Journal: International Journal of Molecular Sciences

Article Title: PGC1α Cooperates with FOXA1 to Regulate Epithelial Mesenchymal Transition through the TCF4-TWIST1

doi: 10.3390/ijms23158247

Figure Lengend Snippet: Suppression of TCF4 and TWIST1 by ID1 is linked to loss of FOXA1-induced EMT. ( A ) Changes of EMT−associated genes by FOXA1 silencing in ectopic GFP or ID1 expressing A549 cells. ( B ) Interaction between TCF4 and TWIST1 in FOXA1 knockdown A549 cells. ( C ) Promoter occupancy of TCF4 and TWIST1 on CDH1 and CDH2 promoter in FOXA1 silenced A549 cells. ( D ) Changes of EMT-associated genes by FOXA1 silencing in control or TCF4 knockdown A549 cells. Values represent mean ± SD (n = 4). * p < 0.05 and ** p < 0.01 by one-way ANOVA with Tukey post-test for multiple comparisons. Not significate ns is normally this meaning.

Article Snippet: Human TCF4 targeting siRNA (sc-61657) was purchased from Santa Cruz Biotechnology (Dallas, TX, USA).

Techniques: Expressing, Knockdown, Control

Journal: International Journal of Molecular Sciences

Article Title: PGC1α Cooperates with FOXA1 to Regulate Epithelial Mesenchymal Transition through the TCF4-TWIST1

doi: 10.3390/ijms23158247

Figure Lengend Snippet: Suppression of TCF4 and TWIST1 by ID1 involves TGFβ1−mediated EMT gene expression. ( A ) Interaction of TCF4 and TWIST1 in the absence or presence of TGFβ1 (20 ng/mL for 24 h). ( B ) EMT-associated gene expression upon TGFβ1 (20 ng/mL for 24 h) in GFP or ID1 stably expressing A549 cells. ( C ) TGFβ1−mediated EMT gene expression in control or TCF4 silenced A549 cells. ( D ) Promoter occupancy of TCF4 on CDH1 and CDH2 promoter in GFP, FOXA1 or PGC1α stably expressing A549 cells. Values represent mean ± SD (n = 3). * p < 0.05 and * * p < 0.01 by one-way ANOVA with Tukey post−test for multiple comparisons. Not significate ns is normally this meaning.

Article Snippet: Human TCF4 targeting siRNA (sc-61657) was purchased from Santa Cruz Biotechnology (Dallas, TX, USA).

Techniques: Gene Expression, Stable Transfection, Expressing, Control

Journal: Drug Design, Development and Therapy

Article Title: Protective effect of Huang Gan formula in 5/6 nephrectomized rats by depressing the Wnt/β-catenin signaling pathway

doi: 10.2147/DDDT.S81157

Figure Lengend Snippet: Average optical density values of protein expressions by immunohistochemistry staining in renal tissues of rats in different groups, x ¯ + s ( n = 3 )

Article Snippet: Rabbit polyclonal antibody Wnt1 (ab85060; Abcam, Inc., Cambridge, MA, USA), β-catenin (BS1982; Bioworld Technology, Inc., St Louis Park, MN, USA), GSK-3β (sc9166; Santa Cruz Biotechnology, Santa Cruz, CA, USA), p-GSK-3β (sc-135653; Santa Cruz Biotechnology Inc., Dallas, TX, USA), transcription factor 4 (TCF4) (BS6172; Bioworld Technology, Inc.), dickkopf 1 (Dkk1) (BS7731; Bioworld Technology, Inc.), fibronectin 1 (Fn1) (ab2413; Abcam, Inc.), and glyceraldehyde 3-phosphate dehydro-genase (GAPDH) (ab9485; Abcam, Inc.) were purchased.

Techniques: Immunohistochemistry, Staining

Journal: Drug Design, Development and Therapy

Article Title: Protective effect of Huang Gan formula in 5/6 nephrectomized rats by depressing the Wnt/β-catenin signaling pathway

doi: 10.2147/DDDT.S81157

Figure Lengend Snippet: Representative micrographs of immunohistochemistry staining in the 5/6 Nx group rat renal cortex sections (400×). Notes: Immunohistochemistry staining of 5/6 Nx group for Wnt1 ( A ), β-catenin ( B ), GSK-3β ( C ), Dkk1 ( D ), TCF4 ( E ), Fn1 ( F ), and IgG negative control ( G ). Abbreviations: Nx, 5/6 nephrectomy; GSK-3β, glycogen synthase kinase-3β; Dkk1, dickkopf 1; TCF4, transcription factor 4; Fn1, fibronectin 1.

Article Snippet: Rabbit polyclonal antibody Wnt1 (ab85060; Abcam, Inc., Cambridge, MA, USA), β-catenin (BS1982; Bioworld Technology, Inc., St Louis Park, MN, USA), GSK-3β (sc9166; Santa Cruz Biotechnology, Santa Cruz, CA, USA), p-GSK-3β (sc-135653; Santa Cruz Biotechnology Inc., Dallas, TX, USA), transcription factor 4 (TCF4) (BS6172; Bioworld Technology, Inc.), dickkopf 1 (Dkk1) (BS7731; Bioworld Technology, Inc.), fibronectin 1 (Fn1) (ab2413; Abcam, Inc.), and glyceraldehyde 3-phosphate dehydro-genase (GAPDH) (ab9485; Abcam, Inc.) were purchased.

Techniques: Immunohistochemistry, Staining, Negative Control

Journal: Drug Design, Development and Therapy

Article Title: Protective effect of Huang Gan formula in 5/6 nephrectomized rats by depressing the Wnt/β-catenin signaling pathway

doi: 10.2147/DDDT.S81157

Figure Lengend Snippet: Wnt1, β-catenin, TCF4, GSK-3β, Dkk1, and Fn1 mRNA expression in renal tissues (n=6). Notes: Compared with the 5/6 Nx group, * P <0.05; compared with Sham and BC groups, # P <0.05; and compared with the losartan group, Δ P <0.05. Abbreviations: TCF4, transcription factor 4; GSK-3β, glycogen synthase kinase-3β; Dkk1, dickkopf 1; Fn1, fibronectin 1; Nx, 5/6 nephrectomy; Sham, sham-operated group; BC, background control group; UCG, uremic clearance granule; HGF-L, Huang Gan formula low dose group; HGF-M, Huang Gan formula middle dose group; HGF-H, Huang Gan formula high dose group.

Article Snippet: Rabbit polyclonal antibody Wnt1 (ab85060; Abcam, Inc., Cambridge, MA, USA), β-catenin (BS1982; Bioworld Technology, Inc., St Louis Park, MN, USA), GSK-3β (sc9166; Santa Cruz Biotechnology, Santa Cruz, CA, USA), p-GSK-3β (sc-135653; Santa Cruz Biotechnology Inc., Dallas, TX, USA), transcription factor 4 (TCF4) (BS6172; Bioworld Technology, Inc.), dickkopf 1 (Dkk1) (BS7731; Bioworld Technology, Inc.), fibronectin 1 (Fn1) (ab2413; Abcam, Inc.), and glyceraldehyde 3-phosphate dehydro-genase (GAPDH) (ab9485; Abcam, Inc.) were purchased.

Techniques: Expressing

Journal: Drug Design, Development and Therapy

Article Title: Protective effect of Huang Gan formula in 5/6 nephrectomized rats by depressing the Wnt/β-catenin signaling pathway

doi: 10.2147/DDDT.S81157

Figure Lengend Snippet: Western blot analysis for detection of Wnt1, β-catenin, TCF4, GSK-3β, Dkk1, and Fn1 expression in the kidney. Notes: Western blot bands of different proteins ( A ); semi-quantitative analysis of the bands (from ( B ) to ( G ) are Wnt1, β-catenin, GSK-3β, Dkk1, TCF4, and Fn1 in sequence). Compared with 5/6 Nx group, * P <0.05; compared with Sham and BC groups, # P <0.05; and compared with the losartan group Δ P <0.05. Abbreviations: TCF4, transcription factor 4; GSK-3β, glycogen synthase kinase-3β; Dkk1, dickkopf 1; Fn1, fibronectin 1; Nx, 5/6 nephrectomy; Sham, sham-operated group; BC, background control group; UCG, uremic clearance granule; HGF-L, Huang Gan formula low dose group; HGF-M, Huang Gan formula middle dose group; HGF-H, Huang Gan formula high dose group; GAPDH, glyceraldehyde 3-phosphate dehydrogenase.

Article Snippet: Rabbit polyclonal antibody Wnt1 (ab85060; Abcam, Inc., Cambridge, MA, USA), β-catenin (BS1982; Bioworld Technology, Inc., St Louis Park, MN, USA), GSK-3β (sc9166; Santa Cruz Biotechnology, Santa Cruz, CA, USA), p-GSK-3β (sc-135653; Santa Cruz Biotechnology Inc., Dallas, TX, USA), transcription factor 4 (TCF4) (BS6172; Bioworld Technology, Inc.), dickkopf 1 (Dkk1) (BS7731; Bioworld Technology, Inc.), fibronectin 1 (Fn1) (ab2413; Abcam, Inc.), and glyceraldehyde 3-phosphate dehydro-genase (GAPDH) (ab9485; Abcam, Inc.) were purchased.

Techniques: Western Blot, Expressing, Sequencing

Journal: Drug Design, Development and Therapy

Article Title: Protective effect of Huang Gan formula in 5/6 nephrectomized rats by depressing the Wnt/β-catenin signaling pathway

doi: 10.2147/DDDT.S81157

Figure Lengend Snippet: Primers used for RT-PCR assay

Article Snippet: Rabbit polyclonal antibody Wnt1 (ab85060; Abcam, Inc., Cambridge, MA, USA), β-catenin (BS1982; Bioworld Technology, Inc., St Louis Park, MN, USA), GSK-3β (sc9166; Santa Cruz Biotechnology, Santa Cruz, CA, USA), p-GSK-3β (sc-135653; Santa Cruz Biotechnology Inc., Dallas, TX, USA), transcription factor 4 (TCF4) (BS6172; Bioworld Technology, Inc.), dickkopf 1 (Dkk1) (BS7731; Bioworld Technology, Inc.), fibronectin 1 (Fn1) (ab2413; Abcam, Inc.), and glyceraldehyde 3-phosphate dehydro-genase (GAPDH) (ab9485; Abcam, Inc.) were purchased.

Techniques: Sequencing, Amplification

Journal: Cell death & disease

Article Title: Aberrant Gcm1 expression mediates Wnt/β-catenin pathway activation in folate deficiency involved in neural tube defects.

doi: 10.1038/s41419-020-03313-z

Figure Lengend Snippet: Fig. 1 Wnt/β-catenin pathway signaling activation in cells with folate deficiency. a Morphology of C57BL/6 mESCs as observed with an optical microscope at magnifications of ×100 and ×200, F6, sixth generation of C57BL/6 mESCs. b Analysis of the cell cycle distribution of C57BL/6 mESCs after six generations of culture under folate deficiency by flow cytometry. 3 × 106 cells were harvested for control and case group respectively; right panel: the results of cell cycle quantification were graded according to the cell cycle phase. c Analysis of apoptosis in C57BL/6 mESCs after six generations of culture under folate deficiency by flow cytometry. 3 × 106 cells were harvested for control and case group, respectively, the number of cells in early apoptosis is shown in a bar graph in the right panel. d Folate concentration in C57BL/6 mESCs after six generations of culture in folate deficiency and in NE-4C after three generations of culture in folate deficiency. e The mRNA expression of Fzd5, Lrp, Wnt6, Ccnd1, Nfat5, Lef1, TCF4, TCF12, Bcl9l, and Axin2 in C57BL/6 mESCs with folate deficiency. f Analysis of luciferase activity by TOP/FOP Flash assays in C57BL/6 mESCs subjected to folate deficiency for six generations and NE-4C subjected to folate deficiency for three generations. Data a–f represent the mean ± SEM (n = 3). The p value was calculated by Student’s t-test, ns was for no significance, *P < 0.05, **P < 0.01, ***P < 0.001.

Article Snippet: siRNA reagents for Gcm1 and TCF4 were purchased from Santa Cruz Biotechnology (USA).

Techniques: Activation Assay, Microscopy, Cytometry, Control, Concentration Assay, Expressing, Luciferase, Activity Assay

Journal: The Journal of Clinical Investigation

Article Title: FOXOs attenuate bone formation by suppressing Wnt signaling

doi: 10.1172/JCI68049

Figure Lengend Snippet: (A) Wnt-target genes determined by microarray analysis in Osx1-GFP calvaria cells. (B) mRNA levels by qRT-PCR in calvaria cells treated with vehicle (veh) or 50 ng/ml Wnt3a overnight (triplicates). (C) mRNA levels by qRT-PCR in femoral shafts from 3-month-old mice (n = 8–9/group), and in periosteal and bone marrow osteoblastic cell cultures described in Figure ​Figure4,4, C and G. (D) mRNA by qRT-PCR (left, triplicates) and protein levels by Western blot (right) in calvaria-derived cells infected with Adeno-Cre to induce Foxo deletion. (E) BrdU incorporation in cells, described in D, treated with veh (–) or 50 ng/ml Wnt3a (triplicates). (F) Lysates from nuclear extracts of calvaria cells, described in D, treated with vehicle or 50 ng/ml Wnt3a, immunoprecipitated with an anti–TCF-4 or anti-IgG antibody, and probed with an anti–β-catenin antibody. (G) mRNA levels by qRT-PCR and (H) BrdU incorporation in calvaria cells infected with Adeno-Cre and with lentivirus expressing shRNA directed against β-catenin or nontargeted shRNA followed by treatment with vehicle or 50 ng/ml Wnt3a overnight (triplicates). #P < 0.05 by Student’s t test; *P < 0.05 by ANOVA with Bonferroni’s test. Bars represent mean + SD.

Article Snippet: To analyze β-catenin–TCF-4 interactions, cell lysates were immunoprecipitated with an anti–TCF-4 antibody (#2569S; Cell Signaling) or an IgG nonspecific control antibody (sc-2003; Santa Cruz Biotechnology Inc.).

Techniques: Microarray, Quantitative RT-PCR, Western Blot, Derivative Assay, Infection, BrdU Incorporation Assay, Immunoprecipitation, Expressing, shRNA

Journal: Chemmedchem

Article Title: 4‐(5‐Chloro‐3‐(3,4,5‐trimethoxybenzoyl)‐1 H ‐indol‐1‐yl)benzenesulfonamide: A Novel Polypharmacology Agent to Target Carbonic Anhydrase IX and XII With Improved Selectivity, Wnt/β‐Catenin Signaling Pathway, and P‐Glycoprotein

doi: 10.1002/cmdc.202500996

Figure Lengend Snippet: Compound 15 inhibits the β‐catenin/TCF‐4 interaction. A coimmunoprecipitation assay was performed in HEK293T cells transiently transfected with FLAG‐tagged β‐catenin (β‐catenin FLAG ) and MYC‐tagged TCF‐4 (TCF‐4 MYC ). Cells were treated with 50 mM LiCl and 60 μM 15 for 24 h. β‐Catenin FLAG was immunoprecipitated from total protein extracts using anti‐FLAG‐conjugated agarose beads (left panel; IP: immunoprecipitation). Coimmunoprecipitated proteins were analyzed by Western blotting using an anti‐MYC‐tag antibody to detect TCF‐4 MYC (left panel; IB: immunoblot). Input shows the levels of TCF‐4 MYC and β‐catenin FLAG in 5% whole‐cell lysates before immunoprecipitation (right panel).

Article Snippet: HEK293T cells were transfected with pcDNA/Myc TCF‐4 (Addgene #16512) and human β‐catenin pcDNA3 (#16828 Addgene).

Techniques: Co-Immunoprecipitation Assay, Transfection, Immunoprecipitation, Western Blot