Journal: Cell death and differentiation

Article Title: Glial cell missing-1 transcription factor is required for the differentiation of the human trophoblast.

doi: 10.1038/cdd.2009.1

Figure Lengend Snippet: Figure 1 Cell membrane integrity of BeWo cells was monitored with anti-human b-catenin immunofluorescence. (a) Immunofluorescent image of BeWo cells undergoing low spontaneous level of fusion (* indicates 3 or more DAPI-stained nuclei within an intact membrane was considered as fusion event). (b) Forskolin-induced excessive cell– cell fusion resulted in membrane dissolution and loss of distinct membrane staining. (c) Silenced BeWo cells and (d) silenced BeWo cells treated with forskolin show no cell– cell fusion shown by intact membrane staining. Scale bar ¼ 100 mm. (e) Relative GCM1 mRNA expression (n ¼ 6) and (f) representative western blot analysis of GCM1 protein expression (n ¼ 6) in BeWo cells stably expressing siRNA to GCM1 in cells treated with 25 mM of forskolin and in cells stably expressing siRNA þ forskolin compared with non-silencing (sense control) siRNA-transfected control BeWo cells. Forskolin treatment of BeWo cells significantly upregulated GCM1 RNA and protein expression. GCM1 silencing eliminated this effect. Bar graph showing the mean±s.e. (n ¼ 6). NS, non-silenced control; F, forskolin (25 mM). (g) The two-color fusion assay shows a significant decrease of de novo fusion in the silenced BeWo group with forskolin treatment compared with that in control and forskolin-treated non-silenced BeWo cells. Difference between the non-forskolin-treated sense control group and stably expressing GCM1 siRNA group is not significant because of the low level of basal fusion (n ¼ 6). (h) A proliferation assay of control BeWo cells, overexpressing GCM1 (25 mM forskolin treatment) and BeWo cells stably expressing siRNA to GCM1 over 3 days in culture. On the third day of culture, cells overexpressing GCM1 show significant reduction in the number of cells, whereas BeWo cells stably expressing GCM1; siRNA show significant increase in cell numbers as compared with control cells (n ¼ 3). Mock controls showed no difference to non-silenced controls in all experiments (data not shown). *Po0.005, **Po0.001

Article Snippet: Membranes were blocked with 5% skimmed milk in 0.1% (v/v) Tween Tris-buffer saline (TBST) for 2 h at room temperature and incubated with anti-GCM1 antibody (1:3000) at 41C overnight (Aviva System Biology, San Diego, CA, USA).

Techniques: Membrane, Staining, Dissolution, Expressing, Western Blot, Stable Transfection, Control, Transfection, Single Vesicle Fusion Assay, Proliferation Assay

Journal: Cell death and differentiation

Article Title: Glial cell missing-1 transcription factor is required for the differentiation of the human trophoblast.

doi: 10.1038/cdd.2009.1

Figure Lengend Snippet: Figure 3 GCM1 antisense oligonucleotides and siRNA treatment after trypsinization of SCT result in an accumulation of 4.9±1.5 layers of CT cells (semi-thin section, a) that proliferate (Ki-67 immunopositive in brown, d) but do not regenerate a new layer of SCT. (b) Treatment of non-denuded first trimester explants with GCM1 antisense oligonucleotide or siRNA results in the accumulation of 3.8±0.4 layers of cytotrophoblast cells underneath a necrotic SCT layer (note the dissolution of syncytial nuclei, r). (e) Accumulating V-CT exhibited a high level of proliferation (Ki-67 positive). (c) Forskolin treatment results in the accumulation of multilayer of syncytial nuclei, but reduced the numbers of V-CT cells (0.9±0.3 layers) and (f) the rate of Ki-67 immunopositivity in the trophoblast layer. Scale bars ¼ 1000 mm (a–c), 50 mm (d–f). (g) Western blot analysis of GCM1 protein levels in siRNA or antisense oligonucleotide-treated explants. (h) Forskolin treatment of first trimester floating villous explants regulates GCM1 protein levels in time-dependent manner. White bars ¼ controls, black bars ¼ forskolin-treated explants. Significant difference from control explants is indicated by *Po0.05

Article Snippet: Membranes were blocked with 5% skimmed milk in 0.1% (v/v) Tween Tris-buffer saline (TBST) for 2 h at room temperature and incubated with anti-GCM1 antibody (1:3000) at 41C overnight (Aviva System Biology, San Diego, CA, USA).

Techniques: Dissolution, Western Blot, Control

Journal: Cell death and differentiation

Article Title: Glial cell missing-1 transcription factor is required for the differentiation of the human trophoblast.

doi: 10.1038/cdd.2009.1

Figure Lengend Snippet: Figure 4 Histological section of (a) control, (b) GCM1-silenced, and (c) forskolin-treated floating villous placental explants stained for BrdU (blue nuclei). The index of proliferation of villous trophoblast cells in explants cultured with (d) GCM1 siRNA, (e) the GCM1 antisense oligonucleotide and (f) forskolin. Scale bars ¼ 40 mm (a, b); 50 mm (c). n ¼ 9; *Po0.05

Article Snippet: Membranes were blocked with 5% skimmed milk in 0.1% (v/v) Tween Tris-buffer saline (TBST) for 2 h at room temperature and incubated with anti-GCM1 antibody (1:3000) at 41C overnight (Aviva System Biology, San Diego, CA, USA).

Techniques: Control, Staining, Cell Culture

Journal: Cell death and differentiation

Article Title: Glial cell missing-1 transcription factor is required for the differentiation of the human trophoblast.

doi: 10.1038/cdd.2009.1

Figure Lengend Snippet: Figure 5 Dissecting light microscope image of (a) sense oligonucleotide-treated first trimester explant cultured on Matrigel. Typical organized extravillous trophoblast outgrowth formation; eight outgrowth fingers are numbered. (b) H&E staining of control explants. (c) The outgrowths are a1-Integrin-positive as well as (d) GCM1- immunopositive. (e) GCM1 antisense oligonucleotide-treated explants do not form extravillous outgrowths, (f) cells do not invade Matrigel (H&E staining). (g) Cells are negative for the invasive marker a1-Integrin. (h) Immunofluorescent staining to GCM1 protein. Scale bars ¼ 100 mm (a, e); 50 mm (b, f); 25 mm (c, d, g, h)

Article Snippet: Membranes were blocked with 5% skimmed milk in 0.1% (v/v) Tween Tris-buffer saline (TBST) for 2 h at room temperature and incubated with anti-GCM1 antibody (1:3000) at 41C overnight (Aviva System Biology, San Diego, CA, USA).

Techniques: Light Microscopy, Cell Culture, Staining, Control, Marker

Journal: Polymers

Article Title: Mechanical and Thermal Behaviours of Weft-Knitted Spacer Fabric Structure with Inlays for Insole Applications

doi: 10.3390/polym14030619

Figure Lengend Snippet: Summary of test methods.

Article Snippet: Surface unevenness , 3D-optical microscope (VR-3000, KEYENCE, Osaka, Japan) , ISO4287:1997 Surface unevenness-Definitions.

Techniques: Microscopy, Permeability

Journal: Cell Host & Microbe

Article Title: Lassa virus glycoprotein nanoparticles elicit neutralizing antibody responses and protection

doi: 10.1016/j.chom.2022.10.018

Figure Lengend Snippet:

Article Snippet: Four-fold serial dilutions of mAbs diluted to 2.5 μg/mL in Casein were then added for 2 h. A 1:3000 dilution of HRP-labeled goat anti-rabbit IgG (Jackson Immunoresearch) in Casein was added for 1 h. Up to now, between each step, plates were washed three times with TBS.

Techniques: Virus, Recombinant, Mass Spectrometry, Sequencing, Lysis, Emulsion, Adjuvant, Plasmid Preparation, High Molecular Weight, Marker, Electron Microscopy, Luciferase, Software, Chromatography, Random Hexamer, Luminex, Expressing, Modification, Nano Differential Scanning Fluorimetry, Clinical Proteomics