Anti-Phospho-Smad3 (S423 + S425) Rabbit Monoclonal Antibody

All lanes use the Antibody at 1:1K dilution for 1 hour at room temperature.

All lanes use the Antibody at 1:1K dilution for 1 hour at room temperature.

SRPX2 regulated TGF-β/SMADs signaling pathways by AP1 and SMAD7. A: Results for Western blot analysis of p-SMAD2, SMAD2, p-SMAD3 and SMAD3 expression in HPFs following TGF-β1 stimulation. B-C : Western blot (B) and RT-PCR (C) analysis of SMAD7 expression in HPFs following TGF-β1 induction. D : Expression of AP1 in HPFs after TGF-β1 stimulation. E : Western blot results for analysis of the levels of P-SMAD2, P-SMAD3 and SMAD7 in HPFs pre-treated with T-5224 (an inhibitor for AP-1) treatment following TGF-β1 induction. The data are represented as the mean ± SEM of three independent experiments. *, p < 0.05; **, p < 0.01; ***, p < 0.001.
Index in PubMed under a CC BY license. PMID: 34093874

Srpx2 promoted FMT in BLM-induced pulmonary fibrosis. A : Western blot analysis of Fibronectin, Col1a1, α-SMA and Srpx2 expression in mice after BLM induction with Scrambled or Srpx2 siRNA-loaded liposomes. B : Representative images of immunostaining of Fibronectin, Col1a1 and α-SMA in the mice lung sections. The nuclei were stained blue by DAPI, and the images were taken under original magnification ×400. C : Western blot analysis of p-Smad2, p-Smad3 and Smad7 expression in mice after BLM induction. D : RT-PCR analysis of AP-1 expression in mice in each group. Six mice were included in each study group. The data are represented as the mean ± SEM. *, p < 0.05; **, p < 0.01; ***, p < 0.001.
Index in PubMed under a CC BY license. PMID: 34093874

TGF-β2 gene expression and TGF-Smad signaling are augmented in Sema7A-HUVECs. a The top 30 upregulated genes in Sema7A-HUVECs compared with Con335-HVUECs. b TGF-β2 mRNA level was analyzed by qPCR normalized to GAPDH. Fold changes are shown. Data are mean ± SEM, N = 3, *** p < 0.001. c The concentration of TGF-β2 in cell supernatant was detected by ELISA. N = 10. Unpaired two-tailed Student’s t tests was used to analysis the data. Data are mean ± SEM, ** p < 0.01. d GSEA based on gene ontology (GO) pathway database showed TGF-β signaling pathway was enrich in Sema7A-HUVECs. e , f Cells were treated with Oxymatrine (Oxy) (20 μmol/l) or T4442 (1 μg/ml) and the lysates were analyzed by western blotting for Smad3 phosphorylation, normalized to total Smad3. Data are mean ± SEM, N = 3, * p < 0.05; ** p < 0.01. g , h CD31 and α-SMA mRNA in cells treated with inhibitors were analyzed by qPCR normalized to GAPDH. Fold changes are shown. Data are mean ± SEM, N = 3, * p < 0.05; ** p < 0.01; *** p < 0.001. i – k CD31 and α-SMA proteins in cells treated with inhibitors were analysis by western blotting, normalized to tubulin. Data are mean ± SEM, N = 3, * p < 0.05; ** p < 0.01; *** p < 0.001. T4442: TGF-β2 blocking antibody; Oxymatrine (Oxy): TGF/Smad signaling pathway inhibitor.
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Inhibition of ATF3 reduced TGF-β2 expression and Sema7A-induced EndMT. a ATF3 mRNA level was analyzed by qPCR normalized to GAPDH. Fold changes are shown. Data are mean ± SEM, N = 3, *** p < 0.001 b ATF3 protein expression were analyzed by western blotting, normalized to tubulin. Data are mean ± SEM, N = 3, *** p < 0.001. c TGF-β2 mRNA level was analyzed by qPCR normalized to GAPDH. Fold changes are shown. Data are mean ± SEM, N = 3, * p < 0.05; ** p < 0.01. d The concentration of TGF-β2 in cell supernatant was detected by ELISA among Con335-HUVECs, Sema7A-HUVECs, and Sema7A-HUVECs + ATF3-siRNA. N = 10. Data are mean ± SEM, * p < 0.05; *** p < 0.001. e Chip-qPCR product in agarose gel electrophoresis. f Chip-qPCR TGF-β2 percentage of input in con335-HUVECs and Sema7A - HUVECs were analyzed by qPCR normalized to IgG. Data are mean ± SEM, N = 3, ** p < 0.01. g Schematic graph of the constructed reporter plasmid. TGF-β2 mut indicates the TGF-β2 mutation promoter region in ATF3 binding site. The mutated nucleotides in TGF-β2 fragments are in red letters. h Luciferase reporter assays were performed on HEK 293 T cells. Data are mean ± SEM, N = 3, ** p < 0.01 vs negative control. i , j ATF3-overexpression plasmid was transfected to HUVECs, and the mRNA levels of TGF - β2 and TGF-β1 were performed by qPCR normalized to GAPDH. Fold changes are shown. Data are mean ± SEM, N = 3, ** p < 0.01. k , l P-Smad3 protein in cells treated with siRNA was analyzed by Western blotting, normalized to total Smad3. Data are mean ± SEM, N = 3, * p < 0.05; ** p < 0.01. m – q CD31 and α-SMA RNA and proteins in cells treated with siRNA or control was analyzed by qPCR and western blotting. Data are mean ± SEM, N = 3, * p < 0.05; ** p < 0.01; *** p < 0.001.
Index in PubMed under a CC BY license. PMID: 32826874

Β1 integrin mediates Sema7A signal to TGF-β2 via ATF3. a Cells were treated with β1 integrin antibody (P5D2), and ATF3 mRNA level was analyzed by qPCR normalized to GAPDH. Fold changes are shown. Data are mean ± SEM, N = 3, * p < 0.05; ** p < 0.0.1. b ATF3 protein expression was analyzed by western blotting, normalized to tubulin. Data are mean ± SEM, N = 3, * p < 0.05; ** p < 0.01. c TGF-β2 mRNA level was analyzed by qPCR normalized to GAPDH. Fold changes are shown. Data are mean ± SEM, N = 3, * p < 0.05; ** p < 0.0.1. d The concentration of TGF-β2 in cell supernatant was detected by ELISA for Con335-HUVECs, Sema7A-HUVECs, and Sema7A-HUVECs + P5D2. Data are mean ± SEM, N = 10, * p < 0.05; *** p < 0.001. e , f Smad3 phosphorylation were analyzed by western blotting, normalized to total Smad3. Data are mean ± SEM, N = 3, * p < 0.05. g , h CD31 and α-SMA mRNA level were analyzed by qPCR normalized to GAPDH. Fold changes are shown. Data are mean ± SEM, N = 3, * p < 0.05; ** p < 0.01; *** p < 0.001. i – k CD31 and α-SMA proteins were analyzed by western blotting, normalized to tubulin. Data are mean ± SEM, N = 3, * p < 0.05; ** p < 0.01; *** p < 0.001. l ATF3 overexpression plasmid was transfected into P5D2 incubated Sema7A-HUVECs, and TGF-β2 mRNA expression in Sema7A-HUVECs + P5D2 and Sema7A-HUVECs + P5D2 + ATF3 were analyzed by qPCR normalized to GAPDH. Fold changes are shown. Data are mean ± SEM, N = 3, ** p < 0.01. m – o CD31 and α-SMA protein expressions were analyzed by western blotting, normalized to GAPDH. Data are mean ± SEM, N = 3, ** p < 0.01 (Sema7A-HUVECs + P5D2 vs Sema7A-HUVECs + P5D2 + ATF3). P5D2 β1 integrin antibody.
Index in PubMed under a CC BY license. PMID: 32826874

Western blot analysis of Phospho-Smad3 (S423/S425) expression in A549 cell lysate treated with TGF-?1.

All lanes use the Antibody at 1:1K dilution for 1 hour at room temperature.

All lanes use the Antibody at 1:1K dilution for 1 hour at room temperature.

Immunohistochemical analysis of paraffin-embedded human gastric adenocarcinoma, using Phospho-Smad3 (S423 + S425) Antibody.

Immunofluorescent analysis of A549 cells treated with TGFβ , using Phospho-Smad3 (S423 + S425) Antibody.