Journal: Oncology reports

Article Title: miRNA-regulated expression of oncogenes and tumor suppressor genes in the cisplatin-inhibited growth of K562 cells.

doi: 10.3892/or_00000813

Figure Lengend Snippet: Figure 2. Changes in mRNA expression of BCL2, E2F1, E2F3, RB1 and P53 in K562 cells after cisplatin treatment. BCL2, E2F1, E2F3, RB1 and P53 were detected by (A) RT-PCR, (B) real-time PCR and (C) ELISA.

Article Snippet: K562 cells were lysed [lysis buffer: 0.15 M NaCl, 5 mM EDTA (pH 8.0), 1% Triton X-100, 10 mM Tris-Cl (pH 7.4), 100 mM PMSF and 5 M DTT) and incubated in a 96-well plate, followed by the addition of goat anti-human antibodies against BCL2 (1:400), E2F1, E2F3 (1:400, 1:400), RB1 (1:500) and P53 (1:300, 1:400) (all from Immunoleader, Boster) according to the manufacturer's instructions.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

Journal: Oncology reports

Article Title: miRNA-regulated expression of oncogenes and tumor suppressor genes in the cisplatin-inhibited growth of K562 cells.

doi: 10.3892/or_00000813

Figure Lengend Snippet: Figure 4. Correlative expression of miRNAs and oncogenes using antisense oligos (ASO). (A) RT-PCR, (B) real-time PCR. Correlative expression of (C) E2F1 and its targeted miR-17-5p, (D) E2F3 and its targeted miRNAs and (E) Bcl-2 and its targeted miRNAs (miR-16, 34a-c) using ELISA is shown. *Significant difference (p<0.05).

Article Snippet: K562 cells were lysed [lysis buffer: 0.15 M NaCl, 5 mM EDTA (pH 8.0), 1% Triton X-100, 10 mM Tris-Cl (pH 7.4), 100 mM PMSF and 5 M DTT) and incubated in a 96-well plate, followed by the addition of goat anti-human antibodies against BCL2 (1:400), E2F1, E2F3 (1:400, 1:400), RB1 (1:500) and P53 (1:300, 1:400) (all from Immunoleader, Boster) according to the manufacturer's instructions.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

Journal: Oncology reports

Article Title: miRNA-regulated expression of oncogenes and tumor suppressor genes in the cisplatin-inhibited growth of K562 cells.

doi: 10.3892/or_00000813

Figure Lengend Snippet: Figure 5. Expression of E2F1, E2F3, BCL2, RB1 and P53 genes regulated by miRNAs. Detection of the expression of (A) E2F1, (B) E2F3, (C) BCL2, (D) RB1 and (E) P53. *Significant difference (p<0.05).

Article Snippet: K562 cells were lysed [lysis buffer: 0.15 M NaCl, 5 mM EDTA (pH 8.0), 1% Triton X-100, 10 mM Tris-Cl (pH 7.4), 100 mM PMSF and 5 M DTT) and incubated in a 96-well plate, followed by the addition of goat anti-human antibodies against BCL2 (1:400), E2F1, E2F3 (1:400, 1:400), RB1 (1:500) and P53 (1:300, 1:400) (all from Immunoleader, Boster) according to the manufacturer's instructions.

Techniques: Expressing

Journal: The Journal of urology

Article Title: A Murine Model of K-RAS and β-Catenin Induced Renal Tumors Expresses High Levels of E2F1 and Resembles Human Wilms Tumor.

doi: 10.1016/j.juro.2015.04.090

Figure Lengend Snippet: Figure 5. E2F1 is up-regulated in murine KrasG12D/CatnbDex3 kidneys. Mice were bred and sacrificed at ages 15 to 20 weeks. Kidneys were snap frozen, RNA was extracted and RT-PCR was performed for E2F1 (A). Results were compared to kidneys from controls and analyzed across groups using 2eDDC T .16 BCat, b-catenin. Asterisk indicates p <0.05. Whole protein lysates were extracted and tested by immunoblot (B). Ctrl, control. Formalin fixed, paraffin embedded kidneys were tested for E2F1 expression by IHC (C to F ). Reduced from 20 (C to F ).

Article Snippet: The antibodies used for IHC and/or immunoblotting were c-Myc (Epitomics ), Actin (Sigma-Aldrich ), Axin2 (Abcam ), E2F1 (Atlas Antibodies, Stockholm, Sweden), survivin and Cyclin D1 (Cell Signaling Technology ).

Techniques: Reverse Transcription Polymerase Chain Reaction, Western Blot, Control, Expressing

Journal: The Journal of urology

Article Title: A Murine Model of K-RAS and β-Catenin Induced Renal Tumors Expresses High Levels of E2F1 and Resembles Human Wilms Tumor.

doi: 10.1016/j.juro.2015.04.090

Figure Lengend Snippet: Figure 6. E2F1 is expressed at high levels in most human WTs. IHC was performed for E2F1 on TMA of 32 human WTs. Strong nuclear staining was seen in 30 of 32 WTs (94%) and seen in blastemal elements at low power (A) and high power (B), in stromal elements (asterisk, B), and epithelial elements at low power (C ) and high power (D). Reduced from 20 (A and C ) and 40 (B and D).

Article Snippet: The antibodies used for IHC and/or immunoblotting were c-Myc (Epitomics ), Actin (Sigma-Aldrich ), Axin2 (Abcam ), E2F1 (Atlas Antibodies, Stockholm, Sweden), survivin and Cyclin D1 (Cell Signaling Technology ).

Techniques: Staining

Journal: Frontiers in Cell and Developmental Biology

Article Title: Safrana l Prevents Prostate Cancer Recurrence by Blocking the Re-activation of Quiescent Cancer Cells via Downregulation of S-Phase Kinase-Associated Protein 2

doi: 10.3389/fcell.2020.598620

Figure Lengend Snippet: The inhibitory effects of safranal on AKT phosphorylation and transcriptional activity of E2F1 during quiescent Pca cell re-entry. Quiescent LNCaP (A) and PC-3 (B) cells were initially stimulated to re-enter the cell cycle, and the effects of safranal (GI 90 ) on protein expression levels of p-AKT (Ser473), AKT, and E2F1 at specific intervals were determined by immunoblotting. GAPDH served as a loading control. The mRNA expression levels of E2F1 in LNCaP (C) and PC-3 (D) cells during cell cycle re-entry in the presence or absence of safranal (GI 90 ) were examined by RT-qPCR. E2F1 levels in cell nuclear and cytoplasm extracts of LNCaP (E) and PC-3 cells (F) were analyzed using immunoblotting after treatment with safranal for 12 and 3 h, respectively, following release from the quiescent state. α-Tubulin and lamin A/C served as loading and purity controls for the cytoplasm and nuclear fractions, respectively. Following a 6-day serum withdrawal for LNCaP and 2-day contact inhibition for PC-3 cells, quiescent LNCaP (G) and PC-3 (H) were transfected with pGM-E2F-Luc and renilla luciferase reporter plasmid pML-SV40-hRluc by using the EZ transfection agent for 15 h. The transfected quiescent cells were then induced to re-enter the cell cycle in the presence or absence of safranal (GI 90 ) for 24 h. Cell lysates were collected to assess firefly and renilla luciferase activities using the dual luciferase reporter assay kit. Renilla luciferase served to normalize the values of the experimental reporter gene and acted as an internal control for transfection efficiency. Data are expressed as mean ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001 vs. dimethyl sulfoxide vehicle control.

Article Snippet: Then, the samples were stained with hematoxylin and eosin and with antibodies against Ki-67 (Abcam, ab16667), NF-κB p65 (Santa Cruz, sc514451), p-IκBα (Santa Cruz, sc8404), p21 (Proteintech, #10355-1-AP), CDK4 (Epitomics, #3830-1), CDK6 (Proteintech, #14052-1-AP), CDK2 (Abcam ab32147), p-Rb (Ser807, Abcam, ab184796), E2F1 (St John’s Laboratory, STJ92807), Skp2 (Santa Cruz, sc7164), c-MYC (Abcam, ab32072), and p27 (sc528, Santa Cruz).

Techniques: Phospho-proteomics, Activity Assay, Expressing, Western Blot, Control, Quantitative RT-PCR, Inhibition, Transfection, Luciferase, Plasmid Preparation, Reporter Assay

Journal: Frontiers in Cell and Developmental Biology

Article Title: Safrana l Prevents Prostate Cancer Recurrence by Blocking the Re-activation of Quiescent Cancer Cells via Downregulation of S-Phase Kinase-Associated Protein 2

doi: 10.3389/fcell.2020.598620

Figure Lengend Snippet: Safranal suppresses the tumor growth of quiescent PC-3 cells in vivo . Quiescent PC-3 cells were xenografted subcutaneously into the left flank of male nude mice. Then, the mice were orally administrated with either the vehicle control or 100 mg/kg of safranal, commencing a day prior to the xenograft procedure. Tumor volume (A) and mice body weight (B) were measured on alternate days. After a 46-day treatment, the mice were sacrificed, and the tumor (C) and major organs (D) were resected and photographed (scale bar: 20 μm). Paraffin-embedded tumor tissues were stained with hematoxylin and eosin and with antibodies against Ki-67, Skp2, E2F1, NF-κB p65, p-IκBα (Ser32), c-MYC, p-Rb (Ser807), CDK4, CDK6, CDK2, E2F1, Skp2, p21, and p27; representative images ( E , ×400) and quantification data (F) are presented. All data are shown as mean ± SD. * P < 0.05, ** P < 0.01, *** P < 0.001 vs. control.

Article Snippet: Then, the samples were stained with hematoxylin and eosin and with antibodies against Ki-67 (Abcam, ab16667), NF-κB p65 (Santa Cruz, sc514451), p-IκBα (Santa Cruz, sc8404), p21 (Proteintech, #10355-1-AP), CDK4 (Epitomics, #3830-1), CDK6 (Proteintech, #14052-1-AP), CDK2 (Abcam ab32147), p-Rb (Ser807, Abcam, ab184796), E2F1 (St John’s Laboratory, STJ92807), Skp2 (Santa Cruz, sc7164), c-MYC (Abcam, ab32072), and p27 (sc528, Santa Cruz).

Techniques: In Vivo, Control, Staining