anti total smad 2 3  (Cell Signaling Technology Inc)

Journal: Journal of Biochemical and Molecular Toxicology

Article Title: Molecular Mechanisms of Neutrophil Extracellular Traps in Promoting Gastric Cancer Epithelial–Mesenchymal Transition Through SERPINE‐1 Expression

doi: 10.1002/jbt.70157

Figure Lengend Snippet: NETs promote EMT in gastric cancer cells. (A) Transwell invasion assay and wound healing assay of HGC‐27 and MKN‐45 cells treated with NETs or DNase‐1‐treated NETs. Representative images at 48 h were shown. (B) Western blot analysis of EMT markers (PAI‐1, Vimentin, N‐cadherin, E‐cadherin) in HGC‐27 and MKN‐45 cells treated with NETs or DNase‐1‐treated NETs. Additionally, immunofluorescence co‐staining of PAI‐1 and EMT typical markers (Vimentin, N‐cadherin, and E‐cadherin) in HGC‐27 and MKN‐45 cells treated with NETs. Scale bar: 20 μm. (C) Sphere formation assay and CCK‐8 proliferation assay of HGC‐27 and MKN‐45 cells treated with NETs or DNase‐1‐treated NETs. Representative images of sphere formation assay at 48 h were shown.

Article Snippet: The membranes were blocked with 5% non‐fat milk and then incubated with primary antibodies against PAI‐1, Vimentin, N‐cadherin, E‐cadherin, TGF‐β1, TGF‐βR1, TGF‐βR2, phospho‐Smad2/3, Smad4 and total Smad2/3 (all 1:1,000, Cell Signaling Technology) overnight at 4°C.

Techniques: Transwell Invasion Assay, Wound Healing Assay, Western Blot, Immunofluorescence, Staining, Tube Formation Assay, CCK-8 Assay, Proliferation Assay

Journal: Journal of Biochemical and Molecular Toxicology

Article Title: Molecular Mechanisms of Neutrophil Extracellular Traps in Promoting Gastric Cancer Epithelial–Mesenchymal Transition Through SERPINE‐1 Expression

doi: 10.1002/jbt.70157

Figure Lengend Snippet: SERPINE‐1 knockdown reverses NET‐induced EMT in gastric cancer cells. (A) Western blot analysis confirming SERPINE‐1 knockdown efficiency in HGC‐27 and MKN‐45 cells. (B) Western blot analysis of EMT markers in control and SERPINE‐1 knockdown cells treated with NETs. Additionally, immunofluorescence co‐staining of PAI‐1 and EMT typical markers (Vimentin, N‐cadherin, and E‐cadherin) in control and SERPINE‐1 knockdown cells treated with NETs. Scale bar: 20 μm. (C) Transwell invasion assay and wound healing assay of control and SERPINE‐1 knockdown cells treated with NETs. Representative images at 48 h were shown. (D) CCK‐8 proliferation assay of control and SERPINE‐1 knockdown cells treated with NETs.

Article Snippet: The membranes were blocked with 5% non‐fat milk and then incubated with primary antibodies against PAI‐1, Vimentin, N‐cadherin, E‐cadherin, TGF‐β1, TGF‐βR1, TGF‐βR2, phospho‐Smad2/3, Smad4 and total Smad2/3 (all 1:1,000, Cell Signaling Technology) overnight at 4°C.

Techniques: Knockdown, Western Blot, Control, Immunofluorescence, Staining, Transwell Invasion Assay, Wound Healing Assay, CCK-8 Assay, Proliferation Assay

Journal: Frontiers in Immunology

Article Title: Potential implications of granzyme B in keloids and hypertrophic scars through extracellular matrix remodeling and latent TGF-β activation

doi: 10.3389/fimmu.2024.1484462

Figure Lengend Snippet: List of primary and secondary antibodies used for immunohistochemistry (IHC), tyramide signal amplification (TSA), co-immunofluorescence (Co-IF) and Western blotting (WB).

Article Snippet: WB , Rabbit anti-human total smad2/3 , , Cell signaling (mAb#3102) , 1/500 in TBS-0.1% Tween + 5% milk , HRP-conjugated goat anti-rabbit , 1/2,500 in TBS-0.1% Tween, 10% milk.

Techniques: Immunohistochemistry, Amplification, Western Blot, Blocking Assay

Journal: Scientific Reports

Article Title: Knockout or inhibition of DHPS suppresses ovarian tumor growth and metastasis by attenuating the TGFβ pathway

doi: 10.1038/s41598-025-85466-5

Figure Lengend Snippet: KO or inhibition of DHPS attenuated the TGFβ signaling pathway in OC cells. a , b . Western blot analysis showing the expression of phospho- and total SMAD2 in DHPS knockout (KO) and control SKOV3 and OVCAR8 cells following treatment with 6 ng/ml TGFβ at the indicated time points. c , d . Western blot analysis showing the expression of phospho- and total SMAD2 in SKOV3 and OVCAR8 cells following treatment with 20 µM GC7 for 12 h and subsequent treatment with 6 ng/ml TGFβ at the indicated time points.

Article Snippet: The membranes were then blocked with 5% nonfat milk for 1 h and incubated with primary antibodies against the following proteins: DHPS (Novus), Hypusine (Millipore Sigma), EIF5A2, Cytokeratin-7 (Abcam), GAPDH (Santa Cruz; St. Louis, MO), Vimentin, E-cadherin, β-catenin, total-Smad2, or p-Smad2 (Cell Signaling).

Techniques: Inhibition, Western Blot, Expressing, Knock-Out, Control

Journal: Scientific Reports

Article Title: Knockout or inhibition of DHPS suppresses ovarian tumor growth and metastasis by attenuating the TGFβ pathway

doi: 10.1038/s41598-025-85466-5

Figure Lengend Snippet: KO or inhibition of DHPS using lentiviral CRISPR/Cas9 nickase vector suppressed primary ovarian tumor growth and metastasis in orthotopic OC mouse model. ( a ) Primary ovarian tumors dissected one month after intrabursal injection of DHPS KO and control OVCAR8 cells. Tumor weight in DHPS KO is significantly lower than in the control group ( n = 5, ** P < 0.01). ( b ) Bioluminescence imaging of metastatic tumors in the liver of mice xenografted with DHPS KO and control cells ( n = 3, ** P < 0.01). ( c ) Histopathological characterization of metastatic tumors from the liver of mice xenografted with DHPS KO and control cells using H&E staining. ( d ) Western blot analysis of DHPS, Hypusine, EIF5A2, p-SMAD2, and EMT markers in primary tumors of mice xenografted with DHPS KO and control cells. ( e ) Significant reduction in primary ovarian tumor weight in mice treated with GC7 compared to control (*** P < 0.001). ( f ) Bioluminescence imaging of metastatic tumors in the liver of mice xenografted with GC7 treatment and the control group (*** P < 0.001). ( g ) Histopathological characterization of metastatic tumors from the liver of mice xenografted with GC7 treatment and the control group using H&E staining. ( h ) Western blot analysis of DHPS, EIF5A2, p-SMAD2, and EMT markers in primary tumors of mice treated with GC7 and the control group (** P < 0.01; *** P < 0.001).

Article Snippet: The membranes were then blocked with 5% nonfat milk for 1 h and incubated with primary antibodies against the following proteins: DHPS (Novus), Hypusine (Millipore Sigma), EIF5A2, Cytokeratin-7 (Abcam), GAPDH (Santa Cruz; St. Louis, MO), Vimentin, E-cadherin, β-catenin, total-Smad2, or p-Smad2 (Cell Signaling).

Techniques: Inhibition, CRISPR, Plasmid Preparation, Injection, Control, Imaging, Staining, Western Blot