related protein markers in MetS human HepG2 hepatocyte model. (A) Representative images of MFF, p-DRP1(Ser616), DRP1; (B) Relative protein expression level of MFF; (C) Relative protein expression level of p-DRP1/DRP1. Data are expressed as mean ± standard deviation (SD) for three independent samples. Bars with no letters in common are significantly different (p < 0.05).

Article Snippet: Human hepatic cancer HepG2 cells were obtained from the American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: Expressing, Standard Deviation

Journal: Scientific reports

Article Title: Ferulic acid restores mitochondrial dynamics and autophagy via AMPK signaling pathway in a palmitate-induced hepatocyte model of metabolic syndrome.

doi: 10.1038/s41598-024-66362-w

Figure Lengend Snippet: Figure 8. Proposed mechanisms of action of FA on targeting mitochondrial activities and dynamics in MetS human HepG2 hepatocyte model. Created with BioRender.com.

Article Snippet: Human hepatic cancer HepG2 cells were obtained from the American Type Culture Collection (ATCC) (Manassas, VA).

Techniques: