Journal: Molecular Cancer

Article Title: Systematic analyses reveal long non-coding RNA (PTAF)-mediated promotion of EMT and invasion-metastasis in serous ovarian cancer

doi: 10.1186/s12943-018-0844-7

Figure Lengend Snippet: miR-25 knockdown promotes OvCa cell migration. a The expression of miR-25 in OvCa cells after treatment with 10 ng/ml TGF-β1 for 48 h. b The mRNA levels of EMT-related markers in SKOV3 cells transfected with a miR-25 inhibitor (AMO-25) or a negative control (NC). c Western blotting analysis of epithelial and mesenchymal markers in SKOV3 cells transfected with AMO-25. Wound-healing ( d ) and migration assays ( e ) were used to determine the effect of miR-25 inhibition on OvCa cell migration. n = 5 independent experiments. * P < 0.05, ** p < 0.01 vs. NC

Article Snippet: In accordance with the manufacturer’s instructions, stable PTAF knockdown or overexpression SKOV3 cell lines and respective control cell lines were generated using lentiviral vectors carrying either sh-PTAF, a PTAF overexpression construct, or the respective negative controls (Biowit Technology, Shenzhen, China).

Techniques: Knockdown, Migration, Expressing, Transfection, Negative Control, Western Blot, Inhibition

Journal: Molecular Cancer

Article Title: Systematic analyses reveal long non-coding RNA (PTAF)-mediated promotion of EMT and invasion-metastasis in serous ovarian cancer

doi: 10.1186/s12943-018-0844-7

Figure Lengend Snippet: Forced expression of miR-25 blunts TGF-β1-induced EMT and migration in OvCa cells. qRT-PCR ( a ) and western blot ( b ) analyses showed the inhibitory effect of miR-25 on EMT in SKOV3 cells treated with TGF-β1. A wound-healing assay displayed the inhibitory effects of miR-25 on TGF-β1-induced migration in SKOV3 cells ( c ) and in A2780 cells ( d ). A migration assay showed that miR-25 attenuated TGF-β1-induced migration in SKOV3 cells ( e ) and A2780 cells ( f ). n = 5 independent experiments. * P < 0.05, ** P < 0.01

Article Snippet: In accordance with the manufacturer’s instructions, stable PTAF knockdown or overexpression SKOV3 cell lines and respective control cell lines were generated using lentiviral vectors carrying either sh-PTAF, a PTAF overexpression construct, or the respective negative controls (Biowit Technology, Shenzhen, China).

Techniques: Expressing, Migration, Quantitative RT-PCR, Western Blot, Wound Healing Assay

Journal: Molecular Cancer

Article Title: Systematic analyses reveal long non-coding RNA (PTAF)-mediated promotion of EMT and invasion-metastasis in serous ovarian cancer

doi: 10.1186/s12943-018-0844-7

Figure Lengend Snippet: PTAF regulates the expression and activity of miR-25. a PTAF contains a sequence domain complementary to the seed motif of miR-25. b Up-regulation of PTAF in OvCa cells treated with 10 ng/ml TGF-β1 for 48 h. n = 6 independent experiments. ** P < 0.01 vs. Control. c - d Overexpression of PTAF inhibited the expression of miR-25 in SKOV3 cells. n = 6 independent experiments. ** P < 0.01 vs. pcDNA3.1. e - f Silencing of PTAF using a specific shRNA up-regulated miR-25 in SKOV3 cells. n = 6 independent experiments. * P < 0.05, ** P < 0.01 vs. sh-Scramble. g - h PTAF binds to miR-25 and regulates its activity. * P < 0.05, ** P < 0.01. i PTAF directly binding to miR-25. SKOV3 cells were transfected with biotin-tagged miR-Ctrl (Bio-miR-Ctrl) or biotin-tagged miR-25 (Bio-miR-25). Forty-eight hours after transfection, the cells were harvested for a biotin-based pull-down assay. PTAF expression levels were analyzed by qRT-PCR. ** P < 0.01. j Luciferase reporter activities of chimeric vectors carrying the luciferase gene and a fragment of PTAF containing the wild-type (WT) binding site or a mutated binding site for miR-25. ** P < 0.01. k - l qRT-PCR were used to examine the expression of PTAF in SKOV3 cells after overexpression or knockdown of miR-25. * P < 0.05, ** P < 0.01. n = 6 independent experiments

Article Snippet: In accordance with the manufacturer’s instructions, stable PTAF knockdown or overexpression SKOV3 cell lines and respective control cell lines were generated using lentiviral vectors carrying either sh-PTAF, a PTAF overexpression construct, or the respective negative controls (Biowit Technology, Shenzhen, China).

Techniques: Expressing, Activity Assay, Sequencing, Control, Over Expression, shRNA, Binding Assay, Transfection, Pull Down Assay, Quantitative RT-PCR, Luciferase, Knockdown

Journal: Molecular Cancer

Article Title: Systematic analyses reveal long non-coding RNA (PTAF)-mediated promotion of EMT and invasion-metastasis in serous ovarian cancer

doi: 10.1186/s12943-018-0844-7

Figure Lengend Snippet: PTAF silencing inhibits tumor progression in an orthotopic mouse model of OvCa. a Representative bioluminescence images of mice injected intraperitoneally with SKOV3 OvCa cells expressing either PTAF or empty pcDNA3.1 ( n = 10 in each group). b More tumor masses (yellow arrows) were formed by SKOV3-PTAF cells than by SKOV3-pcDNA3.1 cells. c Mice injected with SKOV3-PTAF cells showed liver metastases. d Representative images of tumor nodules and quantification of tumor nodules and tumor weights in mice injected intraperitoneally with SKOV3 OvCa cells expressing either PTAF or empty pcDNA3.1 ( n = 10 in each group). ** P < 0.01 vs. pcDNA3.1. e SKOV3-ip tumor samples from mice injected with cells expressing PTAF or empty pcDNA3.1 were immunohistochemically stained for E-cadherin and SNAI2. * P < 0.05 vs. pcDNA3.1. f Representative bioluminescence images in sh-Scramble- and sh-PTAF-treated mice ( n = 10 in each group) that were injected intraperitoneally with SKOV3 OvCa cells. g Fewer tumor masses (yellow arrows) were formed in the mice treated with sh-PTAF than in those treated with sh-Scramble. h Mice injected with sh-Scramble showed liver metastases, whereas very little metastasis was observed in mice injected with sh-PTAF. i Representative images of tumor nodules and quantification of tumor nodules and tumor weights in sh-Scramble- and sh-PTAF-treated mice injected with SKOV3 cells ( n = 10 for each group). ** P < 0.01 vs. sh-Scramble. j SKOV3-ip tumor samples from sh-Scramble- and sh-PTAF-treated mice were immunohistochemically stained for E-cadherin and SNAI2. * P < 0.05 vs. sh-Scramble. Scale bars, 100X = 100 μm; 400X = 20 μm

Article Snippet: In accordance with the manufacturer’s instructions, stable PTAF knockdown or overexpression SKOV3 cell lines and respective control cell lines were generated using lentiviral vectors carrying either sh-PTAF, a PTAF overexpression construct, or the respective negative controls (Biowit Technology, Shenzhen, China).

Techniques: Injection, Expressing, Staining

Journal: American Journal of Cancer Research

Article Title: Overexpression of MEF2D contributes to oncogenic malignancy and chemotherapeutic resistance in ovarian carcinoma

doi:

Figure Lengend Snippet: Analyses of MEF2D expression in OC cell lines and patient samples. A. Data on MEF2D DNA copy number and mRNA expression in ovarian cancer and normal ovarian tissue from several study groups deposited in the Oncomine database (www.oncomine.org). B. The protein expression of MEF2D in normal human ovarian epithelial cells (IOSE) and human OC cells (OVCAR3, SKOV3) was determined by western blot assays. C. The protein expression of MEF2D in normal ovarian tissues (N1-N4) and human OC tissues (T1-T12) was determined by western blot assays. Error bars represent the s.d. of triplicate measurements. *P < 0.05; **P < 0.01; ***P < 0.001. D. IHC analysis of MEF2D protein expression in normal ovarian tissues, and OC tissues Original magnifications: × 200 and × 400. E. IHC analysis of IKBKE and HPSE protein expression in normal ovarian tissues, and OC tissues Original magnifications: × 200 and × 400.

Article Snippet: Cell lines culture and establishment of DDP resistant cells Human ovarian cancer OVCAR3, SKOV3 cell lines and normal human ovarian epithelial cells IOSE were obtained from Sciencell (California, USA), and authenticated by short tandem repeat.

Techniques: Expressing, Western Blot

Journal: American Journal of Cancer Research

Article Title: Overexpression of MEF2D contributes to oncogenic malignancy and chemotherapeutic resistance in ovarian carcinoma

doi:

Figure Lengend Snippet: Effect of MEF2D knockdown on proliferation and apoptosis of OC cells in vitro. A. CCK-8 assays were performed to determine the effects of MEF2D knockdown on the proliferation of SKOV3 and OVCAR3 cells. Cell viability was determined at 0, 12, 24, 48 and 72 h. B. Flowcytometry assays were performed to determine the effects of MEF2D knockdown on the apoptosis of SKOV3 and OVCAR3 cells. C. Effects of MEF2D knockdown on proliferation-associated protein cyclin-D1 and c-myc and apoptosis-related protein caspase3 and cleaved caspase3 were analyzed by western blotting in SKOV3 and OVCAR3 cells. Error bars represent the s.d. of triplicate measurements. *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: Cell lines culture and establishment of DDP resistant cells Human ovarian cancer OVCAR3, SKOV3 cell lines and normal human ovarian epithelial cells IOSE were obtained from Sciencell (California, USA), and authenticated by short tandem repeat.

Techniques: Knockdown, In Vitro, CCK-8 Assay, Western Blot

Journal: American Journal of Cancer Research

Article Title: Overexpression of MEF2D contributes to oncogenic malignancy and chemotherapeutic resistance in ovarian carcinoma

doi:

Figure Lengend Snippet: Effect of MEF2D knockdown on invasion and migration of OC cells in vitro. A. The migration abilities of SKOV3 and OVCAR3 were measured through testing the wound closure after MEF2D knockdown using wound healing assays. B. Transwell assays were used to detect the migration and invasion abilities after MEF2D knockdown in SKOV3 and OVCAR3 cells. Original magnifications, × 200 and × 400. C. Effects of MEF2D knockdown on migration associated protein MMP9 were analyzed by western blotting in SKOV3 and OVCAR3 cells. Error bars represent the s.d. of triplicate measurements. *P < 0.05; **P < 0.01; ***P < 0.001.

Article Snippet: Cell lines culture and establishment of DDP resistant cells Human ovarian cancer OVCAR3, SKOV3 cell lines and normal human ovarian epithelial cells IOSE were obtained from Sciencell (California, USA), and authenticated by short tandem repeat.

Techniques: Knockdown, Migration, In Vitro, Western Blot

Journal: American Journal of Cancer Research

Article Title: Overexpression of MEF2D contributes to oncogenic malignancy and chemotherapeutic resistance in ovarian carcinoma

doi:

Figure Lengend Snippet: Effect of MEF2D knockdown on tumorigenecity of ovarian carcinoma cells in vivo. A, B. OVCAR3 cells were subcutaneously inoculated in SCID mice, which were randomly grouped to NC or siMEF2D001 (n = 6 for each group) and then injected with NC or siMEF2D001 every 3 days. 28 days later tumors were removed for analysis. Each tumor formed was volumed and weighted. The weight and volume of established tumors was measured and is shown in a scatter plot. C. Immunohistochemical analysis of MEF2D expression was performed on OVCAR3 tumor xenografts. The representative images are shown (with original magnification, of × 200 and × 400).

Article Snippet: Cell lines culture and establishment of DDP resistant cells Human ovarian cancer OVCAR3, SKOV3 cell lines and normal human ovarian epithelial cells IOSE were obtained from Sciencell (California, USA), and authenticated by short tandem repeat.

Techniques: Knockdown, In Vivo, Injection, Immunohistochemical staining, Expressing