Journal: Cell cycle (Georgetown, Tex.)

Article Title: PKN activation via transforming growth factor-beta 1 (TGF-beta 1) receptor signaling delays G2/M phase transition in vascular smooth muscle cells.

doi: 10.4161/cc.6.6.3985

Figure Lengend Snippet: Figure 5. PKN phosphorylates Cdc25C in PAC‑1 cells. Control cells were cotransfected with kinase‑dead PKN and wild type, myc‑tagged Cdc25C (left lane), and experimental cells were cotransfected with wild type PKN and wild type, myc‑tagged Cdc25C (right lane). 24 hrs post transfection, Y‑27632 was added to the control to pretreat the cells for 1 hr. Then both groups were penetrated with a‑toxin before exposed to TGF‑b1 and labeled with [g‑32P] ATP. Cdc25C was immunoprecipitated with anti‑myc antibody and applied to autoradiography to detect phosphorylation (upper panel). The immunoprecipitation with anti‑myc antibody without radiolabeling followed by anti‑myc Western blotting was performed to show the loading control (lower panel). A representative image of three independent experiments was shown.

Article Snippet: Lysates were processed for Cdc25C immunoprecipitation with an anti‐myc antibody (Santa Cruz Technology) at 4°C overnight, followed by protein G‐Sepharose (Sigma‐Aldrich) precipitation.

Techniques: Control, Transfection, Labeling, Immunoprecipitation, Autoradiography, Phospho-proteomics, Radioactivity, Western Blot

Journal: Cell cycle (Georgetown, Tex.)

Article Title: PKN activation via transforming growth factor-beta 1 (TGF-beta 1) receptor signaling delays G2/M phase transition in vascular smooth muscle cells.

doi: 10.4161/cc.6.6.3985

Figure Lengend Snippet: Figure 6. PKN coimmunoprecipitates with Cdc25C. (A) PAC‑1 cells were stimulated with vehicle control or TGF‑b1 for 1 and 3 hours. Endogenous Cdc25C or PKN was immunoprecipitated with rabbit anti‑Cdc25C Ab or mouse anti‑PKN Ab, followed by Western blot analysis with mouse anti‑PKN Ab (right upper panel) or rabbit anti‑Cdc25C Ab (right lower panel). Left panels show the input. (B) PAC‑1 cells were transfected with either empty vector (Control), active PKN (PKN‑AF3) or kinase‑dead PKN (K644E). Coimmunoprecipitation assay was performed with mouse anti‑PKN Ab followed by Western blot analysis with rabbit anti‑Cdc25C Ab. (C) PAC‑1 cells were cotransfected with Flag‑PKN (wild type) and Myc‑Cdc25C (wild type). Cdc25C or PKN was immunoprecipi‑ tated with mouse anti‑myc Ab or mouse anti‑Flag Ab, followed by Western blot analysis with mouse anti‑Flag Ab (right upper panel) or mouse anti‑myc Ab (right lower panel). Left panels show the input.

Article Snippet: Lysates were processed for Cdc25C immunoprecipitation with an anti‐myc antibody (Santa Cruz Technology) at 4°C overnight, followed by protein G‐Sepharose (Sigma‐Aldrich) precipitation.

Techniques: Control, Immunoprecipitation, Western Blot, Transfection, Plasmid Preparation, Co-Immunoprecipitation Assay

Journal: Cell cycle (Georgetown, Tex.)

Article Title: PKN activation via transforming growth factor-beta 1 (TGF-beta 1) receptor signaling delays G2/M phase transition in vascular smooth muscle cells.

doi: 10.4161/cc.6.6.3985

Figure Lengend Snippet: Figure 7. PKN and Cdc25C colocalize in dividing cells. PAC‑1 cells were synchronized at G1/S phase border and then released into the cell cycle. At 2 hr (S‑phase) (A) and 6 hr (M‑phase) (B and C) after synchronization release, PKN and Cdc25C were visualized by indirect immunocytochemistry. Nuclei were detected with DAPI staining. Arrows point to dividing cells. (A and B) 40 × Magnification, Fluorescence was visualized by traditional fluo‑ rescence microscopy. (C) 40 × Magnification, Fluorescence was visualized by confocal microscopy.

Article Snippet: Lysates were processed for Cdc25C immunoprecipitation with an anti‐myc antibody (Santa Cruz Technology) at 4°C overnight, followed by protein G‐Sepharose (Sigma‐Aldrich) precipitation.

Techniques: Immunocytochemistry, Staining, Fluorescence, Microscopy, Confocal Microscopy

Journal: Scientific Reports

Article Title: CDC25C downregulation suppresses HCC growth via mitochondrial stress-induced autophagy and apoptosis

doi: 10.1038/s41598-026-36351-2

Figure Lengend Snippet: The suppressed levels of CDC25C in cell line and xenograft model. ( A ) The CDC25C gene was detected via agarose gel electrophoresis, with β-actin serving as an internal control. The marker used was a 1000 bp Gene Ruler, and the expected product sizes were 970 bp for CDC25C and 709 bp for β-actin. The negative control prepared from a sample containing just the Master-Mix, in which there was no DNA contamination. ( B ) The relative expression of CDC25C mRNA was evaluated using qRT-PCR. ( C ) The expression level of CDC25C protein was assessed through Western blotting and quantified using ImageJ software, with GAPDH serving as the internal control. Data are presented as means ± SDs from three independent experiments. c p <0.001.

Article Snippet: CDC25C , Mouse , #66912-1-Ig , Proteintech , , 1:1000.

Techniques: Agarose Gel Electrophoresis, Control, Marker, Negative Control, Expressing, Quantitative RT-PCR, Western Blot, Software

Journal: Scientific Reports

Article Title: CDC25C downregulation suppresses HCC growth via mitochondrial stress-induced autophagy and apoptosis

doi: 10.1038/s41598-026-36351-2

Figure Lengend Snippet: Downregulation of CDC25C inhibited the malignant biological behaviors of Hepa1-6 cells. ( A ) The colony formation ability was assessed using a plate cloning assay at 100× magnification, with the colony formation rate calculated by counting the number of colonies containing more than 50 cells. ( B ) The lateral migration ability of cells was measured using a wound healing assay at 100× magnification, and the migration rate (scratch healing rate) was calculated by measuring the width of the scratch. ( C , D ) The longitudinal migration and invasion abilities of the cells were evaluated using Transwell assays, quantifying the number of cells that passed through the filter. The scale bar represents 20 μm. Data are presented as means ± SDs from three independent experiments. a p <0.05, b p <0.01, c p <0.001.

Article Snippet: CDC25C , Mouse , #66912-1-Ig , Proteintech , , 1:1000.

Techniques: Cloning, Migration, Wound Healing Assay

Journal: Scientific Reports

Article Title: CDC25C downregulation suppresses HCC growth via mitochondrial stress-induced autophagy and apoptosis

doi: 10.1038/s41598-026-36351-2

Figure Lengend Snippet: Downregulation of CDC25C altered the morphology of the subcellular structure of Hepa1-6 cells. The images of Hepa1-6 cells captured under transmission electron microscopy. The scale bar represents a measurement of 2 μm at lower magnification and 200 nm at higher magnification. White arrows indicate healthy mitochondria, orange arrows denote autophagosomes, blue arrow highlight cellular vacuolation, and red arrows signify swollen and dysfunctional mitochondria.

Article Snippet: CDC25C , Mouse , #66912-1-Ig , Proteintech , , 1:1000.

Techniques: Transmission Assay, Electron Microscopy

Journal: Scientific Reports

Article Title: CDC25C downregulation suppresses HCC growth via mitochondrial stress-induced autophagy and apoptosis

doi: 10.1038/s41598-026-36351-2

Figure Lengend Snippet: Downregulation of CDC25C induced a mitochondrial stress response. ( A , B ) Mitochondrial calcium concentrations were measured in Hepa1-6 and AML12 cells using the Rhod-2/AM fluorescent probe, with ImageJ software employed to quantify the red fluorescence intensity. ( C , D ) ROS levels were assessed in Hepa1-6 cells and AML12 using the MitoSOX fluorescent probe, and the red fluorescence intensity was quantified using ImageJ software. ( E ) The relative mRNA expression levels of CHOP, HSP60, ClpP, and LONP1 were evaluated via qRT-PCR. ( F , G ) The relative protein expression levels of CHOP, HSP60, ClpP, and LONP1 were determined through Western blotting and quantified using ImageJ software, with GAPDH serving as the internal control. The scale bar represents 10 μm. Data are presented as means ± SDs from three independent experiments. a p <0.05, b p <0.01, c p <0.001. ‘ns’ indicates no statistical significance.

Article Snippet: CDC25C , Mouse , #66912-1-Ig , Proteintech , , 1:1000.

Techniques: Software, Fluorescence, Expressing, Quantitative RT-PCR, Western Blot, Control

Journal: Scientific Reports

Article Title: CDC25C downregulation suppresses HCC growth via mitochondrial stress-induced autophagy and apoptosis

doi: 10.1038/s41598-026-36351-2

Figure Lengend Snippet: Downregulation of CDC25C induced an autophagic response. ( A ) The relative mRNA expression levels of LC3, p62, and Beclin1 were evaluated using qRT-PCR. ( B , C ) The relative protein expression levels of LC3, p62, and Beclin1 were determined through Western blotting and quantified using ImageJ software, with GAPDH serving as the internal control. Data are presented as means ± SDs from three independent experiments. a p < 0.05, b p < 0.01, c p <0.001.

Article Snippet: CDC25C , Mouse , #66912-1-Ig , Proteintech , , 1:1000.

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Software, Control

Journal: Scientific Reports

Article Title: CDC25C downregulation suppresses HCC growth via mitochondrial stress-induced autophagy and apoptosis

doi: 10.1038/s41598-026-36351-2

Figure Lengend Snippet: Downregulation of CDC25C induced a mitochondria-mediated apoptosis. ( A ) Hoechst 33258 staining was employed to morphologically identify apoptotic cells. ( B ) Annexin V/TMRE costaining was utilized for apoptosis detection via flow cytometry to investigate the apoptosis rate and quantified using ImageJ software. ( C ) The relative mRNA expression levels of Cyt c, Caspase-3 and Caspase-9 were evaluated using qRT-PCR. ( D , E ) The relative protein expression levels of Cyt c, Caspase-3 and Caspase-9 were determined through Western blotting and quantified using ImageJ software, with GAPDH serving as the internal control. The scale bar represents 50 μm. Data are presented as means ± SDs from three independent experiments. a p < 0.05, b p < 0.01, c p <0.001. ‘ns’ indicates no statistical significance.

Article Snippet: CDC25C , Mouse , #66912-1-Ig , Proteintech , , 1:1000.

Techniques: Staining, Flow Cytometry, Software, Expressing, Quantitative RT-PCR, Western Blot, Control

Journal: Cell Death Discovery

Article Title: ABCB1 confers resistance to carboplatin by accumulating stem-like cells in the G2/M phase of the cell cycle in p53 null ovarian cancer

doi: 10.1038/s41420-025-02435-7

Figure Lengend Snippet: A Cell cycle analysis in carR-SKOV3 and -OVCAR3 compared to each nonR group. (SKOV3: nonR; n = 16, carR; n = 13, OVCAR3: nonR; n = 10, carR; n = 12) Cell cycle distribution in each phase of the cell cycle was shown in graphs ( B ). Statistical analyses were conducted between nonR and carR cells that are distributed in the G2/M phase (SKOV3 nonR vs carR: p < 0.001/OVCAR3 nonR vs carR: p = 0.0042). C Immunoblotting analyses of G2/M transition related-proteins (phospho-cdc25c and CCNB1) in carR-SKOV3 and -OVCAR3 compared to each nonR group. Loading control: β-actin. Analysis of stemness profiling of each phase of the cell cycle in carR-SKOV3 (CD133: 1.55-fold, p < 0.001; LGR5: 1.79-fold, p = 0.005; CD117: 1.61-fold, p < 0.001; SOX2: 1.89-fold, p = 0.005) ( D ) and -OVCAR3 (CD133: p = 0.512; LGR5: p = 0.952; CD117: p = 0.839; SOX2: p = 0.082) ( E ) compared to nonR groups, respectively. F Flow cytometry plots showing the Ki67-positive cell proportion in carR-SKOV3 ( p = 0.0087) and -OVCAR3 ( p = 0.9469) compared to each nonR group. Comparison of Ki67-positive proportion in G2/M-gated fraction was displayed in graph ( G ). Data represent the means ± SD from multiple experiments. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001, NS not significant.

Article Snippet: After blocking procedure, membranes were incubated with primary antibodies against p53 (7F5), phospho-p53 (E9Y4U; Ser15), PARP1 (46D11), cleaved-PARP1 (D64E10; Asp214), BCL-XL (54H6), phospho-cdc25c (63F9; Ser216), cyclin B1 (D5C10), ΔNp73 (38C674.2; Novus, USA), and β-actin (all from Cell signaling, USA), respectively, and finally visualized by ECL solution (Thermo, USA).

Techniques: Cell Cycle Assay, Western Blot, Control, Flow Cytometry, Comparison