Journal: Cells

Article Title: Conjugated Bile Acids Promote Lymphangiogenesis by Modulation of the Reactive Oxygen Species–p90RSK–Vascular Endothelial Growth Factor Receptor 3 Pathway

doi: 10.3390/cells12040526

Figure Lengend Snippet: Antibodies used in Western blot, immunohistochemistry, and immunofluorescence.

Article Snippet: YAP1 , WB , 1:1000 , Novus , NB110-58358 , AB_922796.

Techniques: Western Blot, Immunohistochemistry, Immunofluorescence

Journal: bioRxiv

Article Title: KLF7-Regulated ITGA2 as a Therapeutic Target for Inhibiting Oral Cancer Stem Cells

doi: 10.1101/2024.11.04.621805

Figure Lengend Snippet: A The log2-transformed fold change (FC) in RNA levels between the stem-like cluster with the rest of the clusters and the stem-related pathway with the rest of the pathway. B Immunoblot assay of ITGA2, phosphor-ERK1/2, total-ERK1/2, phosphor-AKT, total-AKT, phosphor-YAP1, total-YAP1 protein levels in CAL27 and HSC3 cells after stable silencing ITGA2. C Representative immunofluorescent image showing the expression of YAP1 and DAPI in wt and stable silencing ITGA2 cells. D The sequence and domain in ITGA2 protein. E Co-IP analysis in CAL27 and HSC3 cells transduced with ITGA2 OE and ITGA2 mut plasmid. F Immunoblot assay of phosphor-ERK1/2, total-ERK1/2, phosphor-AKT, total-AKT, phosphor-YAP1, total-YAP1 protein levels in CAL27 and HSC3 cells after transduced with ITGA2 OE and ITGA2 mut plasmid. G Representative immunofluorescent image showing the expression of YAP1 and DAPI in CAL27 and HSC3 cells after transduced with ITGA2 OE and ITGA2 mut plasmid.

Article Snippet: After washing and blocking, the primary antibody against YAP1 (66900-1-Ig, Proteintech) was incubated overnight.

Techniques: Transformation Assay, Western Blot, Expressing, Sequencing, Co-Immunoprecipitation Assay, Transduction, Plasmid Preparation

Journal: bioRxiv

Article Title: KLF7-Regulated ITGA2 as a Therapeutic Target for Inhibiting Oral Cancer Stem Cells

doi: 10.1101/2024.11.04.621805

Figure Lengend Snippet: A The molecular formula of TC-I 15. B Co-IP analysis in CAL27 and HSC3 cells treated with DMSO and TC-I 15. C CAL27 cells were intracardially injected into mice. Seven days later, TC-I 15 was injected (20 mg/kg) into mice via vein every third day for 1 month. D Representative images showing the xenograft model in CA27 cells treated with DMSO or TC-I 15(n = 5 per group) and the growth of tumor grafts was shown. E Immunoblot assay of phosphor-ERK1/2, total-ERK1/2, phosphor-AKT, total-AKT, phosphor-YAP1, and total-YAP1 protein levels in xenograft tissue. F Representative FACS plots and quantification of CD133+cells in the xenograft tissues. G CAL27 cells were intracardially injected into mice. Seven days later, TC-I 15 (20 mg/kg) and plastin(5 mg/kg) were injected into nude mice via vein every third day for 1 month. H Representative images showing the xenograft model in CA27 cells treated with DMSO or TC-I 15(n = 5 per group) and the growth of tumor grafts were shown.

Article Snippet: After washing and blocking, the primary antibody against YAP1 (66900-1-Ig, Proteintech) was incubated overnight.

Techniques: Co-Immunoprecipitation Assay, Injection, Western Blot

Journal: PLoS ONE

Article Title: Yes-Associated Protein (YAP) Modulates Oncogenic Features and Radiation Sensitivity in Endometrial Cancer

doi: 10.1371/journal.pone.0100974

Figure Lengend Snippet: (A) Expression of YAP and phospho-YAP (Ser127) in three EMCA cell lines by western blotting. (B) Immunofluorescent cytochemical staining of endogenous YAP using anti-YAP antibody (YAP: green, DAPI: blue).

Article Snippet: Samples were then incubated in YAP antibody (1∶200, NB110-58358) from Novus Biologicals (Littleton, CO) for 30 min with a subsequent TBST wash and incubated in Dako Envision+ HRP rabbit solution (Dako, Carpinteria, CA) (diluted according to manufacturer's instructions) for 30 minutes.

Techniques: Expressing, Western Blot, Staining

Journal: Frontiers in Molecular Biosciences

Article Title: Folic Acid Alleviates High Glucose and Fat-Induced Pyroptosis via Inhibition of the Hippo Signal Pathway on H9C2 Cells

doi: 10.3389/fmolb.2021.698698

Figure Lengend Snippet: Effect of folic acid on the pyroptosis and Hippo signaling pathway. (A) Representative immunoblot showing expression levels of NLRP3, ACS, Cleaved-Caspase-1 and Caspase-1. (B) Quantification of NLRP3, ACS, Cleaved-Caspase-1 and Caspase-1 with α-tubulin as the loading control. (C) IL-1β and IL-18 levels in the cell supernatant. (D) Representative immunoblot showing expression levels of YAP1 and p-YAP1. (E) Quantification of YAP1 and p-YAP1 with α-tubulin as the loading control. * p < 0.05, compared with NG; # p < 0.05, compared with HGF; & p < 0.05, compared with HGF+50 nM folic acid.

Article Snippet: Each membrane was blocked for 1.5 h at room temperature in 5% non-fat milk, followed by incubation overnight at 4°C with the primary antibodies against NLRP3 (1:1000, abcam, ab263899, United Kingdom), apoptosis associated speck like protein containing a CARD (ASC) (1:500, abcam, ab180799, United Kingdom), Caspase-1 (1:1000, Immunoway,YT5743, United States), cleaved-Caspase-1 (1:1000, abcam, ab179515 United States), YAP1 (1:2000, proteintech, 66900-1-Ig, China), phosphor-YAP1 (ser127) (1:1000, cell signaling technology, 13008s, United States), YAP1 (1:2000, proteintech, 66900-1-Ig, China), phosphor-YAP1 (ser127), LATS (1:1000, proteintech, 17049-1-AP, China), phosphor-LATS1 (Thr1079/1041) (1:1000, immunoway, YP1222, United States), and α-tubulin (1:2000, abcam, ab176560, United Kingdom).

Techniques: Western Blot, Expressing

Journal: Frontiers in Molecular Biosciences

Article Title: Folic Acid Alleviates High Glucose and Fat-Induced Pyroptosis via Inhibition of the Hippo Signal Pathway on H9C2 Cells

doi: 10.3389/fmolb.2021.698698

Figure Lengend Snippet: Verteporfin decreased the protective effect of folic acid on H9C2 cells by inhibiting YAP expression. (A) The effect of verteporfin on the percentage of viable H9C2 cells. (B) Representative immunoblot showing expression levels of YAP and p-YAP in H9C2 cells treated with verteporfin. (D, G) Representative immunoblot showing levels of NLRP3, ACS, Cleaved-Caspase-1, Caspase-1, YAP and p-YAP. (H) IL-1β and IL-18 levels in the cell supernatant. (C, E, F) Quantification of YAP1, p-YAP1, NLRP3, ACS, with α-tubulin as the loading control, for Cleaved-caspase-1, Caspase-1 as the loading control. * p < 0.05, compared with NG; # p < 0.05, compared with HGF, & p < 0.05, compared with HGF + Folic acid.

Article Snippet: Each membrane was blocked for 1.5 h at room temperature in 5% non-fat milk, followed by incubation overnight at 4°C with the primary antibodies against NLRP3 (1:1000, abcam, ab263899, United Kingdom), apoptosis associated speck like protein containing a CARD (ASC) (1:500, abcam, ab180799, United Kingdom), Caspase-1 (1:1000, Immunoway,YT5743, United States), cleaved-Caspase-1 (1:1000, abcam, ab179515 United States), YAP1 (1:2000, proteintech, 66900-1-Ig, China), phosphor-YAP1 (ser127) (1:1000, cell signaling technology, 13008s, United States), YAP1 (1:2000, proteintech, 66900-1-Ig, China), phosphor-YAP1 (ser127), LATS (1:1000, proteintech, 17049-1-AP, China), phosphor-LATS1 (Thr1079/1041) (1:1000, immunoway, YP1222, United States), and α-tubulin (1:2000, abcam, ab176560, United Kingdom).

Techniques: Expressing, Western Blot

Journal: Frontiers in Molecular Biosciences

Article Title: Folic Acid Alleviates High Glucose and Fat-Induced Pyroptosis via Inhibition of the Hippo Signal Pathway on H9C2 Cells

doi: 10.3389/fmolb.2021.698698

Figure Lengend Snippet: The anti-pyroptosis activity of folic acid was depended on YAP1. (A) siRNA significantly inhibits the expression of YAP1. (B) Representative immunoblot levels of NLRP3, ACS, Cleaved-Caspase-1, Caspase-1 in scr and siYAP1 H9C2 cells treated with different medium. (C) IL-1β and IL-18 levels in the cell supernatant. (D–F) Quantification of NLRP3, ACS, with α-tubulin as the loading control; for Cleaved-caspase-1, Caspase-1 as the loading control. ** p < 0.01, compared with NG treated scr cells; # p < 0.01, compared with HGF treated scr cells, & p < 0.05, compared with HGF+ 500 nM folic acid treated siYAP1 cells.

Article Snippet: Each membrane was blocked for 1.5 h at room temperature in 5% non-fat milk, followed by incubation overnight at 4°C with the primary antibodies against NLRP3 (1:1000, abcam, ab263899, United Kingdom), apoptosis associated speck like protein containing a CARD (ASC) (1:500, abcam, ab180799, United Kingdom), Caspase-1 (1:1000, Immunoway,YT5743, United States), cleaved-Caspase-1 (1:1000, abcam, ab179515 United States), YAP1 (1:2000, proteintech, 66900-1-Ig, China), phosphor-YAP1 (ser127) (1:1000, cell signaling technology, 13008s, United States), YAP1 (1:2000, proteintech, 66900-1-Ig, China), phosphor-YAP1 (ser127), LATS (1:1000, proteintech, 17049-1-AP, China), phosphor-LATS1 (Thr1079/1041) (1:1000, immunoway, YP1222, United States), and α-tubulin (1:2000, abcam, ab176560, United Kingdom).

Techniques: Activity Assay, Expressing, Western Blot

Journal: Frontiers in Molecular Biosciences

Article Title: Folic Acid Alleviates High Glucose and Fat-Induced Pyroptosis via Inhibition of the Hippo Signal Pathway on H9C2 Cells

doi: 10.3389/fmolb.2021.698698

Figure Lengend Snippet: Folic acid reduces myocardial pyroptosis by inhibiting activation of Hippo signaling pathway in HFD + STZ mice. (A) Blood glucose curve; (B) body weight curve; (C) Representative immunoblot showing levels of NLRP3, ACS, Cleaved-Caspase-1, LATS, p-LATS, YAP and p-YAP; (D) Quantification of NLRP3, ACS, Cleaved-caspase-1, LATS, p-LATS, YAP1, p-YAP1, α-tubulin was used as the loading control. * p < 0.05, compared with control group; # p < 0.05, compared with HFD + STZ group. CON: normal diet mice; HFD + STZ: HFD + STZ mice treated with PBS; HFD + STZ + folic acid: HFD + STZ mice treated with folic acid.

Article Snippet: Each membrane was blocked for 1.5 h at room temperature in 5% non-fat milk, followed by incubation overnight at 4°C with the primary antibodies against NLRP3 (1:1000, abcam, ab263899, United Kingdom), apoptosis associated speck like protein containing a CARD (ASC) (1:500, abcam, ab180799, United Kingdom), Caspase-1 (1:1000, Immunoway,YT5743, United States), cleaved-Caspase-1 (1:1000, abcam, ab179515 United States), YAP1 (1:2000, proteintech, 66900-1-Ig, China), phosphor-YAP1 (ser127) (1:1000, cell signaling technology, 13008s, United States), YAP1 (1:2000, proteintech, 66900-1-Ig, China), phosphor-YAP1 (ser127), LATS (1:1000, proteintech, 17049-1-AP, China), phosphor-LATS1 (Thr1079/1041) (1:1000, immunoway, YP1222, United States), and α-tubulin (1:2000, abcam, ab176560, United Kingdom).

Techniques: Activation Assay, Western Blot

Journal: Theranostics

Article Title: Remodeling cancer stemness by collagen/fibronectin via the AKT and CDC42 signaling pathway crosstalk in glioma

doi: 10.7150/thno.50613

Figure Lengend Snippet: The integrin αvβ3/CDC42/F-actin/YAP/NUPR1/Nestin signaling pathway is activated in collagen/FN-cultured glioma cell (A) Expression of CDC42, F-actin, and β-actin in LN229 cells and T98G cells cultured in a flask or 3D Collagen/FN gel. (B) Expression of CDC42, F-actin, and β-actin in LN229 cells and T98G cells cultured in 3D Collagen/FN gel treated with PBS or SB273005 (5 nM, 24). (C) Cell fraction of cytosol (C) and nucleus (N) was analyzed by Western blotting. Expression of YAP1 in LN229 cells or LN229 CDC42 OE cells cultured in a flask or 3D Collagen/FN gel and treated with PBS or SB273005 (5 nM, 24 h). (D) Western blotting analysis of proteins immunoprecipitated (IP) with anti-YAP1 from LN229 cells cultured in a flask or 3D Collagen/FN gel for 48 h. (E) ChIP analysis of YAP1 or TEAD4 binding to the NUPR1 promoter in LN229 cells. YAP1 and TEAD4 increased the luciferase activity of the NUPR1 promoter in LN229 cells. (F) Expression of NUPR1 and Nestin in LN229 cells and T98G cells cultured in a flask or 3D Collagen/FN gel. (G) Expression of NUPR1in LN229 and T98G cells cultured in 3D Collagen/FN gel, treated with PBS or SB273005 (5 nM, 24 h) and transfected with CDC42 OE lentivirus. (H) Representative photographs of LN229-NC cells and LN229-CDC42 OE cells cultured in 3D Collagen/FN gel. Scale bar represents 20 µm (I) Representative photographs of LN229-NC cells and LN229-Nestin shRNA cultured in 3D Collagen/FN gel. Scale bar represents 20 µm (J) Quantification of colony sizes in (H) and (I) (K) Proliferation of 3D collagen/FN pre-cultured LN229-vec cells, LN229-CDC42 OE cells, LN229-NC cells, and LN229-Nestin shRNA cells (L) Colony formation of 3D collagen/FN pre-cultured LN229-vec cells, LN229-CDC42 OE cells, LN229-NC cells, and LN229-Nestin shRNA cells. Mean ± SEM, n.s, no significant difference, *p < 0.05, **p < 0.01.

Article Snippet: LN229 cells were harvested, lysed, and immunoprecipitated with anti-YAP1 (MAB8094, RD, USA) or IgG conjugated to protein G agarose (11243233001, Roche, USA) at 4 ºC overnight.

Techniques: Cell Culture, Expressing, Western Blot, Immunoprecipitation, Binding Assay, Luciferase, Activity Assay, Transfection, shRNA