Journal: Oncology Reports

Article Title: Pituitary tumor‑transforming gene 1 regulates the senescence and apoptosis of oral squamous cell carcinoma in a p21‑dependent DNA damage response manner

doi: 10.3892/or.2024.8794

Figure Lengend Snippet: PTTG1 expression in relation to cell proliferation, cell cycle and apoptosis in OSCC cells. (A) PTTG1 expression was analyzed by reverse transcription quantitative PCR in OSCC cells. (B) Protein expression of PTTG1 and PCNA in OSCC cell lines revealed by western blotting. The left panel shows the membranes stained with antibodies, with GAPDH as an internal control, and the right panel shows the relative quantification of protein expression. (C and D) Representative images of cell proliferation ability (left) and quantification (right) of PTTG1 using the EdU assay in the (C) HSC-2 and (D) SCC-9 cell lines (scale bars, 100 µm and 50 µm, respectively; original magnification, ×40). The percentages of HSC-2 and SCC-9 cells treated with VC or siR-PTTG1 were determined by EdU incorporation (green) and DAPI (blue). (E) Protein expression (left) and the fold change (right) of cell cycle markers, including cyclins D1, E, and B1 in OSCC cells. (F) Protein expression (left) and fold change (right) related to apoptosis markers, including Cas-7, c-Cas-7, and c-PARP in OSCC cells. *P<0.05, **P<0.01 and ***P<0.001 vs. VC using Student's t-test. All experiments were performed in triplicate. PTTG1, pituitary tumor-transforming gene 1; OSCC, oral squamous cell carcinoma; PCNA, proliferating cell nuclear antigen; VC, vehicle control; siR-PTTG1, small interfering RNA-PTTG1; EdU, 5-ethynyl-2′-deoxyuridine; Cas-7, Caspase-7; c-, cleaved-; PARP, poly (ADP-ribose) polymerase.

Article Snippet: Next, the membranes were incubated with the following primary antibodies overnight at 4°C: Rabbit anti-PTTG1 (cat. no. GTX111938; GeneTex, Inc.), anti-p21 (cat. no. 2947; Cell Signaling Technology, Inc.), anti-p53 (cat. no. 2527; Cell Signaling Technology, Inc.), anti-retinoblastoma (Rb; cat. no. 9390; Cell Signaling Technology, Inc.), anti-phosphorylated Rb (cat. no. 8180; Cell Signaling Technology, Inc.), anti-proliferating cell nuclear antigen (PCNA; cat. no. 2586; Cell Signaling Technology, Inc.), mouse anti-Caspase-7 (Cas-7; cat no. sc-56063; Santa Cruz Biotechnology, Inc.), anti-cleaved (c-) Cas-7 (cat. no. 8438; Cell Signaling Technology, Inc.), anti-c-poly (ADP-ribose) polymerase (PARP; cat. no. 5625; Cell Signaling Technology, Inc.), anti-PARP (cat. no. 9542; Cell Signaling Technology, Inc.), mouse anti-cyclin D1 (cat. no. sc-450; Santa Cruz Biotechnology, Inc.), mouse anti-cyclin E (cat. no. sc-247; Santa Cruz Biotechnology, Inc.), mouse anti-cyclin B1 (cat. no. sc-245; Santa Cruz Biotechnology, Inc.), anti-phosphorylated histone H2AX (γH2AX; cat. no. 80312; Cell Signaling Technology, Inc.), anti-phosphorylated ATR (cat. no. 2853; Cell Signaling Technology, Inc.) anti-ATR (cat. no. 2790; Cell Signaling Technology, Inc.), anti-phosphorylated ataxia telangiectasia mutant (p-ATM; cat. no. 13050; Cell Signaling Technology, Inc.) and anti-ATM (cat. no. 2873; Cell Signaling Technology, Inc.); all primary antibodies were diluted at 1:1,000 with BSA.

Techniques: Expressing, Reverse Transcription, Real-time Polymerase Chain Reaction, Western Blot, Staining, Control, EdU Assay, Small Interfering RNA

Journal: Cell Division

Article Title: Overexpression of ZFP69B promotes hepatocellular carcinoma growth by upregulating the expression of TLX1 and TRAPPC9

doi: 10.1186/s13008-024-00131-z

Figure Lengend Snippet: Overexpression of TRAPPC9 overturns the effects of TLX1 knockdown on Huh-7 and MHCC97H cells. ( A ) Expression of TRAPPC9 mRNA in Huh-7 and MHCC97H in response to sh-TLX1 + oe-NC or sh-TLX1 + oe-TRAPPC9 was analyzed using RT-qPCR. ( B ) The viability of Huh-7 and MHCC97H cells was examined using CCK-8 assays. ( C ) The colony formation of Huh-7 and MHCC97H cells was determined using colony formation assays. ( D D) The cell apoptosis of Huh-7 and MHCC97H cells was determined using flow cytometry. ( E ) The protein expression of p-P65, Bcl-2, and Cyclin D1 in Huh-7 and MHCC97H cells was determined using western blot analysis. Data represent the mean ± SEM of at least three independent experiments. ** p < 0.01, *** p < 0.001, **** p < 0.0001. Differences were tested using an unpaired t-test ( A , C , D ) and the two-way ANOVA ( B , E )

Article Snippet: After blocking, membranes were incubated first with primary antibodies to p-P65 (1:1000, ab76302, Abcam), Bcl2 (1:2000, ab182858, Abcam), Cyclin D1 (1:20, MA5-16356, Thermo Fisher Scientific) and β-actin (1:5000, GTX110564, GeneTex) and then with horseradish peroxidase-coupled goat anti-rabbit IgG antibody (1:100, GTX213110-01, GeneTex) for 2 h at room temperature.

Techniques: Over Expression, Knockdown, Expressing, Quantitative RT-PCR, CCK-8 Assay, Flow Cytometry, Western Blot