Journal: Journal of Translational Medicine

Article Title: Hsa_circ_0016662 acts as a prognostic biomarker and promotes the progression of hepatocellular carcinoma via autophagy regulation

doi: 10.1186/s12967-025-06763-1

Figure Lengend Snippet: Validation of hsa_circ_0016662 regulatory network in real-world cohort and HCC cell lines. (A) - (B) The expression of hsa_circ_0016662 and hsa-miR-22-3p validated in Guangxi local HCC samples by qRT-PCR ( n = 31). (C) - (D) Representative IHC staining images of YWHAZ in local HCC tissues and adjacent normal liver tissues. (E) The IHC scores of YWHAZ were compared between local HCC tissues and adjacent normal liver tissues ( n = 36). (F) - (H) qRT-PCR revealed the mRNA expression of hsa_circ_0016662, hsa-miR-22-3p and YWHAZ in HCC cell lines comparing to the normal liver cell L02. (I) The protein expression of YWHAZ in HCC cell lines and L02 detected by western blot. * : P < 0.05, *** : P < 0.001

Article Snippet: Six HCC cell lines (Huh7, HCCLM3, HepG2, MHCC-97 H, MHCC-97 L and SNU-449) and normal human liver cell L02 were obtained from Procell Life Science & Technology (Wuhan, China) and cultured in Dulbecco’s modified Eagle’s medium (DMEM, Invitrogen) or RPMI-1640 containing 10% fetal bovine serum (Procell Life Science & Technology).

Techniques: Biomarker Discovery, Expressing, Quantitative RT-PCR, Immunohistochemistry, Western Blot

Journal: Journal of Translational Medicine

Article Title: Hsa_circ_0016662 acts as a prognostic biomarker and promotes the progression of hepatocellular carcinoma via autophagy regulation

doi: 10.1186/s12967-025-06763-1

Figure Lengend Snippet: Knockdown of hsa_circ_0016662 suppressed HCC progression in vivo. (A) Subcutaneous xenografts of Huh7 cells dissected from nude mice. (B) - (C) The tumor volume and tumor weight were calculated and compared in samples from the subcutaneous xenograft model. (D) Hsa_circ_0016662 expression in the subcutaneous xenograft model was detected by qRT-PCR. (E) Representative staining images of HE and Ki-67 in tumors from the subcutaneous xenograft model. (F) Representative HE images of lung metastatic foci derived from the mice of metastatic model. (G) The metastatic proportion was calculated in the lung metastatic model. ** P < 0.001, *** : P < 0.001, **** : P < 0.0001

Article Snippet: Six HCC cell lines (Huh7, HCCLM3, HepG2, MHCC-97 H, MHCC-97 L and SNU-449) and normal human liver cell L02 were obtained from Procell Life Science & Technology (Wuhan, China) and cultured in Dulbecco’s modified Eagle’s medium (DMEM, Invitrogen) or RPMI-1640 containing 10% fetal bovine serum (Procell Life Science & Technology).

Techniques: Knockdown, In Vivo, Expressing, Quantitative RT-PCR, Staining, Derivative Assay

Journal: Frontiers in Immunology

Article Title: PSMD12 promotes hepatocellular carcinoma progression by stabilizing CDK1

doi: 10.3389/fimmu.2025.1581398

Figure Lengend Snippet: PSMD12 promotes HCC cell cycle progression and interacts with CDK1. (A) GSEA analysis of the TCGA-HCC cohort highlighting cell cycle signaling pathways. (B-E) Cell cycle distribution in HCC cells following PSMD12 knockdown or overexpression, presented as peak plots and quantitative analysis of cell distribution across G0/G1, S, and G2/M phases. (F, G) Western blot analysis of PSMD12, PCNA, Cyclin D1, CDK1, and GAPDH protein expression in PSMD12-silenced MHCC97H cells (F) and PSMD12-overexpressing HCCLM3 cells (G) , with GAPDH as the control. (H) GSEA analysis of the TCGA-HCC cohort revealing the Ubiquitin-mediated proteolysis and G2/M checkpoint signaling pathways. (I, J) Mass spectrometry detected co-precipitated PSMD12 and CDK1 proteins. (K-N) Co-immunoprecipitation (Co-IP) assays in MHCC97H and HCCLM3 cells confirmed the interaction between PSMD12 and CDK1. (O, P) Co-localization of PSMD12 (red) and CDK1 (green) in HCC cells, followed by DAPI nuclear counterstaining (blue). Scale bar: 20 μm. *p < 0.05, **p < 0.01, ***p < 0.001; ns, not significant.

Article Snippet: Additional human HCC cell lines HCCLM3 (CL-0042) and SMCC7721 (CL-0216) were obtained from Procell (Wuhan, China), and the MHCC97H (ZB001) cell line was acquired from Bei Zhi Creatures (Shanghai, China).

Techniques: Protein-Protein interactions, Knockdown, Over Expression, Western Blot, Expressing, Control, Ubiquitin Proteomics, Mass Spectrometry, Immunoprecipitation, Co-Immunoprecipitation Assay

Journal: Frontiers in Immunology

Article Title: PSMD12 promotes hepatocellular carcinoma progression by stabilizing CDK1

doi: 10.3389/fimmu.2025.1581398

Figure Lengend Snippet: PSMD12 regulates HCC proliferation through CDK1. (A) Western blot analysis of PSMD12, CDK1, PLK1, p-PLK1, AKT, p-AKT, and GAPDH expression in MHCC97H cells transfected with shNC, Vector, shPSMD12#1, or CDK1. (B) Western blot analysis showing protein expression of PSMD12, CDK1, PLK1, p-PLK1, AKT, p-AKT, and GAPDH in HCCLM3 cells transfected with Vector, PSMD12, or shCDK1. (C) CCK-8 assays demonstrated that restoration of CDK1 expression counteracted the growth-inhibitory effect of PSMD12 knockdown in MHCC97H cells. (D) Knockdown of CDK1 expression inhibited the pro-proliferative effect induced by PSMD12 overexpression in HCCLM3 cells, as assessed by CCK-8. (E, F) Representative images and quantification of colony formation assays in hepatocellular carcinoma cells following PSMD12 or CDK1 knockdown or overexpression. (G, H) EdU assays were performed to assess the proliferative capacity of cells (scale bar: 200 μm). (I, J) Quantification of EdU assays in hepatocellular carcinoma cells following PSMD12 or CDK1 knockdown or overexpression. Data are presented as mean ± standard deviation from three independent experiments and analyzed using one-way ANOVA followed by Tukey’s multiple comparisons test. **p < 0.01, ***p <0.001. (K) Representative tumor morphology in BALB/c nude mice. (L, M) Statistical analysis of tumor volume (L) and tumor weight (M) across different groups. Tumor volume (V) was calculated as V = 0.52 × length × width2, and data were analyzed by one-way ANOVA followed by Tukey’s multiple comparisons test. Tumor volumes are presented as mean ± SD, n = 4, **p < 0.01, ***p < 0.001. (N) Representative H&E and IHC staining of PSMD12, CDK1, and Ki67 in tumor tissues from different nude mouse groups. Scale bar: 50 μm.

Article Snippet: Additional human HCC cell lines HCCLM3 (CL-0042) and SMCC7721 (CL-0216) were obtained from Procell (Wuhan, China), and the MHCC97H (ZB001) cell line was acquired from Bei Zhi Creatures (Shanghai, China).

Techniques: Western Blot, Expressing, Transfection, Plasmid Preparation, CCK-8 Assay, Knockdown, Over Expression, Standard Deviation, Immunohistochemistry

Journal: Frontiers in Immunology

Article Title: PSMD12 promotes hepatocellular carcinoma progression by stabilizing CDK1

doi: 10.3389/fimmu.2025.1581398

Figure Lengend Snippet: PSMD12 stabilizes CDK1 protein levels by reducing ubiquitin-mediated degradation of CDK1 in HCC cells. (A, B) Western blot analysis of PSMD12 and CDK1 protein levels in HCC cells following PSMD12 knockdown or overexpression, with GAPDH as a loading control. (C, D) qRT-PCR assessment of PSMD12 and CDK1 mRNA levels in HCC cells after PSMD12 knockdown or overexpression, with GAPDH as a control. (E, F) MHCC97H and HCCLM3 cells were treated with cycloheximide (CHX, 50 μg/mL), and Western blot analysis was performed to detect CDK1 protein levels at different time points. (G, H) MHCC97H and HCCLM3 cells were treated with CHX (50 μg/mL) for specified durations, with or without the addition of the PSMD12 overexpression plasmid, followed by Western blot analysis to assess CDK1 protein levels. (I, J) HCC cells were treated with 15 μmol/L MG132 and collected at 0/3/6/9 hours, followed by Western blot to analyze protein expression levels. (K, L) MG132 (15 μM) treatment of MHCC97H and HCCLM3 cells altered PSMD12 expression, with PSMD12 and CDK1 protein levels assessed by Western blot analysis. (M, N) Co-immunoprecipitation (Co-IP) assays in MHCC97H and HCCLM3 cells confirmed the interaction between ubiquitin (Ub) and CDK1. (O) Colocalization studies in HCC cells using an anti-CDK1 antibody (1:100, red) and anti-ubiquitin antibody (1:100, green), followed by DAPI nuclear counterstaining (blue). Scale bar: 20 mm. (P, Q) MG132 (15 μM) was added to MHCC97H and HCCLM3 cells, which were simultaneously transfected with shPSMD12#1 or HA-PSMD12 plasmids. Co-IP assays were performed to detect ubiquitin binding to CDK1. (R) After 48 hours of transfection of Myc-PSMD12, Flag-CDK1, and HA-Ub WT/K48/K63, cells were lysed, and then immunoprecipitation was performed with Flag antibody, and immunoblot analysis was performed with HA antibody. (S) The docking conformation and three-dimensional structure of PSMD12 and CDK1.PSMD12 and CDK1 are shown in orange and cyan respectively. Statistical significance was determined using Student's t-tests and one-way ANOVA followed by Tukey's multiple comparisons test. *p < 0.05, **p < 0.01, ***p < 0.001; ns, not significant.

Article Snippet: Additional human HCC cell lines HCCLM3 (CL-0042) and SMCC7721 (CL-0216) were obtained from Procell (Wuhan, China), and the MHCC97H (ZB001) cell line was acquired from Bei Zhi Creatures (Shanghai, China).

Techniques: Ubiquitin Proteomics, Western Blot, Knockdown, Over Expression, Control, Quantitative RT-PCR, Plasmid Preparation, Expressing, Immunoprecipitation, Co-Immunoprecipitation Assay, Transfection, Binding Assay

Journal: International Journal of Preventive Medicine

Article Title: Effect of Zebularine on Apoptotic Pathways in Hepatocellular Carcinoma Cell Lines

doi: 10.4103/ijpvm.ijpvm_191_21

Figure Lengend Snippet: The effect of zebularine (0, 10, 25, 50, 75, and 100 μM) on the viability of HCC cell lines, HCCLM3, MHCC97H, and MHCC97L. The HCC cell lines were treated without and with different doses of zebularine for 24 and 48 h, and the cell viability was investigated by MTT assay. Asterisks (*) indicate significant differences between treated and untreated cells

Article Snippet: HCC cell lines (HCCLM3, MHCC97H, and MHCC97L) were obtained from the National Cell Bank of Iran-Pasteur Institute.

Techniques: MTT Assay