Journal: Acta Neuropathologica

Article Title: Analyzing microglial phenotypes across neuropathologies: a practical guide

doi: 10.1007/s00401-021-02370-8

Figure Lengend Snippet: Commonly used immunomarkers for the characterization of different microglial states in the healthy and diseased murine and human CNS are shown

Article Snippet: SLC2A5 , Glucose transporter type 5 labelling parenchymal myeloid cells , IHC , Abcam, Atlas Antibodies, HPA005449, 1:250.

Techniques: Flow Cytometry, Isolation, Marker, Activation Assay, Activity Assay

Journal: Frontiers in Cell and Developmental Biology

Article Title: Loss of the fructose transporter SLC2A5 inhibits cancer cell migration

doi: 10.3389/fcell.2022.896297

Figure Lengend Snippet: Attenuation of the SLC2A5 gene expression in fibrosarcoma cells. (A) . Images of scratch test (left) and quantitative analysis of closure (right) as a function of time for HT1080tdT and HT1080tdT-ε2A5 fibrosarcoma cells (n = 3). Scale bar 10 µm. (B) . Images of colonies formed by HT1080tdT and HT1080tdT-ε2A5 cells and analysis of colony number (500 cells/well plated at time 0 h). * p = 0.007; n = 3. Colonies were fixed with 70% ethanol and visualized by staining with 1% trypan blue. (C) . Proliferation assay of HT1080tdT and HT1080tdT-ε2A5 cells. HT1080tdT and HT1080tdT-ε2A5 cells were plated at a density of 5.0 × 10 3 cells/well of a 96 well plates. Fructose was added to the media as indicated in the Figure. Cells were cultured for 48 h followed by MTS assay (n = 3). All data in the Figure is representative of more than 3 experiments with 3 biological replicates.

Article Snippet: Anti-SLC2A5 antibodies were purchased from Boster (Cat# PA 2064) and used at 1:500 dilution.

Techniques: Gene Expression, Staining, Proliferation Assay, Cell Culture, MTS Assay

Journal: Frontiers in Cell and Developmental Biology

Article Title: Loss of the fructose transporter SLC2A5 inhibits cancer cell migration

doi: 10.3389/fcell.2022.896297

Figure Lengend Snippet: Editing of the SLC2A5 gene. (A) . Inactivation of the SLC2A5 gene in human MIA-PaCa-2 and HT1080tdT cells using CRISPR/Cas9-mediated in situ gene editing. Guide RNAs were targeted to Exon 3. Genomic DNA of edited cells was sequenced, and positive clones were identified as having a 62 base pair disruption, leading to an early stop codon 14 amino acids downstream of the disruption point. (B) . qPCR analysis of SLC2A5 mRNA abundance in MIA-PaCa-2 and MIA-PaCa-ε2A5 cells (clones B3 and F11) (n = 3). DNA primers were designed to amplify Exon 12 of the SLC2A5 gene. Immunoblot analysis was carried out with antibodies directed at the C-terminus of SLC2A5. Anti-β-tubulin or anti-GAPDH antibodies were used to assess amounts of loaded protein samples. (C) . qPCR analysis of SLC2A5 mRNA abundance in HT1080tdT and HT1080tdT-ε2A5 cells. DNA primers and control antibodies were as described above. (D) . qPCR analyses of SLC2A1, SLC2A2, and SLC2A4 mRNA abundance in MIA-PaCa-2 and MIA-PaCa-ε2A5 cells (n = 3). (E) . qPCR analyses of SLC2A1, SLC2A2, and SLC2A4 mRNA abundance in HT1080tdT and HT1080tdT-ε2A5 cells (n = 3). All data in the Figure is representative of more than 3 biological replicates. NS, not significant.

Article Snippet: Anti-SLC2A5 antibodies were purchased from Boster (Cat# PA 2064) and used at 1:500 dilution.

Techniques: CRISPR, In Situ, Clone Assay, Disruption, Western Blot, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Loss of the fructose transporter SLC2A5 inhibits cancer cell migration

doi: 10.3389/fcell.2022.896297

Figure Lengend Snippet: Attenuation of the SLC2A5 gene expression in pancreatic cancer cells. (A) . Images of scratch test (left panel) and quantitative analysis of closure (right panel) for MIA-PaCa-2 and MIA-PaCa-ε2A5 cells (clone B3; ** p = 0.002, n = 3). Scale bar 10 µm. (B) . Scratch test analysis for MIA-PaCa-2 and MIA-PaCa-ε2A5 cell lines clone F11 (** p = 0.001, n = 3). Scale bar 10 µm. (C) . Colony formation images and colony number for MIA-PaCa-ε2A5 cells, clone B3 (left panel; 1000 cells/well plated at time 0 h; * p = 0.0001; n = 3) and clone F11 (right panel; 500 cells/well plated at time 0 h; * p = 0.001; n = 3). (D) . Fructose-dependent proliferation of the MIA-PaCa-2 and MIA-PaCa-ε2A5 cells. MIA-PaCa-2 and MIA-PaCa-ε2A5 (clones B3 and F11) cells were plated at a density of 1.0 × 10 4 cells/well of a 96 well plates. Fructose was added to the culture medium as indicated in the figure. The absorbance at 490 nm (OD 490 ) of the culture medium after MTS assay at time 0 and 48 h in culture are shown. Experiments were performed a minimum of three times in triplicates. * p = 0.0002; n = 3.

Article Snippet: Anti-SLC2A5 antibodies were purchased from Boster (Cat# PA 2064) and used at 1:500 dilution.

Techniques: Gene Expression, Clone Assay, MTS Assay

Journal: Frontiers in Cell and Developmental Biology

Article Title: Loss of the fructose transporter SLC2A5 inhibits cancer cell migration

doi: 10.3389/fcell.2022.896297

Figure Lengend Snippet: Restoration of SLC2A5 function via SLC2A5 transgene expression. Proliferation assay (A) and scratch test assay (B) of MIA-PaCa-2, MIA-PaCa-ε2A5 (clone F11) and MIA-PaCa-ε2A5 cells expressing wild-type SLC2A5 or the non-functional SLC2A5-E401A mutant (* p = 0.002; n = 3). In (A) , fold change in cell number is shown at day 0 to day 3. In (B) , the percentage of open scratch is shown at time 0 and 48 h. Proliferation assay (C) and scratch test assay (D) of HT1080tdT, HT1080tdT-ε2A5 cells transfected with expression vectors encoding wild-type SLC2A5 or the non-functional SLC2A5-E401A mutant (* p = 0.001, n = 3). In (C) , the absorbance at 490 nm (OD 490 ) of the culture medium after MTS assay from time 0 and 7 h in culture is shown. In (D) , the percentage of open scratch is shown at time 0 and 7 h. NS, not significant. All data in the Figure is representative of more than 3 experiments with 3 replicates.

Article Snippet: Anti-SLC2A5 antibodies were purchased from Boster (Cat# PA 2064) and used at 1:500 dilution.

Techniques: Expressing, Proliferation Assay, Wound Healing Assay, Functional Assay, Mutagenesis, Transfection, MTS Assay

Journal: Frontiers in Cell and Developmental Biology

Article Title: Loss of the fructose transporter SLC2A5 inhibits cancer cell migration

doi: 10.3389/fcell.2022.896297

Figure Lengend Snippet: Mitochondria localization and dynamics in SLC2A5-deficient cancer cells. (A) . Electron microscopy analysis of MIA-PaCa-2 cells shows that mitochondria were localized in the center of these cells in the perinuclear space (encircled). In MIA-PaCa-ε2A5 cells, mitochondria were spread throughout the cell towards the cell periphery, and appeared elongated (arrows) (n = 3). (B) . and (C) . Mitochondria area (B) and length (C) analyses in MIA-PaCa-2 and MIA-PaCa-ε2A5 cells. * p < 0.0006; ** p < 0.0001 (n = 3). (D) . Confocal images of mitochondria stained with MitoTracker ® Green in red fluorescent protein-labelled HT1080tdT cells,HT1080tdT-ε2A5 cells or HT1080tdT-ε2A5 cells expressing wild-type SLC2A5. Time-lapse video is shown in the (arrowhead). (E) . Mitochondria number and (F) . mitochondria length in fibrosarcoma HT1080tdT cells, HT1080tdT-ε2A5 or HT1080tdT-ε2A5 cells expressing wild-type SLC2A5. *** p < 0.0002 (n = 3).

Article Snippet: Anti-SLC2A5 antibodies were purchased from Boster (Cat# PA 2064) and used at 1:500 dilution.

Techniques: Electron Microscopy, Staining, Expressing

Journal: Frontiers in Cell and Developmental Biology

Article Title: Loss of the fructose transporter SLC2A5 inhibits cancer cell migration

doi: 10.3389/fcell.2022.896297

Figure Lengend Snippet: SLC2A5 function is necessary for polarized localization of mitochondria in HT1080tdT human fibrosarcoma cells in vitro and in vivo . (A) mitochondria (green) localization in HT1080tdT or HT1080tdT-ε2A5 (vasculature, grey; blue, DAPI) cells that are extravasating out of the chicken CAM vasculature ( in vivo ). Right panels show all three channels: red fluorescent protein (red), mitochondria (green), nuclei (blue) and vasculature (grey). Time-lapse video is shown in the . (B) . Fluorescent channel intensity along the line scans (dashed arrows in (A) (right panel) indicate the direction of the scan). Green line depicts mitochondria localization. The short arrows in (A) and (B) indicate vascular membrane breaches.

Article Snippet: Anti-SLC2A5 antibodies were purchased from Boster (Cat# PA 2064) and used at 1:500 dilution.

Techniques: In Vitro, In Vivo, Membrane