Journal: World Journal of Surgical Oncology

Article Title: LAIR-1 overexpression inhibits epithelial–mesenchymal transition in osteosarcoma via GLUT1-related energy metabolism

doi: 10.1186/s12957-020-01896-7

Figure Lengend Snippet: LAIR-1 inhibits Glut1-related glucose uptake in OS cells. a Heatmap showing the levels of differentially expressed mRNAs. b Top 20 KEGG pathway annotation categories for target gene functions of predicted mRNAs. c Selected significantly differentially expressed mRNA-related to EMT in RNA-seq data between two groups, *** P < 0.001. d qPCR validation of differentially expressed EMT-related genes in LV-NC and LV-LAIR-1-overexpressing OS cells, ** P < 0.01. e Glut1 expression analyzed by western blotting. f Immunofluorescence staining of Glut1 in the LV-LAIR-1-overexpressing OS cells. Scale bar = 50 μm. Data were obtained from at least two independent experiments.

Article Snippet: Total protein was extracted using a routine procedure and blotted with the following primary antibodies: LAIR-1 (sc-398141; Santa Cruz Biotechnology), phospho-Foxo1 (Ser256) (84192; Cell Signaling Technology, Danvers, MA, USA), Foxo1 (2880; Cell Signaling Technology), phospho-Akt (Ser473) (AF8355; Affinity Biosciences, Cincinnati, OH, USA), Akt (9272; Cell Signaling Technology), proliferating cell nuclear antigen (PCNA; BM0104; Boster Biotech Co., Ltd., Wuhan, China), Twist1 (ab50581; Abcam, Cambridge, UK), Glut1 (NB110-39113, Novus Biologicals, Littleton, CO, USA), and β-actin (30101ES50; Yeasen Biotech Co., Ltd., Shanghai, China).

Techniques: RNA Sequencing, Biomarker Discovery, Expressing, Western Blot, Immunofluorescence, Staining

Journal: Viruses

Article Title: Cytolytic Recombinant Vesicular Stomatitis Viruses Expressing STLV-1 Receptor Specifically Eliminate STLV-1 Env-Expressing Cells in an HTLV-1 Surrogate Model In Vitro

doi: 10.3390/v14040740

Figure Lengend Snippet: Amino acid sequence identity between human GLUT1 and Japanese macaque GLUT1.

Article Snippet: At 1 day post-infection, the cells were stained with mouse monoclonal anti-Human GLUT1 antibody (Ab) (R&D Systems, Minneapolis, MN, USA), followed by incubation with a FITC-conjugated goat anti-mouse IgG (H + L) Ab (Jackson ImmunoResearch, West Grove, PA, USA) in accordance with the manufacturer’s recommendations.

Techniques: Sequencing

Journal: Viruses

Article Title: Cytolytic Recombinant Vesicular Stomatitis Viruses Expressing STLV-1 Receptor Specifically Eliminate STLV-1 Env-Expressing Cells in an HTLV-1 Surrogate Model In Vitro

doi: 10.3390/v14040740

Figure Lengend Snippet: Schematic showing the structure of rVSVs expressing an STLV-1 receptor molecule and AcGFP1 as a reporter protein. The gene orders in the wildtype VSV and the rVSV constructs used in this study are illustrated. The deleted G gene was replaced with the JM GLUT1 or AcGFP1 genes, yielding the VSVΔG-JmGL or VSVΔG-AcGFP constructs, respectively. In addition, a JM GLUT1 gene was inserted upstream of the AcGFP1 gene in VSVΔG-AcGFP, yielding VSVΔG-JmGL-AcGFP.

Article Snippet: At 1 day post-infection, the cells were stained with mouse monoclonal anti-Human GLUT1 antibody (Ab) (R&D Systems, Minneapolis, MN, USA), followed by incubation with a FITC-conjugated goat anti-mouse IgG (H + L) Ab (Jackson ImmunoResearch, West Grove, PA, USA) in accordance with the manufacturer’s recommendations.

Techniques: Expressing, Construct

Journal: Viruses

Article Title: Cytolytic Recombinant Vesicular Stomatitis Viruses Expressing STLV-1 Receptor Specifically Eliminate STLV-1 Env-Expressing Cells in an HTLV-1 Surrogate Model In Vitro

doi: 10.3390/v14040740

Figure Lengend Snippet: Expression of the STLV-1 receptor molecule on the rVSV-infected target cell surface. The surface expression of JM GLUT1 protein after VSV infection was confirmed by immunofluorescence staining. VSV-permissive BHK-21 cells were infected with G-complemented VSVΔG-JmGL at an MOI of 0.1. After 1 day of culture, JM GLUT1 expressed by the viral genome was stained with anti-GLUT1-specific mouse monoclonal antibody (MAb) followed by FITC-conjugated anti-mouse IgG. The stained cells were observed by fluorescence microscopy and photographed at a constant magnification. Scale bars in panels represent 400 µm.

Article Snippet: At 1 day post-infection, the cells were stained with mouse monoclonal anti-Human GLUT1 antibody (Ab) (R&D Systems, Minneapolis, MN, USA), followed by incubation with a FITC-conjugated goat anti-mouse IgG (H + L) Ab (Jackson ImmunoResearch, West Grove, PA, USA) in accordance with the manufacturer’s recommendations.

Techniques: Expressing, Infection, Immunofluorescence, Staining, Fluorescence, Microscopy

Journal: Viruses

Article Title: Cytolytic Recombinant Vesicular Stomatitis Viruses Expressing STLV-1 Receptor Specifically Eliminate STLV-1 Env-Expressing Cells in an HTLV-1 Surrogate Model In Vitro

doi: 10.3390/v14040740

Figure Lengend Snippet: Functional assessment of infectivity and specificity of rVSVs. ( A , B ) STLV-1 Env-dependent infectivity of rVSVs was evaluated. BHK-21 cells were initially mock-transfected (empty vector) or transfected with expression plasmids for STLV-1 Env derived from Si-2 cells or from STLV-1-infected JM PBMCs. After 24 h, transfected BHK-21 cells were infected with non-G-complemented rVSVs (0.25 mL each). After 3 days of culture, the cells were fixed. Subsequently, VSVΔG-JmGL-infected cells were stained with anti-GLUT1-specific MAb to detect rVSV-infected cells. On the other hand, VSVΔG-JmGL-AcGFP- or VSVΔG-AcGFP-infected cells were detected by GFP expression. Representative results from each group of rVSV-infected BHK-21 cells are shown in panel ( A ). Areas enclosed with squares are enlarged in panel ( B ). Syncytia with enlarged cell size in VSVΔG-JmGL- or VSVΔG-JmGL-AcGFP-infected cells are shown in panel ( B ). Scale bars in panels ( A , B ) represent 400 µm and 200 µm, respectively. ( C ) STLV-1 Env-dependent rVSV infection was assessed by neutralizing assay. BHK-21 cells were initially transfected with the Si-2 strain STLV-1 Env expression plasmid. After 24 h, transfected BHK-21 cells were infected with G-complemented VSVΔG-AcGFP (MOI of 0.01) or non-G-complemented rVSVs (0.25 mL each), in the absence or presence of HTLV-1 envelope glycoprotein-specific neutralizing antibody (LAT-27; 10 µg/mL). The cells infected with VSVΔG-JmGL or VSVΔG-huGL were stained and examined by VSV N-specific immunofluorescence to detect the rVSV-infected cells at 48 h post-infection. The culture of each virus without LAT-27 was defined as a control. The total number of fluorescent cells per well was determined, and the relative infectivity of each virus was calculated. The data are expressed as a percentage of the control (mean ± SD) from four independent experiments. ( D ) Viral growth kinetic assay results are shown. Si-2 strain STLV-1 Env-transfected BHK-21 cells were infected with non-G-complemented VSVΔG-AcGFP (square) or VSVΔG-JmGL-AcGFP (circle) (0.25 mL each). The total number of GFP-positive cells per well was counted to determine the quantitative viral titer (in infectious units per milliliter). The data are expressed as the mean ± SD from four independent experiments. Asterisks in panel ( C ) represent significant differences versus control (** p < 0.001 significance, by two-tailed Student’s t -test with equal variance).

Article Snippet: At 1 day post-infection, the cells were stained with mouse monoclonal anti-Human GLUT1 antibody (Ab) (R&D Systems, Minneapolis, MN, USA), followed by incubation with a FITC-conjugated goat anti-mouse IgG (H + L) Ab (Jackson ImmunoResearch, West Grove, PA, USA) in accordance with the manufacturer’s recommendations.

Techniques: Functional Assay, Infection, Transfection, Plasmid Preparation, Expressing, Derivative Assay, Staining, Neutralizing Assay, Immunofluorescence, Virus, Control, Kinetic Assay, Two Tailed Test