Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: EEF1D overexpression promotes osteosarcoma cell proliferation by facilitating Akt-mTOR and Akt-bad signaling

doi: 10.1186/s13046-018-0715-5

Figure Lengend Snippet: EEF1D is upregulated in osteosarcoma tissues and cell lines. a , b , c EEF1D mRNA and protein expression levels in osteosarcoma cell lines (MNNG/HOS, MG63 and U2OS) and human normal osteoblast cell line (hFOB 1.19) were measured by qRT-PCR and western blotting, respectively. d , e EEF1D expression levels were measured in 20 pairs of osteosarcoma and corresponding adjacent non-tumor tissues. For qRT-PCR, EEF1D expression was normalized to β-actin

Article Snippet: After blocking in 5% nonfat milk, the membranes were incubated with the following primary antibodies: EEF1D (1:500, Proteintech) [ ], mTOR (total, 1:1000; Cell Signaling Technology), phospho-mTOR (Ser2448, 1:1000; Cell Signaling Technology), Akt (total, 1:1000; Cell Signaling Technology), phospho-Akt (Thr308, 1:1000; Cell Signaling Technology), Bad (total, 1:500; BBI Life Sciences), phospho-Bad (Ser112, 1:500; BBI Life Sciences), or β-actin (1:20,000, Sigma-Aldrich).

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: EEF1D overexpression promotes osteosarcoma cell proliferation by facilitating Akt-mTOR and Akt-bad signaling

doi: 10.1186/s13046-018-0715-5

Figure Lengend Snippet: EEF1D Knockdown inhibits osteosarcoma cell growth in vitro. a - f Expression levels of EEF1D mRNA and protein in MNNG/HOS, U2OS and MG63 cells were measured after EEF1D siRNA (si-EEF1D) transfection by qRT-PCR and western blotting, respectively. g - i Cell Counting Kit-8 (CCK-8) assays were performed to measure cell proliferation after siRNA transfection. j - o Colony-formation assays were performed for EEF1D-silenced osteosarcoma and control cells. Data are representative of results from three independent experiments. * P < 0.05. For qRT-PCR, EEF1D expression was normalized to β-actin

Article Snippet: After blocking in 5% nonfat milk, the membranes were incubated with the following primary antibodies: EEF1D (1:500, Proteintech) [ ], mTOR (total, 1:1000; Cell Signaling Technology), phospho-mTOR (Ser2448, 1:1000; Cell Signaling Technology), Akt (total, 1:1000; Cell Signaling Technology), phospho-Akt (Thr308, 1:1000; Cell Signaling Technology), Bad (total, 1:500; BBI Life Sciences), phospho-Bad (Ser112, 1:500; BBI Life Sciences), or β-actin (1:20,000, Sigma-Aldrich).

Techniques: Knockdown, In Vitro, Expressing, Transfection, Quantitative RT-PCR, Western Blot, Cell Counting, CCK-8 Assay, Control

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: EEF1D overexpression promotes osteosarcoma cell proliferation by facilitating Akt-mTOR and Akt-bad signaling

doi: 10.1186/s13046-018-0715-5

Figure Lengend Snippet: EEF1D Knockdown inhibits osteosarcoma cell cycle G2/M transition. Representative images of the cell cycle assays in MNNG/HOS ( a , b ), U2OS ( d , e ) and MG63 cells ( g , h ) after transfection with nonspecific control siRNA (si-NC) or EEF1D siRNA (si-EEF1D). c , f , i Diagrams showing the results of cell cycle assay in MNNG/HOS, U2OS and MG63 cells

Article Snippet: After blocking in 5% nonfat milk, the membranes were incubated with the following primary antibodies: EEF1D (1:500, Proteintech) [ ], mTOR (total, 1:1000; Cell Signaling Technology), phospho-mTOR (Ser2448, 1:1000; Cell Signaling Technology), Akt (total, 1:1000; Cell Signaling Technology), phospho-Akt (Thr308, 1:1000; Cell Signaling Technology), Bad (total, 1:500; BBI Life Sciences), phospho-Bad (Ser112, 1:500; BBI Life Sciences), or β-actin (1:20,000, Sigma-Aldrich).

Techniques: Knockdown, Transfection, Control, Cell Cycle Assay

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: EEF1D overexpression promotes osteosarcoma cell proliferation by facilitating Akt-mTOR and Akt-bad signaling

doi: 10.1186/s13046-018-0715-5

Figure Lengend Snippet: EEF1D Knockdown inhibits Akt-mTOR and Akt-Bad signaling pathways in osteosarcoma cells. a , b MNNG/HOS and U2OS cell extracts were prepared and analyzed using PathScan® intracellular signaling array kit. Images were captured with Odyssey® Infrared Imaging System (LI-COR). c , d Quantification of results from MNNG/HOS and U2OS cells shown in ( a , b ) e , f Western blotting analysis of Akt-mTOR and Akt-Bad signaling pathway molecules in MNNG/HOS and U2OS cells transfected with nonspecific control siRNA (si-NC) or EEF1D siRNA (si-EEF1D). g Western blotting analysis of Akt-mTOR and Akt-Bad signaling pathway molecules, including mTOR, Akt, and Bad, in hFOB 1.19 cells transfected with pcDNA 3.1 or pcDNA 3.1-EEF1D. Quantification of results in ( e , f , g ) were shown in ( h , i and j ) respectively

Article Snippet: After blocking in 5% nonfat milk, the membranes were incubated with the following primary antibodies: EEF1D (1:500, Proteintech) [ ], mTOR (total, 1:1000; Cell Signaling Technology), phospho-mTOR (Ser2448, 1:1000; Cell Signaling Technology), Akt (total, 1:1000; Cell Signaling Technology), phospho-Akt (Thr308, 1:1000; Cell Signaling Technology), Bad (total, 1:500; BBI Life Sciences), phospho-Bad (Ser112, 1:500; BBI Life Sciences), or β-actin (1:20,000, Sigma-Aldrich).

Techniques: Knockdown, Protein-Protein interactions, Imaging, Western Blot, Transfection, Control

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: EEF1D overexpression promotes osteosarcoma cell proliferation by facilitating Akt-mTOR and Akt-bad signaling

doi: 10.1186/s13046-018-0715-5

Figure Lengend Snippet: Clinical significance of EEF1D in osteosarcoma patients. a Representative IHC image of EEF1D in non-tumor tissues. b Representative IHC image of EEF1D in osteosarcoma. Original magnification: 50×, 200×

Article Snippet: After blocking in 5% nonfat milk, the membranes were incubated with the following primary antibodies: EEF1D (1:500, Proteintech) [ ], mTOR (total, 1:1000; Cell Signaling Technology), phospho-mTOR (Ser2448, 1:1000; Cell Signaling Technology), Akt (total, 1:1000; Cell Signaling Technology), phospho-Akt (Thr308, 1:1000; Cell Signaling Technology), Bad (total, 1:500; BBI Life Sciences), phospho-Bad (Ser112, 1:500; BBI Life Sciences), or β-actin (1:20,000, Sigma-Aldrich).

Techniques:

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: EEF1D overexpression promotes osteosarcoma cell proliferation by facilitating Akt-mTOR and Akt-bad signaling

doi: 10.1186/s13046-018-0715-5

Figure Lengend Snippet: Correlation analyses of EEF1D protein expression in relation to clinicopathologic variables of 50 patients with osteosarcoma

Article Snippet: After blocking in 5% nonfat milk, the membranes were incubated with the following primary antibodies: EEF1D (1:500, Proteintech) [ ], mTOR (total, 1:1000; Cell Signaling Technology), phospho-mTOR (Ser2448, 1:1000; Cell Signaling Technology), Akt (total, 1:1000; Cell Signaling Technology), phospho-Akt (Thr308, 1:1000; Cell Signaling Technology), Bad (total, 1:500; BBI Life Sciences), phospho-Bad (Ser112, 1:500; BBI Life Sciences), or β-actin (1:20,000, Sigma-Aldrich).

Techniques: Expressing