Journal: eLife

Article Title: Beta-catenin signaling regulates barrier-specific gene expression in circumventricular organ and ocular vasculatures

doi: 10.7554/eLife.43257

Figure Lengend Snippet:

Article Snippet: The following antibodies were used for tissue immunohistochemistry: rabbit anti-GLUT1 (Thermo Fisher Scientific RB-9052-P1); rat anti-mouse PLVAP/MECA-32 (BD Biosciences 553849); mouse anti-CLDN5, Alexa Fluor 488 conjugate (Thermo Fisher Scientific 352588); rabbit anti-GFP, Alexa Fluor 488 conjugate (Thermo Fisher Scientific A21311); rabbit anti-6xMyc (JH6204), rabbit mAb anti-LEF1 (Cell Signaling Technologies C12A5), rabbit anti-ZO-1 (Invitrogen 40–2200), rabbit anti-Occludin (Invitrogen 406100), rabbit mAb anti-MDR1 (E1Y7S; Cell Signaling Technology 13978S), chicken anti-Vimentin (EMD Millipore Corp AB5733), and rabbit anti-MFSD2A (a kind gift of David Silver, Duke-NUS Medical School).

Techniques: Recombinant, Software, Plasmid Preparation

Journal: Advanced science (Weinheim, Baden-Wurttemberg, Germany)

Article Title: Identification and Characterization of the Wilms Tumor Cancer Stem Cell.

doi: 10.1002/advs.202206787

Figure Lengend Snippet: Figure 3. SIX2+CITED1+ cell self-renewal: the role of the extracellular matrix niche. A,B) Representative immunofluorescence staining showing the distribution of ITG𝛽1 (green) and SIX2 (red) in hFK (10 WGA) and WT (WT#12, favorable stage III) A) and for ITG𝛽1 (red) and CITED1 (green) in hFK (10 WGA) and WT (WT#8 favorable stage II. B) Nuclei stained with DAPI (blue); scale bars 50 and 75 μm, respectively. C) Heatmap showing gene expression profile for integrins in SIX2+CITED1+ cells from hFK (17, 17.2, and 17.5 WGA) and WT (WT#3 anaplastic stage I, WT#4: non-anaplastic, stage III, and WT#5: non-anaplastic chemo-treated, stage IV). D) Densitometric analysis by western blot (WB) of ITG𝛽1 expression in freshly isolated SIX2+CITED1+ cells from WT (WT#8,11,12, favorable stage II, favorable stage III, and favorable stage II) versus SIX2+CITED1+ cells from hFK (15,16,18 WGA) showing higher expression of ITG𝛽1 in hFK cells; 𝛽-actin was used as housekeeping protein for normalization. WB bands are presented below the graph, *p < 0.05. E) Representative immunofluorescence staining of SIX2 (red) and CITED1 (green) in SIX2+CITED1+ cells from hFK (17 WGA) cultured for 72 h with/without 1 μg mL−1 anti-ITG𝛽1 or 0.5 μg mL−1 anti-ITG𝛽4 neutralizing antibody showing increased expression of CITED1 in cells treated with anti-ITG𝛽1. Nuclei stained with DAPI (blue). Scale bar = 50 μm. F,G) Percentage of SIX2+CITED1+ cells and total SIX2+ cells from hFK (17 WGA) by flow cytometry analysis after F) 5 d or G) 28 d of culture with/without anti-ITG𝛽1 or anti-ITG𝛽4 neutralizing antibody or a combination of both. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001; mean ± SEM. H) Densitometric analysis by WB of LEF1 protein expression in hFK SIX2+CITED1+ cells (17 WGA) cultured for 28 d with/without anti-ITG𝛽1 or anti-ITG𝛽4 neutralizing antibody 𝛽-actin was used as housekeeping control. WB bands are presented below the graph. *p < 0.05. I) Kaplan-Meier survival analysis of mice injected with WT SIX2+CITED1+ cells, without treatment (control, n = 4) or with the treatment of anti-ITG𝛽1 (n = 5) or anti-ITG𝛽4 (n = 4), endpoint tumor size 1.5 cm. J) Schematic representation showing the proposed role of ITG𝛽1 and ITG𝛽4 in WT SIX2+CITED1+ cells.

Article Snippet: #9315 1:1000 WB Cyclin D1 Abcam # 16663 1:1000 WB LEF1 (C12A5) Cell Signaling Tech.

Techniques: Staining, Gene Expression, Western Blot, Expressing, Isolation, Cell Culture, Cytometry, Control, Injection

Journal: Cell Proliferation

Article Title: METTL3 ‐m 6 A methylase regulates the osteogenic potential of bone marrow mesenchymal stem cells in osteoporotic rats via the Wnt signalling pathway

doi: 10.1111/cpr.13234

Figure Lengend Snippet: Expression of m 6 A methylation level and Wnt signalling pathway were downregulated in OP‐BMSCs. (A) The mRNA methylation level of OP‐BMSCs was lower than that of BMSCs, methylene blue staining (MB) was used as a loading control. The gene expression of Mettl3 , Mettl14 , Wtap , Fto and Ythdf1 was lower in OP‐BMSCs compared to BMSCs. (B) Gene and protein expression of Mettl3, Runx2, Opn, Lef1, P‐Gsk‐3β and β‐Catenin were decreased after osteogenic induction for 3 days in OP‐BMSCs. (C) Gene and protein expression of Mettl3, Runx2, Opn, Lef1, P‐Gsk‐3β and β‐Catenin were also decreased after osteogenic induction for 5 days in OP‐BMSCs. Data represent the mean ± SD ( n ≥ 3). (* p < 0.05,** p < 0.01, *** p < 0.001)

Article Snippet: The proteins were migrated to polyvinylidene difluoride (PVDF) membranes and combined with specific antibodies to GAPDH (ab181602), OPN (ab8448), RUNX2 (ab92336), β‐CATENIN (ab32572), METTL3 (ab195352) (all Abcam), LEF1 (C12A5) GSK (9832S) and P‐GSK (9323S) (Cell Signalling Technology).

Techniques: Expressing, Methylation, Staining, Control, Gene Expression

Journal: Cell Proliferation

Article Title: METTL3 ‐m 6 A methylase regulates the osteogenic potential of bone marrow mesenchymal stem cells in osteoporotic rats via the Wnt signalling pathway

doi: 10.1111/cpr.13234

Figure Lengend Snippet: Overexpression of Mettl3 improved the osteogenic ability of OP‐BMSCs by activating the canonical Wnt signalling pathway in vitro . (A) Mettl3 overexpression lentivirus transfected into OP‐BMSCs for 72 h with an MOI value of 60. (B) Relative gene and protein expression of Mettl3, Runx2, Opn, Lef1, P‐Gsk‐3β and β‐Catenin were increased after Mettl3 overexpressing in OP‐BMSCs for osteogenic induction of 3 days. (C) After Mettl3 was overexpressed in OP‐BMSCs, the relative gene and protein expression of Mettl3, Runx2, Opn, Lef1, P‐Gsk‐3β and β‐Catenin were increased after osteogenic induction for 5 days. Data represent the mean ± SD ( n ≥ 3). (* p < 0.05,** p < 0.01, *** p < 0.001)

Article Snippet: The proteins were migrated to polyvinylidene difluoride (PVDF) membranes and combined with specific antibodies to GAPDH (ab181602), OPN (ab8448), RUNX2 (ab92336), β‐CATENIN (ab32572), METTL3 (ab195352) (all Abcam), LEF1 (C12A5) GSK (9832S) and P‐GSK (9323S) (Cell Signalling Technology).

Techniques: Over Expression, In Vitro, Transfection, Expressing

Journal: Cell reports

Article Title: TCF1 and LEF1 Control Treg Competitive Survival and Tfr Development to Prevent Autoimmune Diseases

doi: 10.1016/j.celrep.2019.05.061

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit monoclonal anti-LEF1 (C12A5) Al488 , Cell Signaling , Cat#8490S; RRID:AB_10949503.

Techniques: Virus, Plasmid Preparation, Recombinant, Staining, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Diagnostic Assay, Software, Extraction, Gene Expression