recombinant mouse wnt2b  (R&D Systems)

Journal: Oncology Reports

Article Title: Wnt2b and Wnt5a expression is highly associated with M2 TAMs in non-small cell lung cancer

doi: 10.3892/or.2022.8404

Figure Lengend Snippet: Immunostaining of lung cancer. Carcinoma with (A) positive Wnt2b expression in tumor cells and (B) high density of M2 TAMs. Carcinoma with (C) positive Wnt2b expression in stromal cells and (D) high density of M2 TAMs. Carcinoma with (E) negative Wnt2b expression in tumor and stromal cells and (F) low density of M2 TAMs. Carcinoma with (G) positive Wnt5a expression in tumor and weak Wnt5a expression in stromal cells and (H) high density of M2 TAMs. Carcinoma with (I) high and (J) low density of M1 TAMs. TAM, tumor-associated macrophage.

Article Snippet: The following antibodies were prepared: Mouse monoclonal anti-human Wnt2b (clone C-2; 1:30; Santa Cruz Biotechnology, Inc.), Wnt5a (clone 3A4; 1:100; Sigma-Aldrich; Merck KGaA) and CD163 (clone 760-4437; prediluted; Ventana Medical Systems; Roche Diagnostics) and rabbit polyclonal anti-human iNOS (cat. no. ab3523; 1:50; Abcam) and monoclonal anti-human Ki-67 (clone 30-9; prediluted; Ventana Medical Systems; Roche Diagnostics).

Techniques: Immunostaining, Expressing

Journal: Oncology Reports

Article Title: Wnt2b and Wnt5a expression is highly associated with M2 TAMs in non-small cell lung cancer

doi: 10.3892/or.2022.8404

Figure Lengend Snippet: Wnt2b expression in patients with non-small cell lung cancer.

Article Snippet: The following antibodies were prepared: Mouse monoclonal anti-human Wnt2b (clone C-2; 1:30; Santa Cruz Biotechnology, Inc.), Wnt5a (clone 3A4; 1:100; Sigma-Aldrich; Merck KGaA) and CD163 (clone 760-4437; prediluted; Ventana Medical Systems; Roche Diagnostics) and rabbit polyclonal anti-human iNOS (cat. no. ab3523; 1:50; Abcam) and monoclonal anti-human Ki-67 (clone 30-9; prediluted; Ventana Medical Systems; Roche Diagnostics).

Techniques: Expressing

Journal: Oncology Reports

Article Title: Wnt2b and Wnt5a expression is highly associated with M2 TAMs in non-small cell lung cancer

doi: 10.3892/or.2022.8404

Figure Lengend Snippet: M2 TAM density and stromal M1 TAM density in relation to Wnt status. M2 TAM density in relation to (A) tumoral and (B) stromal Wnt2b and (C) tumoral and (D) stromal Wnt5a status. Stromal M1 TAM density in relation to (E) tumoral and (F) stromal Wnt2b and (G) tumoral and (H) stromal Wnt5a status. TAM, tumor-associated macrophage.

Article Snippet: The following antibodies were prepared: Mouse monoclonal anti-human Wnt2b (clone C-2; 1:30; Santa Cruz Biotechnology, Inc.), Wnt5a (clone 3A4; 1:100; Sigma-Aldrich; Merck KGaA) and CD163 (clone 760-4437; prediluted; Ventana Medical Systems; Roche Diagnostics) and rabbit polyclonal anti-human iNOS (cat. no. ab3523; 1:50; Abcam) and monoclonal anti-human Ki-67 (clone 30-9; prediluted; Ventana Medical Systems; Roche Diagnostics).

Techniques:

Journal: Oncology Reports

Article Title: Wnt2b and Wnt5a expression is highly associated with M2 TAMs in non-small cell lung cancer

doi: 10.3892/or.2022.8404

Figure Lengend Snippet: Ki-67 proliferation index in relation to Wnt status. (A) tumoral and (B) stromal Wnt2b and (C) tumoral and (D) stromal Wnt5a status.

Article Snippet: The following antibodies were prepared: Mouse monoclonal anti-human Wnt2b (clone C-2; 1:30; Santa Cruz Biotechnology, Inc.), Wnt5a (clone 3A4; 1:100; Sigma-Aldrich; Merck KGaA) and CD163 (clone 760-4437; prediluted; Ventana Medical Systems; Roche Diagnostics) and rabbit polyclonal anti-human iNOS (cat. no. ab3523; 1:50; Abcam) and monoclonal anti-human Ki-67 (clone 30-9; prediluted; Ventana Medical Systems; Roche Diagnostics).

Techniques:

Journal: Oncology Reports

Article Title: Wnt2b and Wnt5a expression is highly associated with M2 TAMs in non-small cell lung cancer

doi: 10.3892/or.2022.8404

Figure Lengend Snippet: Overall survival of patients with resected NSCLC. (A) in relation to Wnt2b status. (B) in relation to Wnt5a status. (C) in relation to M2 TAM status. TAM, tumor-associated macrophage.

Article Snippet: The following antibodies were prepared: Mouse monoclonal anti-human Wnt2b (clone C-2; 1:30; Santa Cruz Biotechnology, Inc.), Wnt5a (clone 3A4; 1:100; Sigma-Aldrich; Merck KGaA) and CD163 (clone 760-4437; prediluted; Ventana Medical Systems; Roche Diagnostics) and rabbit polyclonal anti-human iNOS (cat. no. ab3523; 1:50; Abcam) and monoclonal anti-human Ki-67 (clone 30-9; prediluted; Ventana Medical Systems; Roche Diagnostics).

Techniques:

Journal: Frontiers in Cell and Developmental Biology

Article Title: Intravenous Transplantation of Human Hair Follicle-Derived Mesenchymal Stem Cells Ameliorates Trabecular Bone Loss in Osteoporotic Mice

doi: 10.3389/fcell.2022.814949

Figure Lengend Snippet: Expression of bone metabolism regulators in serum and bone marrow in OVX-induced osteoporotic mice with intravenous transplantation of hHF-MSCs. (A) Representative image of antibody array against bone metabolism regulators in peripheral blood serum from OVX-induced osteoporotic mice with high-dose hHF-MSCs injection. (B) The levels of bone metabolism regulators in peripheral blood serum from OVX-induced osteoporotic mice with high-dose hHF-MSCs injection determined by antibody array. ( n = 12 for each group). (C) OPG, VCAM-1, RANKL, Noggin, and Wnt2b levels in femur bone marrow of OVX-induced osteoporotic mice in response to high-dose hHF-MSCs injection determined by immunohistochemistry. All the data were obtained from three independent experiments. Data were shown as the means ± s.e.m. *: p < 0.05, **: p < 0.01, ***: p < 0.001, NS: not significant.

Article Snippet: After treatment with Protein Block (Dako Cytomation, Glostrup, Denmark), sections were incubated with anti-mouse OPG (Thermo Fisher Scientific, Ann Arbor, MI, United States), anti-mouse VCAM-1 (Cell Signaling Technology, Danvers, MA, United States), anti-mouse RANKL (Thermo Fisher Scientific), anti-mouse Noggin (Thermo Fisher Scientific), and anti-mouse Wnt2b (Thermo Fisher Scientific) overnight at 4°C.

Techniques: Expressing, Transplantation Assay, Ab Array, Injection, Immunohistochemistry

Journal: Scientific Reports

Article Title: Wnt2b attenuates HSCs activation and liver fibrosis through negative regulating TLR4 signaling

doi: 10.1038/s41598-017-04374-5

Figure Lengend Snippet: Wnt2b is elevated in hepatic fibrosis. ( a ) Representative Sirius Red staining, IHC staining of Wnt2b in liver tissue microarrays containing healthy controls (n = 9, with a mean age of 43.3 ± 1.4 years) and patients with fibrosis (n = 10, with a mean age of 48.8 ± 2.9 years). Hepatic fibrosis mouse model was induced by TAA, and then the following analyses were performed. ( b ) H&E, Sirius Red staining. ( c ) Immunofluorescence staining of α-SMA and IHC staining of Wnt2b. ( d ) RT-PCR (upper) and Western blotting (lower) of Wnt2b. Cropped blots are displayed; Full-length blots are presented in Supplementary Fig. . ( e ) ELISA analysis of Wnt2b in liver homogenate. Statistical analyses provided the mean ± SE (n = 8/group); * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The shRNA plasmid targeting mouse Wnt2b was purchased from OriGene (Gene ID 22414, Beijing, China).

Techniques: Staining, Immunohistochemistry, Immunofluorescence, Reverse Transcription Polymerase Chain Reaction, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Scientific Reports

Article Title: Wnt2b attenuates HSCs activation and liver fibrosis through negative regulating TLR4 signaling

doi: 10.1038/s41598-017-04374-5

Figure Lengend Snippet: Wnt2b is mainly produced by hepatocytes during liver fibrogenesis. ( a ) mRNA and protein levels of Wnt2b in hepatocytes (left) and NPCs (right) from control and TAA-treated mice, respectively. ( b ) mRNA (upper) and protein (lower) levels of Wnt2b in primary mouse hepatocytes treated with CCl 4 in vitro . ( c ) Immunofluorescence staining of Wnt2b in HSCs from control and TAA-treated mice. ( d ) RT-PCR analysis of Wnt2b in cultured HSCs from naive mice at the indicated points of time. Mouse embryonic tissues served as positive controls. Cropped blots are displayed; Full-length blots are presented in Supplementary Fig. . Data are mean ± SEM of three independent experiments; * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The shRNA plasmid targeting mouse Wnt2b was purchased from OriGene (Gene ID 22414, Beijing, China).

Techniques: Produced, Control, In Vitro, Immunofluorescence, Staining, Reverse Transcription Polymerase Chain Reaction, Cell Culture

Journal: Scientific Reports

Article Title: Wnt2b attenuates HSCs activation and liver fibrosis through negative regulating TLR4 signaling

doi: 10.1038/s41598-017-04374-5

Figure Lengend Snippet: Wnt2b protects against liver fibrosis. ( a ) The schedule for Wnt2b knockdown with shRNA targeting mouse Wnt2b (sh-Wnt2b) or Wnt2b-overexpression with plasmid pRK5-mWnt2b (Over-Wnt2b) under TAA treatment. ( b ) The effects of sh-Wnt2b (left) and pRK5-mWnt2b (right) in TAA-challenged mice livers. ( c ) H&E and Sirius Red staining, Western blotting of Collagen-I, ( d ) Western blotting (upper) and immunofluorescence staining of α-SMA (lower) for liver tissues from mice treated with sh-Wnt2b or control vector. ( e ) H&E and Sirius Red staining, Western blotting of Collagen-I, ( f ) Western blotting (upper) and immunofluorescence staining of α-SMA (lower) for liver tissues from mice treated with pRK5-mWnt2b construct or control vector. Cropped blots are displayed; Full-length blots are presented in Supplementary Fig. . Statistical analysis are mean ± SE (n = 6/group); * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The shRNA plasmid targeting mouse Wnt2b was purchased from OriGene (Gene ID 22414, Beijing, China).

Techniques: Knockdown, shRNA, Over Expression, Plasmid Preparation, Staining, Western Blot, Immunofluorescence, Control, Construct

Journal: Scientific Reports

Article Title: Wnt2b attenuates HSCs activation and liver fibrosis through negative regulating TLR4 signaling

doi: 10.1038/s41598-017-04374-5

Figure Lengend Snippet: Wnt2b exerts a direct inhibitory effect on HSCs activation. ( a ) RT-PCR analysis of Fzds and LRP5/6 in cultured HSCs from naive mice at the indicated points of time. Mouse embryonic tissues served as positive controls. ( b , c ) mRNA and protein levels of α-SMA and Collagen-I in LX2 cells cultured in conditioned medium (CM) collected from HEK293 cells transfected with active Wnt2B-V5 (CM-Wnt2b) or control plasmid (CM-Control) ( b ), as well as in LX2 cells transfected with active Wnt2B-V5 or control plasmid ( c ). Cropped blots are displayed; Full-length blots are presented in Supplementary Fig. . Data are mean ± SEM of three independent experiments; * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The shRNA plasmid targeting mouse Wnt2b was purchased from OriGene (Gene ID 22414, Beijing, China).

Techniques: Activation Assay, Reverse Transcription Polymerase Chain Reaction, Cell Culture, Transfection, Control, Plasmid Preparation

Journal: Scientific Reports

Article Title: Wnt2b attenuates HSCs activation and liver fibrosis through negative regulating TLR4 signaling

doi: 10.1038/s41598-017-04374-5

Figure Lengend Snippet: Wnt2b suppresses TLR4 activation-mediated pro-fibrogenic effects. ( a ) Representative images for bacterial growth of jejunum (left) and liver tissues (right) after cultivation on Blood Agar Plates. ( b ) Representative H&E (upper) and Sirius Red staining (lower) of liver tissues from WT mice and TLR4 −/− mice after 12 intraperitoneal injections of TAA. ( c ) H&E and Sirius Red staining,Western blotting of α-SMA in liver tissues from mice treated with TAA alone, or combined with sh-Wnt2b construct/TLR4 inhibitor TAK242 (4 mg/kg, i.p.), or all of the three factors given above in combination for 4 weeks. ( d ) Protein levels of α-SMA and Collagen-I in LX2 cells stimulated with LPS (10, 100 ng/ml) for 24 h. ( e ) Effects of Wnt2b on the α-SMA and Collagen-I expressions in LX2 cells stimulated with LPS (100 ng/ml). ( f ) Protein levels of α-SMA and Collagen-I in LX2 cells stimulated with LPS (100 ng/ml) or vehicle for 24 h, and TGF-β (500 pg/ml) or vehicle for an additional 48 h. ( g ) LX2 cells were transfected with active Wnt2B-V5 or control plasmids for 24 h, followed by treatment with LPS ± TGF-β as described in Fig. 5f. The expression of α-SMA and Collagen-I were then detected by Western blot analysis. Cropped blots are displayed; Full-length blots are presented in Supplementary Fig. . Statistical analysis provided the mean ± SE (n = 6/group), * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The shRNA plasmid targeting mouse Wnt2b was purchased from OriGene (Gene ID 22414, Beijing, China).

Techniques: Activation Assay, Staining, Western Blot, Construct, Transfection, Control, Expressing

Journal: Scientific Reports

Article Title: Wnt2b attenuates HSCs activation and liver fibrosis through negative regulating TLR4 signaling

doi: 10.1038/s41598-017-04374-5

Figure Lengend Snippet: Wnt2b disturbs the TLR4 signaling transduction. ( a , b ) Protein levels of TLR4, p-NF-κB p65 (Ser536) and NF-κB p65 (upper), and mRNA levels of RELA and TNF (lower) in LX2 cells cultured in Wnt2b-CM ( a ), as well as in LX2 cells transfected with active Wnt2B-V5 or control plasmids ( b ). ( c , d ) Western blotting of the phosphorylation of MAPKs in LX2 cells cultured in Wnt2b-CM ( c) , and in LX2 cells transfected with active Wnt2B-V5 or control plasmids ( d ). ( e,f ) Protein levels of TLR4, p-NF-κB p65 (Ser536) and NF-κB p65 ( e ), and mRNA levels of RELA and TNF ( f ) in liver tissues from mice challenged with TAA combined with HD injection of pRK5-mWnt2b/sh-Wnt2b construct or control vector as depicted in Fig. . ( g ) Immunofluorescence staining of NF-κB p65 in HSCs isolated from mice challenged with TAA combined with HD injection of pRK5-mWnt2b (upper) / sh-Wnt2b (lower) construct or control vector as depicted in Fig. . Cropped blots are displayed; Full-length blots are presented in Supplementary Fig. . Statistical analysis provided the mean ± SE (n = 6/group), * P < 0.05, ** P < 0.01, *** P < 0.001.

Article Snippet: The shRNA plasmid targeting mouse Wnt2b was purchased from OriGene (Gene ID 22414, Beijing, China).

Techniques: Transduction, Cell Culture, Transfection, Control, Western Blot, Phospho-proteomics, Injection, Construct, Plasmid Preparation, Immunofluorescence, Staining, Isolation

Journal: Scientific Reports

Article Title: Wnt2b attenuates HSCs activation and liver fibrosis through negative regulating TLR4 signaling

doi: 10.1038/s41598-017-04374-5

Figure Lengend Snippet: A schematic model for the inhibitory effects of Wnt2b on HSCs activation and liver fibrosis through negative regulating TLR4 signaling.

Article Snippet: The shRNA plasmid targeting mouse Wnt2b was purchased from OriGene (Gene ID 22414, Beijing, China).

Techniques: Activation Assay

Journal: Cell reports

Article Title: Wnt Ligands Secreted by Subepithelial Mesenchymal Cells Are Essential for the Survival of Intestinal Stem Cells and Gut Homeostasis.

doi: 10.1016/j.celrep.2016.03.088

Figure Lengend Snippet: Figure 3. Wnt2b Ligand Secreted by Extra- epithelial Cells Drives b-Catenin Signaling Outputs and Is Essential for the Mainte- nance of Intestinal Homeostasis (A) (Upper left) Scheme depicting how the intes- tinal organoids were generated. Intestinal crypts were isolated from villin-WlscKO (or control) ani- mals and cultivated in Matrigel as intestinal orga- noids with standard cultivation medium (SCM) or SCM containing either 100 ng/ml Wnt2b or 100 ng/ml Wnt5a. (Right) Whereas organoids from control animals grew normally, it was not possible to establish growing organoids from villin- WlscKO crypts. Addition of Wnt2b changed the morphology of control organoids to large spher- oids (compare with finger-like protrusions in SCM). External Wnt2b enables organoids from villin- WlscKO crypts to be established. Wnt5a promoted survival of villin-WlscKO crypts but did not promote growth. Organoid representing typical shape of organoids (apparent in more than 75% of the or- ganoids) is shown. DAPI (blue) counterstains the nuclei. (Bottom left) The fraction of living organoids after 7 days (d) in culture is shown; the proportion of various shapes of organoids is indicated. Crypts were seeded at the same initial density. Number of control organoids growing in SCM is set as 100%, each column summarizes data from two indepen- dent experiments (three replicates each). Error bars indicate SD. Scale bar, 100 mm. i.p., intra- peritoneal. (B) Scheme of external Wnt2b application regimen. (C) Injected Wnt2b partially restores intestinal expression of Axin2 indicating active Wnt/b-cat- enin signaling. For real-time qRT PCR, y axes show normalized relative mRNA abundance; control levels were set to 1. Error bars indicate SD. (D) Externally delivered Wnt2b preserves intes- tinal morphology and proliferation determined by Ki67 in R26-WlscKO animals. R26-WlscKO indicates Rosa26-CreERT2,Wntlessflox/flox. Immunohisto- chemistry: DAPI marks nuclei, and b-catenin de- notes epithelial cells. Scale bar, 100 mm.

Article Snippet: External mouse Wnt3a (Abcam) or mouse Wnt2b (R&D Systems) was injected intraperitoneally (50 mg/kg) twice a day, starting 12 hr after the first tamoxifen injection.

Techniques: Generated, Isolation, Control, Injection, Expressing, Quantitative RT-PCR, Immunohistochemistry

Journal: Cell reports

Article Title: Wnt Ligands Secreted by Subepithelial Mesenchymal Cells Are Essential for the Survival of Intestinal Stem Cells and Gut Homeostasis.

doi: 10.1016/j.celrep.2016.03.088

Figure Lengend Snippet: Figure 4. Subepithelial Mesenchymal Cells Expressing High Levels of Wnt2b Are Pre- dominantly Gli1 Positive and the Expression of Either Gli1 or Acta2 Marks the Majority of Wnt2b-Secreting Cells (A) Murine duodenum tissue sections were hy- bridized with smFISH probe libraries for Wnt2b (red dots), Lgr5 (green dots), and Gli1 (cyan dots) single-mRNA molecules. Nuclei were stained with DAPI (blue), and E-cadherin protein was stained with a FITC-coupled antibody (gray) to visualize cell membranes of epithelial cells. Paneth cell granules exhibit non-specific fluorescence ap- pearing in multiple channels. Scale bar, 100 mm. (B) smFISH of duodenum probed for Wnt2b (red dots), Gli1 (green dots), and Acta2 (aSMA) (cyan dots) single-mRNA molecules. The shape of the epithelial cells is visualized by E-cadherin staining; DAPI marks nuclei (similarly as in A). Scale bar, 100 mm. (C) Venn diagram showing overlap of Wnt2b, Gli1, and Acta2 (aSMA). Single-crypt-associated stro- mal cells were stratified into groups of expressing (>0 molecules per cubic micron) or non-expressing cells (0 molecules per cubic micron) for each RNA of interest according to smFISH analyses. The probability of random co-expression was as- sessed with the hypergeometric enrichment test. (D and E) High expression levels of Wnt2b correlate with Gli1 positivity but not with expression of Acta2 (aSMA). Wnt2b-expressing subepithelial cells were divided into two groups based on their expression of Gli1 (Gli1+ versus Gli1) (D) or Acta2 (aSMA; Acta2+ versus Acta2) (E). Cells express- ing high levels of Wnt2b (more Wnt2b-RNA mole- cules per cubic micron) are Gli1 positive. No significant difference based on Acta2 (aSMA) expression was identified. The y axes show numbers of Wnt2b-RNA molecules per cubic micron. Data shown as Tukey-style boxplots.

Article Snippet: External mouse Wnt3a (Abcam) or mouse Wnt2b (R&D Systems) was injected intraperitoneally (50 mg/kg) twice a day, starting 12 hr after the first tamoxifen injection.

Techniques: Expressing, Staining