Journal: Journal of Cancer

Article Title: Ligand-independent EphB1 signaling mediates TGF-β-activated CDH2 and promotes lung cancer cell invasion and migration

doi: 10.7150/jca.44576

Figure Lengend Snippet: Primer sequence for real-time PCR

Article Snippet: Antibodies to proteins were obtained from the following sources: EphB1 (#ab129103) and phos-EphB1 (#ab61791) for Western blot were purchased from Abcam; EphB1 (#AF542) for immunohistochemistry was purchased from Novus Biologicals; N-cadherin (#13116) and E-cadherin (#3195) were from Cell Signaling Technology; β-actin (#60008-1-Ig), Smad2 (#12570-1-AP) and GAPDH (#60004-1-Ig) were from Proteintech.

Techniques: Sequencing

Journal: Journal of Cancer

Article Title: Ligand-independent EphB1 signaling mediates TGF-β-activated CDH2 and promotes lung cancer cell invasion and migration

doi: 10.7150/jca.44576

Figure Lengend Snippet: Ligand-independent EphB1 mediates TGF-β-activated N-cadherin. (A) Expression of mesenchymal molecules in lung cancer cells after transfection of EphB1 or knockdown of EphB1 quantified by real-time PCR. *p<0.05; **p<0.01; ***p<0.001. (B) Expression of CDH2 upon transfection of EphB1 measured by Western blot. (C) Expression of EphB1 upon treatment of TGF-β measured by Western blot. (D) Western blot was conducted to test the expression EphB1 after transfection of Smad2 or knockdown of Smad2. (E) Expression of EphB1 after transfection of Smad2 or knockdown of Smad2 quantified by real-time PCR. (F) The putative Smad2/3 binding sites in EphB1 promoter. (G) ChIP assay in 293 cells to detect the binding of Smad2 to the promoter of EphB1. Mean values± SD of three independent experiments are shown on the right side. IgG indicates nonspecific Ab.

Article Snippet: Antibodies to proteins were obtained from the following sources: EphB1 (#ab129103) and phos-EphB1 (#ab61791) for Western blot were purchased from Abcam; EphB1 (#AF542) for immunohistochemistry was purchased from Novus Biologicals; N-cadherin (#13116) and E-cadherin (#3195) were from Cell Signaling Technology; β-actin (#60008-1-Ig), Smad2 (#12570-1-AP) and GAPDH (#60004-1-Ig) were from Proteintech.

Techniques: Expressing, Transfection, Real-time Polymerase Chain Reaction, Western Blot, Binding Assay

Journal: Journal of Toxicologic Pathology

Article Title: Anti-Thy-1 Antibody-mediated Complement-dependent Cytotoxicity is Regulated by the Distribution of Antigen, Antibody and Membrane Complement Regulatory Proteins in Rats

doi: 10.1293/tox.26.41

Figure Lengend Snippet: Distribution of Thy-1.1 antigen in normal rats. A) The distribution of the Thy-1.1 antigen is shown by the intensity and frequency of positive staining. Red, orange, yellow and white indicate strong, moderate, weak and negative staining intensity, respectively. The frequency of staining is shown as the number of colored columns. Each column stands for 25%. *Other organs and tissue elements. B) Immunohistochemistry of Thy-1.1 in the kidney, adrenal gland and thymus. Positive reactions can be seen in mesangial cells of the kidney, medullary cells of the adrenal gland and lymphocytes of the cortex in the thymus. Bar = 50 µm.

Article Snippet: Antibodies against Thy-1.1 (CD90, OX-7, Cedarlane Laboratories Ltd., Burlington, ON, Canada, 10 μg/mL), Crry (512, BD PharMingen, San Jose, CA, USA, 0.7 μg/mL), CD55 (I-19, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA, 8 μg/mL) and C3 (Cappel, Aurora, OH, USA, 10 μg/mL) were used as the primary antibodies and applied to tissues processed by the PLP-AMeX method.

Techniques: Staining, Negative Staining, Immunohistochemistry

Journal: Journal of Toxicologic Pathology

Article Title: Anti-Thy-1 Antibody-mediated Complement-dependent Cytotoxicity is Regulated by the Distribution of Antigen, Antibody and Membrane Complement Regulatory Proteins in Rats

doi: 10.1293/tox.26.41

Figure Lengend Snippet: Immunohistochemical and histopathological findings in the PBS- or anti-Thy-1.1 antibody-injected rats. Immunohistochemistry for the anti-Thy-1.1 antibody and C3 and HE staining are shown. Distribution of the injected anti-Thy-1.1 antibody can be seen in the kidney, adrenal gland and thymus of the antibody-treated rat but not in the PBS-treated animal. C3 deposition in the mesangial cells of the kidney can be seen in the antibody-treated animal but cannot be seen in the PBS-treated animal. There is no positive staining in the adrenal gland or thymus of the rat injected with anti-Thy-1.1 antibody. Mesangial cell death can be seen even though cell death cannot be seen in the adrenal gland and thymus of the rat injected with anti-Thy-1.1 antibody. Karyolysis in the mesangial cell (arrowheads) and infiltrations of a small number of neutrophils (arrows) in anti-Thy-1.1 antibody-injected rat are also shown. The asterisks indicate the blood vessel in the thymus. All images are from animals at 0.5 hours after injection of PBS or the antibody. Bar = 50 µm.

Article Snippet: Antibodies against Thy-1.1 (CD90, OX-7, Cedarlane Laboratories Ltd., Burlington, ON, Canada, 10 μg/mL), Crry (512, BD PharMingen, San Jose, CA, USA, 0.7 μg/mL), CD55 (I-19, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA, 8 μg/mL) and C3 (Cappel, Aurora, OH, USA, 10 μg/mL) were used as the primary antibodies and applied to tissues processed by the PLP-AMeX method.

Techniques: Immunohistochemical staining, Injection, Immunohistochemistry, Staining

Journal: Journal of Toxicologic Pathology

Article Title: Anti-Thy-1 Antibody-mediated Complement-dependent Cytotoxicity is Regulated by the Distribution of Antigen, Antibody and Membrane Complement Regulatory Proteins in Rats

doi: 10.1293/tox.26.41

Figure Lengend Snippet: Changes in the mesangial cells of the kidney at 8 and 24 hours after injection of the anti-Thy-1.1 antibody in HE staining. In the animal sacrificed at 8 hours after injection, the number of mesangial cells is decreased with reduced karyolysis and increased neutrophil infiltration (arrows). In the animal sacrificed at 24 hours after injection, the mesangial area is decreased and is accompanied by capillary dilatation of the glomerulus (arrowheads). Bar = 50 µm.

Article Snippet: Antibodies against Thy-1.1 (CD90, OX-7, Cedarlane Laboratories Ltd., Burlington, ON, Canada, 10 μg/mL), Crry (512, BD PharMingen, San Jose, CA, USA, 0.7 μg/mL), CD55 (I-19, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA, 8 μg/mL) and C3 (Cappel, Aurora, OH, USA, 10 μg/mL) were used as the primary antibodies and applied to tissues processed by the PLP-AMeX method.

Techniques: Injection, Staining

Journal: Journal of Toxicologic Pathology

Article Title: Anti-Thy-1 Antibody-mediated Complement-dependent Cytotoxicity is Regulated by the Distribution of Antigen, Antibody and Membrane Complement Regulatory Proteins in Rats

doi: 10.1293/tox.26.41

Figure Lengend Snippet: Categories of CDC activation in Thy-1.1 antigen-expressing cells. A) The antibody binds to the antigen, which leads to activation of the CDC pathway and then to cell death. B) The antibody binds to the antigen, but the CDC pathway is not activated because of mCRP expression. Cell death is not induced. C) The antibody does not bind to antigen, and the CDC pathway is not activated; thus, and there is no cell death.

Article Snippet: Antibodies against Thy-1.1 (CD90, OX-7, Cedarlane Laboratories Ltd., Burlington, ON, Canada, 10 μg/mL), Crry (512, BD PharMingen, San Jose, CA, USA, 0.7 μg/mL), CD55 (I-19, Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA, 8 μg/mL) and C3 (Cappel, Aurora, OH, USA, 10 μg/mL) were used as the primary antibodies and applied to tissues processed by the PLP-AMeX method.

Techniques: Activation Assay, Expressing

Journal: Cell Death Discovery

Article Title: SMAD3 and FTO are involved in miR-5581-3p-mediated inhibition of cell migration and proliferation in bladder cancer

doi: 10.1038/s41420-022-01010-8

Figure Lengend Snippet: a Heatmap assessed from mRNA-seq data listed a part of potential downstream genes of miR-5581-3p. b Volcano plot illustrated enrichment of remarkably differentially expressed genes in miR-5581-3p overexpressed vs. control UM-UC3 cells. c KEGG pathways enrichment from mRNA-seq data. d Venn diagram illustrated the intersected genes from mRNA-seq and online databases including miRWalk, TargetScan, and ENCORI. e The relative content of SMAD3 and FTO in BCa were associated with miR-5581-3p in ENCORI database. f A significant downregulation of SMAD3 and FTO were detected via qRT-PCR assay in UM-UC3 and T24 cell lines post over-expression of miR-5581-3p. g Dual-luciferase enzyme reporter assay illustrated that miR-5581-3p remarkably reduced the luciferase enzyme activity of vectors harboring the 3′-UTRs of SMAD3 and FTO. h Schematic diagram illustrating the miR-5581-3p-targeting sites of SMAD3 and FTO with seed matching. i Western blot data demonstrating decreased protein concentration of SMAD3, FTO, and other proteins in cells transfected with miR-5581-3p mimics. j Western blot data demonstrated the protein levels of SMAD3 and FTO changed in BCa cells transfected of miR-5581-3p mimics at the concentration of 0, 25, 50, and 75 nM. Error bars designate SD acquired from three independent experiments; *P < 0.05 , **P < 0.01 , ***P < 0.001 .

Article Snippet: The primary antibodies utilized consisted of: anti-N-cadherin, anti-MMP9 (Proteintech Group); anti-β-actin, anti-E-cadherin, anti-VIMENTIN, anti-CCND1, anti-CDK4, anti-SMAD3, anti-p-SMAD3, anti-FTO, and anti-SNAIL (Cell Signaling Technology).

Techniques: Quantitative RT-PCR, Over Expression, Luciferase, Reporter Assay, Activity Assay, Western Blot, Protein Concentration, Transfection, Concentration Assay

Journal: Cell Death Discovery

Article Title: SMAD3 and FTO are involved in miR-5581-3p-mediated inhibition of cell migration and proliferation in bladder cancer

doi: 10.1038/s41420-022-01010-8

Figure Lengend Snippet: a Western blot assays showed co-transfection of miR-5581-3p and SMAD3 rescued the protein levels of SMAD3 in UM-UC3 and T24 cell lines. b Western blot data showed co- transfection of miR-5581-3p and FTO rescued the protein levels of FTO in UM-UC3 and T24 cell lines. c Transwell assays consistently showed miR-5581-3p-induced cell migration inhibition was partly rescued by SMAD3 over-expression. d Transwell assays consistently showed miR-5581-3p-induced cell migration inhibition was partially rescued by FTO over-expression. e The wound-healing assays exhibited that the miR-5581-3p-triggered cell mobility dampening was partly rescued via SMAD3 over-expression. f The wound-healing assays illustrated that the miR-5581-3p-triggered cell mobility dampening was partly rescued via FTO over-expression. g Colony formation assays illustrated that the co-transfection of miR-5581-3p and FTO rescued the growth dampening triggered by miR-5581-3p over-expression. Error bars designate SD acquired from three independent experiments; *P < 0.05, **P < 0.01, *** P < 0.001.

Article Snippet: The primary antibodies utilized consisted of: anti-N-cadherin, anti-MMP9 (Proteintech Group); anti-β-actin, anti-E-cadherin, anti-VIMENTIN, anti-CCND1, anti-CDK4, anti-SMAD3, anti-p-SMAD3, anti-FTO, and anti-SNAIL (Cell Signaling Technology).

Techniques: Western Blot, Cotransfection, Migration, Inhibition, Over Expression

Journal: Cell Death Discovery

Article Title: SMAD3 and FTO are involved in miR-5581-3p-mediated inhibition of cell migration and proliferation in bladder cancer

doi: 10.1038/s41420-022-01010-8

Figure Lengend Snippet: miR-5581-3p inhibits the expression of SMAD3 and FTO via binding to the 3′-UTR of their mRNAs. Inhibition of SMAD3 suppresses the migration of BCa cells, and inhibition of FTO suppresses the migration and proliferation of BCa cells via m 6 A modification.

Article Snippet: The primary antibodies utilized consisted of: anti-N-cadherin, anti-MMP9 (Proteintech Group); anti-β-actin, anti-E-cadherin, anti-VIMENTIN, anti-CCND1, anti-CDK4, anti-SMAD3, anti-p-SMAD3, anti-FTO, and anti-SNAIL (Cell Signaling Technology).

Techniques: Expressing, Binding Assay, Inhibition, Migration, Modification