fzd 2 Thermo Fisher Search Results


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  • 87
    Thermo Fisher gene exp fzd2 mm02524776 s1
    Expression of Lrp5 and <t>Fzd2</t> mRNAs and proteins in α7 + hSMSC-OB cells, MC3T3-E1 cells, and α7 + hSMSCs. ( A – F ) Cells were stimulated with IL-1β (0, 0.1, 0.3 and 1 ng/mL) prior to real-time qPCR analysis of Lrp5 and Fzd2 expression relative to the control (18S rRNA). Data are the mean ± SD of four independent experiments. * p
    Gene Exp Fzd2 Mm02524776 S1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 87/100, based on 38 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gene exp fzd2 mm02524776 s1/product/Thermo Fisher
    Average 87 stars, based on 38 article reviews
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    85
    Thermo Fisher human fzd 2
    Enhanced Wnt signaling occurs in the absence of NHERF1 ( a ) CHO-N10 cells were co-transfected with the indicated Fzd receptors and either TOP or FOP luciferase reporter plasmid. NHERF1 expression blunted Wnt-induced luciferase expression via Fzd 2, 4 and 7 but had no effect on Wnt signaling through Fzd3 (** p
    Human Fzd 2, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/human fzd 2/product/Thermo Fisher
    Average 85 stars, based on 2 article reviews
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    88
    Thermo Fisher gene exp fzd2 hs00361432 s1
    mRNA expression analyses of Wnt ligands, Fzd receptors and Wnt antagonists . RT-PCR detection of mRNAs for Wnt ligands, Fzd receptors and Wnt antagonists in BM B progenitor cells. The + and - symbols indicate the presence and absence of reverse transcriptase in the reaction mix, respectively. One representative of two experiments is shown. Amplicon sizes: Wnt2B: 328 bp, Wnt5B:279 bp, Wnt8A: 400 bp, Wnt10A: 296 bp, Wnt16: 520 bp, <t>Fzd2:</t> 306 bp, Fzd3: 622 bp, Fzd4: 605 bp, Fzd5: 197 bp, Fzd6: 300 bp, Fzd9: 210 bp, sFRP4: 243 bp, WIF1: 200 bp, Dkk1: 235 bp, Dkk4: 241 bp. M: Size marker 1 kb Plus DNA ladder (Invitrogen, USA). Where two different bands are detected, an arrow marks the correct band.
    Gene Exp Fzd2 Hs00361432 S1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 88/100, based on 24 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gene exp fzd2 hs00361432 s1/product/Thermo Fisher
    Average 88 stars, based on 24 article reviews
    Price from $9.99 to $1999.99
    gene exp fzd2 hs00361432 s1 - by Bioz Stars, 2020-08
    88/100 stars
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    86
    Thermo Fisher gene exp fzd2 rn00597004 s1
    mRNA expression analyses of Wnt ligands, Fzd receptors and Wnt antagonists . RT-PCR detection of mRNAs for Wnt ligands, Fzd receptors and Wnt antagonists in BM B progenitor cells. The + and - symbols indicate the presence and absence of reverse transcriptase in the reaction mix, respectively. One representative of two experiments is shown. Amplicon sizes: Wnt2B: 328 bp, Wnt5B:279 bp, Wnt8A: 400 bp, Wnt10A: 296 bp, Wnt16: 520 bp, <t>Fzd2:</t> 306 bp, Fzd3: 622 bp, Fzd4: 605 bp, Fzd5: 197 bp, Fzd6: 300 bp, Fzd9: 210 bp, sFRP4: 243 bp, WIF1: 200 bp, Dkk1: 235 bp, Dkk4: 241 bp. M: Size marker 1 kb Plus DNA ladder (Invitrogen, USA). Where two different bands are detected, an arrow marks the correct band.
    Gene Exp Fzd2 Rn00597004 S1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gene exp fzd2 rn00597004 s1/product/Thermo Fisher
    Average 86 stars, based on 1 article reviews
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    85
    Thermo Fisher gene exp fzd2 mm01250498 s1
    mRNA expression analyses of Wnt ligands, Fzd receptors and Wnt antagonists . RT-PCR detection of mRNAs for Wnt ligands, Fzd receptors and Wnt antagonists in BM B progenitor cells. The + and - symbols indicate the presence and absence of reverse transcriptase in the reaction mix, respectively. One representative of two experiments is shown. Amplicon sizes: Wnt2B: 328 bp, Wnt5B:279 bp, Wnt8A: 400 bp, Wnt10A: 296 bp, Wnt16: 520 bp, <t>Fzd2:</t> 306 bp, Fzd3: 622 bp, Fzd4: 605 bp, Fzd5: 197 bp, Fzd6: 300 bp, Fzd9: 210 bp, sFRP4: 243 bp, WIF1: 200 bp, Dkk1: 235 bp, Dkk4: 241 bp. M: Size marker 1 kb Plus DNA ladder (Invitrogen, USA). Where two different bands are detected, an arrow marks the correct band.
    Gene Exp Fzd2 Mm01250498 S1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/gene exp fzd2 mm01250498 s1/product/Thermo Fisher
    Average 85 stars, based on 7 article reviews
    Price from $9.99 to $1999.99
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    93
    Thermo Fisher rabbit anti fzd2 antibodies
    Expression of CFIm25 and its targets in lung fibroblasts isolated from Col1a1-CreER-CFIm25 fl/fl mice. Four- to six-week-old Col1a1-CreER-CFIm25 fl/fl mice and age- and sex-matched littermate controls were administrated 75 mg/kg (i.p.) tamoxifen daily for 5 days to induce Cre expression. n = 4 biological replications. One week later, fibroblasts were isolated from the lungs of these mice. ( A ) Western blotting was used to confirm the expression of CFIm25, Cre, and CFIm25 targets in fibroblasts, and ( B ) qRT-PCR was performed to determine the dPAS usage of CFIm25 targets ( Col1a1, Tgfbr1, <t>Fzd2</t> , and Wnt5a ). n = 5 biological replicates. * P
    Rabbit Anti Fzd2 Antibodies, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 93/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/rabbit anti fzd2 antibodies/product/Thermo Fisher
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    90
    Thermo Fisher antibodies against fzd2
    <t>FZD2</t> blockade interferes with canonical and non-canonical signaling in NB tissue A. mRNA expression of SK-N-AS and SK-N-DZ human NB tumor tissue treated with FZD2 siRNA (FZD2si) or scrambled siRNA (Co) on day 24. Graphs show the results of qRT-PCR for human FZD2, Wnt3a, Wnt5a, MYC and cyclin D1 as well as for murine CD31. Data are mean ± SD (n = 8 per group). B. Representative Western blot images and C. quantification of immunoblots stained with FZD2, phospho-LRP6 (p-LRP6), total β-catenin, active β-catenin, MYC, cyclin D1, pan-phospho-PKC, (p-PKC), total JNK, phospho-JNK (p-JNK), total AKT, phospho-AKT (p-AKT), total ERK, phospho-ERK (p-ERK) antibodies. FZD2 blockade resulted in decreased levels of FZD2, p-LRP6, active β-catenin, MYC and cyclin D1 associated with increased levels of p-PKC, p-AKT and p-ERK in NB tissue. Data are mean ± SD (n = 8 per group). Asterisks (*) indicate P
    Antibodies Against Fzd2, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/antibodies against fzd2/product/Thermo Fisher
    Average 90 stars, based on 6 article reviews
    Price from $9.99 to $1999.99
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    Image Search Results


    Expression of Lrp5 and Fzd2 mRNAs and proteins in α7 + hSMSC-OB cells, MC3T3-E1 cells, and α7 + hSMSCs. ( A – F ) Cells were stimulated with IL-1β (0, 0.1, 0.3 and 1 ng/mL) prior to real-time qPCR analysis of Lrp5 and Fzd2 expression relative to the control (18S rRNA). Data are the mean ± SD of four independent experiments. * p

    Journal: International Journal of Molecular Sciences

    Article Title: Wnt16 Signaling Is Required for IL-1β-Induced Matrix Metalloproteinase-13-Regulated Proliferation of Human Stem Cell-Derived Osteoblastic Cells

    doi: 10.3390/ijms17020221

    Figure Lengend Snippet: Expression of Lrp5 and Fzd2 mRNAs and proteins in α7 + hSMSC-OB cells, MC3T3-E1 cells, and α7 + hSMSCs. ( A – F ) Cells were stimulated with IL-1β (0, 0.1, 0.3 and 1 ng/mL) prior to real-time qPCR analysis of Lrp5 and Fzd2 expression relative to the control (18S rRNA). Data are the mean ± SD of four independent experiments. * p

    Article Snippet: The following primer/probe sets were used: human Wnt16, Mm00437347_m1; mouse Wnt16, Mm00437347_m1; human Wnt5a, Mm00437347_m1; mouse Wnt5a, Mm00437347_m1; human Wnt5b, Mm00437347_m1; mouse Wnt5b, Mm01183986_m1; human Lrp5, Mm01227476_m1; mouse Lrp5, Mm01227476_m1; human Fzd2, Mm02524776_s1; mouse Fzd2, Mm02524776_s1; human Fzd9, Mm01206511_s1; mouse Fzd9, Mm01206511_s1; human Lrp6, Hs00233945_m1; mouse Lrp6, Mm00999795_m1; human Ror1, Hs00938677_m1; mouse Ror1, Mm00443462_m1; human Ror2, Hs00896176_m1; mouse Ror2, Mm00443470_m1; human Ryk Hs00243196_m1; mouse Ryk, Mm01238551_m1 (Assays-On-Demand™, Applied Biosystems, Foster City, CA, USA).

    Techniques: Expressing, Real-time Polymerase Chain Reaction

    Enhanced Wnt signaling occurs in the absence of NHERF1 ( a ) CHO-N10 cells were co-transfected with the indicated Fzd receptors and either TOP or FOP luciferase reporter plasmid. NHERF1 expression blunted Wnt-induced luciferase expression via Fzd 2, 4 and 7 but had no effect on Wnt signaling through Fzd3 (** p

    Journal: Oncogene

    Article Title: Direct interaction between NHERF1 and Frizzled regulates ?-catenin signaling

    doi: 10.1038/onc.2010.389

    Figure Lengend Snippet: Enhanced Wnt signaling occurs in the absence of NHERF1 ( a ) CHO-N10 cells were co-transfected with the indicated Fzd receptors and either TOP or FOP luciferase reporter plasmid. NHERF1 expression blunted Wnt-induced luciferase expression via Fzd 2, 4 and 7 but had no effect on Wnt signaling through Fzd3 (** p

    Article Snippet: Human Fzd 2, 3, and 7 were purchased from OpenBiosystems, Inc. and subcloned into pcDNA3.1(+) using EcoRI and XhoI.

    Techniques: Transfection, Luciferase, Plasmid Preparation, Expressing

    mRNA expression analyses of Wnt ligands, Fzd receptors and Wnt antagonists . RT-PCR detection of mRNAs for Wnt ligands, Fzd receptors and Wnt antagonists in BM B progenitor cells. The + and - symbols indicate the presence and absence of reverse transcriptase in the reaction mix, respectively. One representative of two experiments is shown. Amplicon sizes: Wnt2B: 328 bp, Wnt5B:279 bp, Wnt8A: 400 bp, Wnt10A: 296 bp, Wnt16: 520 bp, Fzd2: 306 bp, Fzd3: 622 bp, Fzd4: 605 bp, Fzd5: 197 bp, Fzd6: 300 bp, Fzd9: 210 bp, sFRP4: 243 bp, WIF1: 200 bp, Dkk1: 235 bp, Dkk4: 241 bp. M: Size marker 1 kb Plus DNA ladder (Invitrogen, USA). Where two different bands are detected, an arrow marks the correct band.

    Journal: BMC Immunology

    Article Title: Wnt expression and canonical Wnt signaling in human bone marrow B lymphopoiesis

    doi: 10.1186/1471-2172-7-13

    Figure Lengend Snippet: mRNA expression analyses of Wnt ligands, Fzd receptors and Wnt antagonists . RT-PCR detection of mRNAs for Wnt ligands, Fzd receptors and Wnt antagonists in BM B progenitor cells. The + and - symbols indicate the presence and absence of reverse transcriptase in the reaction mix, respectively. One representative of two experiments is shown. Amplicon sizes: Wnt2B: 328 bp, Wnt5B:279 bp, Wnt8A: 400 bp, Wnt10A: 296 bp, Wnt16: 520 bp, Fzd2: 306 bp, Fzd3: 622 bp, Fzd4: 605 bp, Fzd5: 197 bp, Fzd6: 300 bp, Fzd9: 210 bp, sFRP4: 243 bp, WIF1: 200 bp, Dkk1: 235 bp, Dkk4: 241 bp. M: Size marker 1 kb Plus DNA ladder (Invitrogen, USA). Where two different bands are detected, an arrow marks the correct band.

    Article Snippet: Predeveloped assay reagents including primers and probes for LRP5 (Hs00182031_m1), LRP6 (Hs00233935_m1), Fzd2 (Hs00361432_s1), Fzd5 (Hs00361869_g1), Fzd6 (Hs00171574_m1), Fzd9 (Hs00268954_s1), β-catenin (CTNNB1, Hs00170025_m1), plakoglobin (JUP, Hs00158408_m1), LEF-1 (Hs00212390_m1) and TCF-4 (Hs00181036_m1) mRNAs as well as the endogenous control phosphoglycerate kinase 1 (PGK1) (Hs99999906_m1) were supplied by Applied Biosystems and the PCR reactions were performed according to the manufacturer's instructions using Taqman Universal PCR Master Mix.

    Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Amplification, Marker

    Expression of CFIm25 and its targets in lung fibroblasts isolated from Col1a1-CreER-CFIm25 fl/fl mice. Four- to six-week-old Col1a1-CreER-CFIm25 fl/fl mice and age- and sex-matched littermate controls were administrated 75 mg/kg (i.p.) tamoxifen daily for 5 days to induce Cre expression. n = 4 biological replications. One week later, fibroblasts were isolated from the lungs of these mice. ( A ) Western blotting was used to confirm the expression of CFIm25, Cre, and CFIm25 targets in fibroblasts, and ( B ) qRT-PCR was performed to determine the dPAS usage of CFIm25 targets ( Col1a1, Tgfbr1, Fzd2 , and Wnt5a ). n = 5 biological replicates. * P

    Journal: The Journal of Clinical Investigation

    Article Title: Cleavage factor 25 deregulation contributes to pulmonary fibrosis through alternative polyadenylation

    doi: 10.1172/JCI122106

    Figure Lengend Snippet: Expression of CFIm25 and its targets in lung fibroblasts isolated from Col1a1-CreER-CFIm25 fl/fl mice. Four- to six-week-old Col1a1-CreER-CFIm25 fl/fl mice and age- and sex-matched littermate controls were administrated 75 mg/kg (i.p.) tamoxifen daily for 5 days to induce Cre expression. n = 4 biological replications. One week later, fibroblasts were isolated from the lungs of these mice. ( A ) Western blotting was used to confirm the expression of CFIm25, Cre, and CFIm25 targets in fibroblasts, and ( B ) qRT-PCR was performed to determine the dPAS usage of CFIm25 targets ( Col1a1, Tgfbr1, Fzd2 , and Wnt5a ). n = 5 biological replicates. * P

    Article Snippet: Western blotting was performed as previously described ( ) using primary rabbit anti-CFIm25 antibodies (Proteintech, 10322-1-AP); rabbit anti–Cre recombinase antibodies (Cell Signaling Technology, 15036S); rabbit anti–TGF-βR1 antibodies (Sigma-Aldrich, SAB1300113); mouse anti–β-actin antibodies (Sigma-Aldrich, A2228); rabbit anti-FZD2 antibodies (Thermo Fisher Scientific, 38-4700); rabbit anti-Wnt5A antibodies (Thermo Fisher Scientific, PA5-23178); rabbit anti-CFIm59 antibodies (Proteintech, A301-360A); rabbit anti-CFIm68 antibodies (Proteintech, A301-356A); and mouse anti-GAPDH antibodies (Life Technologies, Thermo Fisher Scientific, AM4300) and then incubated with the corresponding secondary antibodies conjugated to HRP (Cell Signaling Technology, 7076 or 7074).

    Techniques: Expressing, Isolation, Mouse Assay, Western Blot, Quantitative RT-PCR

    TGF-β and Wnt5A pathways are activated in CFIm25-knockdown fibroblasts. ( A ) The dPAS usage of CFIm25 targets involved in TGF-β ( TGFBR1 ) and Wnt ( WNT5A and FZD2 ) pathways was verified by qRT-PCR. n = 3 biological replicates. * P

    Journal: The Journal of Clinical Investigation

    Article Title: Cleavage factor 25 deregulation contributes to pulmonary fibrosis through alternative polyadenylation

    doi: 10.1172/JCI122106

    Figure Lengend Snippet: TGF-β and Wnt5A pathways are activated in CFIm25-knockdown fibroblasts. ( A ) The dPAS usage of CFIm25 targets involved in TGF-β ( TGFBR1 ) and Wnt ( WNT5A and FZD2 ) pathways was verified by qRT-PCR. n = 3 biological replicates. * P

    Article Snippet: Western blotting was performed as previously described ( ) using primary rabbit anti-CFIm25 antibodies (Proteintech, 10322-1-AP); rabbit anti–Cre recombinase antibodies (Cell Signaling Technology, 15036S); rabbit anti–TGF-βR1 antibodies (Sigma-Aldrich, SAB1300113); mouse anti–β-actin antibodies (Sigma-Aldrich, A2228); rabbit anti-FZD2 antibodies (Thermo Fisher Scientific, 38-4700); rabbit anti-Wnt5A antibodies (Thermo Fisher Scientific, PA5-23178); rabbit anti-CFIm59 antibodies (Proteintech, A301-360A); rabbit anti-CFIm68 antibodies (Proteintech, A301-356A); and mouse anti-GAPDH antibodies (Life Technologies, Thermo Fisher Scientific, AM4300) and then incubated with the corresponding secondary antibodies conjugated to HRP (Cell Signaling Technology, 7076 or 7074).

    Techniques: Quantitative RT-PCR

    CFIm25 overexpression results in 3′-UTR lengthening and decreased protein expression of target genes. Primary fibroblasts isolated from healthy and IPF lungs were electroporated with empty or CFIm25-overexpressing pCDNA3.1 plasmids. Two or three days after transfection, cells were collected for ( A ) Western blotting to determine the protein levels of CFIm25 and its target genes and ( B ) qRT-PCR to determine the dPAS usage of the CFIm25 targets COL1A1, TGFBR1, WNT5A , and FZD2 . n = 3 biological replicates. * P

    Journal: The Journal of Clinical Investigation

    Article Title: Cleavage factor 25 deregulation contributes to pulmonary fibrosis through alternative polyadenylation

    doi: 10.1172/JCI122106

    Figure Lengend Snippet: CFIm25 overexpression results in 3′-UTR lengthening and decreased protein expression of target genes. Primary fibroblasts isolated from healthy and IPF lungs were electroporated with empty or CFIm25-overexpressing pCDNA3.1 plasmids. Two or three days after transfection, cells were collected for ( A ) Western blotting to determine the protein levels of CFIm25 and its target genes and ( B ) qRT-PCR to determine the dPAS usage of the CFIm25 targets COL1A1, TGFBR1, WNT5A , and FZD2 . n = 3 biological replicates. * P

    Article Snippet: Western blotting was performed as previously described ( ) using primary rabbit anti-CFIm25 antibodies (Proteintech, 10322-1-AP); rabbit anti–Cre recombinase antibodies (Cell Signaling Technology, 15036S); rabbit anti–TGF-βR1 antibodies (Sigma-Aldrich, SAB1300113); mouse anti–β-actin antibodies (Sigma-Aldrich, A2228); rabbit anti-FZD2 antibodies (Thermo Fisher Scientific, 38-4700); rabbit anti-Wnt5A antibodies (Thermo Fisher Scientific, PA5-23178); rabbit anti-CFIm59 antibodies (Proteintech, A301-360A); rabbit anti-CFIm68 antibodies (Proteintech, A301-356A); and mouse anti-GAPDH antibodies (Life Technologies, Thermo Fisher Scientific, AM4300) and then incubated with the corresponding secondary antibodies conjugated to HRP (Cell Signaling Technology, 7076 or 7074).

    Techniques: Over Expression, Expressing, Isolation, Transfection, Western Blot, Quantitative RT-PCR

    FZD2 blockade interferes with canonical and non-canonical signaling in NB tissue A. mRNA expression of SK-N-AS and SK-N-DZ human NB tumor tissue treated with FZD2 siRNA (FZD2si) or scrambled siRNA (Co) on day 24. Graphs show the results of qRT-PCR for human FZD2, Wnt3a, Wnt5a, MYC and cyclin D1 as well as for murine CD31. Data are mean ± SD (n = 8 per group). B. Representative Western blot images and C. quantification of immunoblots stained with FZD2, phospho-LRP6 (p-LRP6), total β-catenin, active β-catenin, MYC, cyclin D1, pan-phospho-PKC, (p-PKC), total JNK, phospho-JNK (p-JNK), total AKT, phospho-AKT (p-AKT), total ERK, phospho-ERK (p-ERK) antibodies. FZD2 blockade resulted in decreased levels of FZD2, p-LRP6, active β-catenin, MYC and cyclin D1 associated with increased levels of p-PKC, p-AKT and p-ERK in NB tissue. Data are mean ± SD (n = 8 per group). Asterisks (*) indicate P

    Journal: Oncotarget

    Article Title: Frizzled2 signaling regulates growth of high-risk neuroblastomas by interfering with β-catenin-dependent and β-catenin-independent signaling pathways

    doi: 10.18632/oncotarget.10070

    Figure Lengend Snippet: FZD2 blockade interferes with canonical and non-canonical signaling in NB tissue A. mRNA expression of SK-N-AS and SK-N-DZ human NB tumor tissue treated with FZD2 siRNA (FZD2si) or scrambled siRNA (Co) on day 24. Graphs show the results of qRT-PCR for human FZD2, Wnt3a, Wnt5a, MYC and cyclin D1 as well as for murine CD31. Data are mean ± SD (n = 8 per group). B. Representative Western blot images and C. quantification of immunoblots stained with FZD2, phospho-LRP6 (p-LRP6), total β-catenin, active β-catenin, MYC, cyclin D1, pan-phospho-PKC, (p-PKC), total JNK, phospho-JNK (p-JNK), total AKT, phospho-AKT (p-AKT), total ERK, phospho-ERK (p-ERK) antibodies. FZD2 blockade resulted in decreased levels of FZD2, p-LRP6, active β-catenin, MYC and cyclin D1 associated with increased levels of p-PKC, p-AKT and p-ERK in NB tissue. Data are mean ± SD (n = 8 per group). Asterisks (*) indicate P

    Article Snippet: The blots were probed with antibodies against FZD2 (Thermo Fisher Scientific, Waltham, MA), phospho-LRP6 (Ser1490) (Cell Signaling Technology, Danvers, MA), β-catenin (Abcam, Cambridge, UK), non-phospho β-catenin (Ser33/37/Thr41), MYC, cyclin D1, phospho-PKC (pan) (βII Ser660), JNK, phospho-JNK (Thr183/Tyr185), AKT, phospho-AKT (Ser473), ERK1/2 and phospho-44/42 ERK1/2(Thr202/Tyr204) (Cell Signaling Technology) before incubation with horseradish peroxidase–conjugated secondary antibodies (GE Healthcare).

    Techniques: Expressing, Quantitative RT-PCR, Western Blot, Staining

    Wnt3a and Wnt5a signal via FZD2 in NB cell lines Representative Western blot images and quantification of immunoblots. SK-N-AS A. and SK-N-DZ B. NB cells were left untreated (Control; Co) or treated with Wnt3a (3a) and Wnt5a (5a) for 15 min, with or without FZD2 siRNA (si) pretreatment. Nonspecific scrambled siRNA (scr si, scrCo) served as an additional control. Then cells were harvested to analyze the levels of FZD2, phospho-LRP6 (LRP6 phosphorylated at Ser1490; p-LRP6), total β-catenin, active β-catenin (non-phospho β-catenin; Ser33/37/Thr41), MYC, cyclin D1, pan-phospho-PKC, (βII Ser660); p-PKC), total JNK, phospho-JNK (JNK phosphorylated at Thr183/Tyr185; p-JNK), total AKT, phospho-AKT (AKT phosphorylated at Ser473serine-473; p-AKT), total ERK and phospho-ERK (ERK1/2 phosphorylated at Thr202/Tyr204; p-ERK) by Western blotting. Activation levels of signaling factors associated with β-catenin-dependent signaling pathway (p-LRP6, active β-catenin) and β-catenin target genes (MYC, cyclin D1) were reduced below baseline levels in Wnt stimulated and unstimulated NB cells pretreated with FZD2 siRNA in contrast to activation levels of signaling factors associated with β-catenin-independent signaling (p-PKC, p-AKT, p-JNK, p-ERK). Graphs represent the mean of 3 independent replicates ± SD. Asterisks (*) indicate P

    Journal: Oncotarget

    Article Title: Frizzled2 signaling regulates growth of high-risk neuroblastomas by interfering with β-catenin-dependent and β-catenin-independent signaling pathways

    doi: 10.18632/oncotarget.10070

    Figure Lengend Snippet: Wnt3a and Wnt5a signal via FZD2 in NB cell lines Representative Western blot images and quantification of immunoblots. SK-N-AS A. and SK-N-DZ B. NB cells were left untreated (Control; Co) or treated with Wnt3a (3a) and Wnt5a (5a) for 15 min, with or without FZD2 siRNA (si) pretreatment. Nonspecific scrambled siRNA (scr si, scrCo) served as an additional control. Then cells were harvested to analyze the levels of FZD2, phospho-LRP6 (LRP6 phosphorylated at Ser1490; p-LRP6), total β-catenin, active β-catenin (non-phospho β-catenin; Ser33/37/Thr41), MYC, cyclin D1, pan-phospho-PKC, (βII Ser660); p-PKC), total JNK, phospho-JNK (JNK phosphorylated at Thr183/Tyr185; p-JNK), total AKT, phospho-AKT (AKT phosphorylated at Ser473serine-473; p-AKT), total ERK and phospho-ERK (ERK1/2 phosphorylated at Thr202/Tyr204; p-ERK) by Western blotting. Activation levels of signaling factors associated with β-catenin-dependent signaling pathway (p-LRP6, active β-catenin) and β-catenin target genes (MYC, cyclin D1) were reduced below baseline levels in Wnt stimulated and unstimulated NB cells pretreated with FZD2 siRNA in contrast to activation levels of signaling factors associated with β-catenin-independent signaling (p-PKC, p-AKT, p-JNK, p-ERK). Graphs represent the mean of 3 independent replicates ± SD. Asterisks (*) indicate P

    Article Snippet: The blots were probed with antibodies against FZD2 (Thermo Fisher Scientific, Waltham, MA), phospho-LRP6 (Ser1490) (Cell Signaling Technology, Danvers, MA), β-catenin (Abcam, Cambridge, UK), non-phospho β-catenin (Ser33/37/Thr41), MYC, cyclin D1, phospho-PKC (pan) (βII Ser660), JNK, phospho-JNK (Thr183/Tyr185), AKT, phospho-AKT (Ser473), ERK1/2 and phospho-44/42 ERK1/2(Thr202/Tyr204) (Cell Signaling Technology) before incubation with horseradish peroxidase–conjugated secondary antibodies (GE Healthcare).

    Techniques: Western Blot, Activation Assay

    Effects of Wnt3a and Wnt5a stimulation and FZD2 blockade on cell proliferation, migration and Rac1 activity in NB cells A. Relative density of cancer cells up to 72 h following stimulation with 100 ng/ml recombinant Wnt3a, Wnt5a or pretreatment with FZD2 siRNA (FZD2si) or nonspecific scrambled siRNA (scrCo) was measured using the WST-1 cell proliferation assay. FZD2 siRNA suppressed NB cell proliferation. Graphs represent the mean of 3 independent experiments ± SD. Asterisks (*) indicate P

    Journal: Oncotarget

    Article Title: Frizzled2 signaling regulates growth of high-risk neuroblastomas by interfering with β-catenin-dependent and β-catenin-independent signaling pathways

    doi: 10.18632/oncotarget.10070

    Figure Lengend Snippet: Effects of Wnt3a and Wnt5a stimulation and FZD2 blockade on cell proliferation, migration and Rac1 activity in NB cells A. Relative density of cancer cells up to 72 h following stimulation with 100 ng/ml recombinant Wnt3a, Wnt5a or pretreatment with FZD2 siRNA (FZD2si) or nonspecific scrambled siRNA (scrCo) was measured using the WST-1 cell proliferation assay. FZD2 siRNA suppressed NB cell proliferation. Graphs represent the mean of 3 independent experiments ± SD. Asterisks (*) indicate P

    Article Snippet: The blots were probed with antibodies against FZD2 (Thermo Fisher Scientific, Waltham, MA), phospho-LRP6 (Ser1490) (Cell Signaling Technology, Danvers, MA), β-catenin (Abcam, Cambridge, UK), non-phospho β-catenin (Ser33/37/Thr41), MYC, cyclin D1, phospho-PKC (pan) (βII Ser660), JNK, phospho-JNK (Thr183/Tyr185), AKT, phospho-AKT (Ser473), ERK1/2 and phospho-44/42 ERK1/2(Thr202/Tyr204) (Cell Signaling Technology) before incubation with horseradish peroxidase–conjugated secondary antibodies (GE Healthcare).

    Techniques: Migration, Activity Assay, Recombinant, Proliferation Assay

    FZD-2 knockdown reduces growth of SK-N-AS and SK-N-DZ xenografts A. Tumor volume curves in SK-N-AS and SK-N-DZ tumor xenograft bearing mice treated with FZD2 siRNA (FZD2si) or scrambled siRNA as control (Co). NB cells were injected s.c. into athymic nude mice and tumor growth was monitored in the presence (red shade) or absence (green shade) of siRNA against FZD2 or scrambled siRNA (n = 8 per group). Data are mean ± SD. *, significantly different from control tumors ( P

    Journal: Oncotarget

    Article Title: Frizzled2 signaling regulates growth of high-risk neuroblastomas by interfering with β-catenin-dependent and β-catenin-independent signaling pathways

    doi: 10.18632/oncotarget.10070

    Figure Lengend Snippet: FZD-2 knockdown reduces growth of SK-N-AS and SK-N-DZ xenografts A. Tumor volume curves in SK-N-AS and SK-N-DZ tumor xenograft bearing mice treated with FZD2 siRNA (FZD2si) or scrambled siRNA as control (Co). NB cells were injected s.c. into athymic nude mice and tumor growth was monitored in the presence (red shade) or absence (green shade) of siRNA against FZD2 or scrambled siRNA (n = 8 per group). Data are mean ± SD. *, significantly different from control tumors ( P

    Article Snippet: The blots were probed with antibodies against FZD2 (Thermo Fisher Scientific, Waltham, MA), phospho-LRP6 (Ser1490) (Cell Signaling Technology, Danvers, MA), β-catenin (Abcam, Cambridge, UK), non-phospho β-catenin (Ser33/37/Thr41), MYC, cyclin D1, phospho-PKC (pan) (βII Ser660), JNK, phospho-JNK (Thr183/Tyr185), AKT, phospho-AKT (Ser473), ERK1/2 and phospho-44/42 ERK1/2(Thr202/Tyr204) (Cell Signaling Technology) before incubation with horseradish peroxidase–conjugated secondary antibodies (GE Healthcare).

    Techniques: Mouse Assay, Injection

    Proposed mechanism by which FZD2 blockade affects NB cell behavior Simplified schematic view of β-catenin-dependent and -independent signaling pathways and the crosstalk between each other. Blue boxes indicate inhibited signaling factors and actions, red boxes depict activated signaling factors and actions following FZD2 blockade, which were revealed in this study. Wnt3a activates the β-catenin pathway by interacting with FZD2 and LRP6. When Wnt5a acts on FZD2 it may compete with Wnt3a, thereby inhibiting the β-catenin pathway. However, Wnt5a activates the Rac/JNK pathways. Downregulation of FZD2 inhibits the β-catenin pathway and the Rac/JNK pathways in NB cells. Blocking the action of Wnt5a on these pathways by blocking FZD2 could promote the activation of other β-catenin-independent Wnt pathways. Consequently, increased availability of Wnt5a protein may interact with receptor ROR1/2 or FZD to activate the PI3K/AKT and PKC pathway, leading to the activation of target genes, which normally suppress proliferation and migration. These actions, perhaps along with modulation of the activity of other non-canonical pathways (e.g. Rac/JNK), mediate the anti-proliferative and anti-migratory effects of Wnts following FZD2 blockade. In addition, Wnts acting via FZD and growth factor receptors also activate ERK. It is unclear, whether Wnt3a may also activate a β-catenin-independent pathway and/or contributes to ERK-activation. Phosphorylation of ERK alone or alteration of the p-ERK/p-AKT ratio in turn may regulate proliferation depending on the signaling strength thus counteracting or supporting the anti-proliferative effects of FZD2 blockade.

    Journal: Oncotarget

    Article Title: Frizzled2 signaling regulates growth of high-risk neuroblastomas by interfering with β-catenin-dependent and β-catenin-independent signaling pathways

    doi: 10.18632/oncotarget.10070

    Figure Lengend Snippet: Proposed mechanism by which FZD2 blockade affects NB cell behavior Simplified schematic view of β-catenin-dependent and -independent signaling pathways and the crosstalk between each other. Blue boxes indicate inhibited signaling factors and actions, red boxes depict activated signaling factors and actions following FZD2 blockade, which were revealed in this study. Wnt3a activates the β-catenin pathway by interacting with FZD2 and LRP6. When Wnt5a acts on FZD2 it may compete with Wnt3a, thereby inhibiting the β-catenin pathway. However, Wnt5a activates the Rac/JNK pathways. Downregulation of FZD2 inhibits the β-catenin pathway and the Rac/JNK pathways in NB cells. Blocking the action of Wnt5a on these pathways by blocking FZD2 could promote the activation of other β-catenin-independent Wnt pathways. Consequently, increased availability of Wnt5a protein may interact with receptor ROR1/2 or FZD to activate the PI3K/AKT and PKC pathway, leading to the activation of target genes, which normally suppress proliferation and migration. These actions, perhaps along with modulation of the activity of other non-canonical pathways (e.g. Rac/JNK), mediate the anti-proliferative and anti-migratory effects of Wnts following FZD2 blockade. In addition, Wnts acting via FZD and growth factor receptors also activate ERK. It is unclear, whether Wnt3a may also activate a β-catenin-independent pathway and/or contributes to ERK-activation. Phosphorylation of ERK alone or alteration of the p-ERK/p-AKT ratio in turn may regulate proliferation depending on the signaling strength thus counteracting or supporting the anti-proliferative effects of FZD2 blockade.

    Article Snippet: The blots were probed with antibodies against FZD2 (Thermo Fisher Scientific, Waltham, MA), phospho-LRP6 (Ser1490) (Cell Signaling Technology, Danvers, MA), β-catenin (Abcam, Cambridge, UK), non-phospho β-catenin (Ser33/37/Thr41), MYC, cyclin D1, phospho-PKC (pan) (βII Ser660), JNK, phospho-JNK (Thr183/Tyr185), AKT, phospho-AKT (Ser473), ERK1/2 and phospho-44/42 ERK1/2(Thr202/Tyr204) (Cell Signaling Technology) before incubation with horseradish peroxidase–conjugated secondary antibodies (GE Healthcare).

    Techniques: Blocking Assay, Activation Assay, Migration, Activity Assay

    Characterization of gene expression in MYCN -unamplified SK-N-AS and MYCN -amplified SK-N-DZ NB cells A. Graphs show the results of qRT-PCR for FZD2, Wnt3a, Wnt5a, MYC and cyclin D1 performed on RNA from human SK-N-AS (AS) and SK-N-DZ (DZ) NB cells. FZD2 , Wnt3a , MYC and cyclin D1 mRNA expression is different between SK-N-AS and SK-N-DZ cells. B. Representative Western blot images and C. quantitative determination of protein expression in SK-N-AS and SK-N-DZ cells. Untreated cells were harvested to analyze the levels of FZD2, phospho-LRP6 (LRP6 phosphorylated at Ser1490; p-LRP6), total β-catenin, active β-catenin (non-phospho β-catenin; Ser33/37/Thr41), MYC, cyclin D1, pan-phospho-PKC, (βII Ser660; p-PKC), total JNK, phospho-JNK (JNK phosphorylated at Thr183/Tyr185; p-JNK), total AKT, phospho-AKT (AKT phosphorylated at Ser473; p-AKT), total ERK and phospho-ERK (ERK1/2 phosphorylated at Thr202/Tyr204; p-ERK) by Western blotting. Comparison of protein expression profiles between SK-N-AS and SK-N-DZ cells revealed differences in basal expression and activation levels of investigated signaling proteins. Graphs represent the mean of 3 independent experiments ± SD (* P

    Journal: Oncotarget

    Article Title: Frizzled2 signaling regulates growth of high-risk neuroblastomas by interfering with β-catenin-dependent and β-catenin-independent signaling pathways

    doi: 10.18632/oncotarget.10070

    Figure Lengend Snippet: Characterization of gene expression in MYCN -unamplified SK-N-AS and MYCN -amplified SK-N-DZ NB cells A. Graphs show the results of qRT-PCR for FZD2, Wnt3a, Wnt5a, MYC and cyclin D1 performed on RNA from human SK-N-AS (AS) and SK-N-DZ (DZ) NB cells. FZD2 , Wnt3a , MYC and cyclin D1 mRNA expression is different between SK-N-AS and SK-N-DZ cells. B. Representative Western blot images and C. quantitative determination of protein expression in SK-N-AS and SK-N-DZ cells. Untreated cells were harvested to analyze the levels of FZD2, phospho-LRP6 (LRP6 phosphorylated at Ser1490; p-LRP6), total β-catenin, active β-catenin (non-phospho β-catenin; Ser33/37/Thr41), MYC, cyclin D1, pan-phospho-PKC, (βII Ser660; p-PKC), total JNK, phospho-JNK (JNK phosphorylated at Thr183/Tyr185; p-JNK), total AKT, phospho-AKT (AKT phosphorylated at Ser473; p-AKT), total ERK and phospho-ERK (ERK1/2 phosphorylated at Thr202/Tyr204; p-ERK) by Western blotting. Comparison of protein expression profiles between SK-N-AS and SK-N-DZ cells revealed differences in basal expression and activation levels of investigated signaling proteins. Graphs represent the mean of 3 independent experiments ± SD (* P

    Article Snippet: The blots were probed with antibodies against FZD2 (Thermo Fisher Scientific, Waltham, MA), phospho-LRP6 (Ser1490) (Cell Signaling Technology, Danvers, MA), β-catenin (Abcam, Cambridge, UK), non-phospho β-catenin (Ser33/37/Thr41), MYC, cyclin D1, phospho-PKC (pan) (βII Ser660), JNK, phospho-JNK (Thr183/Tyr185), AKT, phospho-AKT (Ser473), ERK1/2 and phospho-44/42 ERK1/2(Thr202/Tyr204) (Cell Signaling Technology) before incubation with horseradish peroxidase–conjugated secondary antibodies (GE Healthcare).

    Techniques: Expressing, Amplification, Quantitative RT-PCR, Western Blot, Activation Assay