anti bmpr ii antibody  (R&D Systems)

Journal: Regenerative Therapy

Article Title: Bone morphogenetic protein-2-derived osteogenic peptide promotes bone regeneration via osteoblastogenesis

doi: 10.1016/j.reth.2025.09.006

Figure Lengend Snippet: The osteogenic signal transduction of OP5. (A) The binding affinity of the peptide to BMP receptor types IA and II (BMPR-IA and II). BMPR-IA or BMPR-II in lysate of human mesenchymal stromal cells (hMSCs) was immobilized on a microplate using a specific BMP receptor antibody in an ELISA. Data represents mean ± S.D. Binding activity of BMP-2 (2 μg/ml) and OP5 (20 μg/ml) are detected using anti-BMP-2 monoclonal antibody and 1:2000 horseradish peroxidase (HRP)-conjugated rabbit anti-mouse IgG as a secondary antibody binding to primary antibody. Each bar shows the mean ± S.D. ∗ P < 0.05, ∗∗ P < 0.01. (B) Western blot analysis of β-catenin, phosphorylated cAMP response element-binding protein (P-CREB), and phosphorylated SMAD (P-SMAD) expression in hMSCs treated with BMP-2 and OP5 for 3 and 6 h. Cell lysates were prepared and analyzed by western blot using antibodies to β-catenin, P-CREB, and P-SMAD (all of 1:1000). The expression level of each group was normalized with the density % of β-actin in the same group. O.D., optical density; None, without peptide or protein group; BMP, bone morphogenetic protein; OP, osteogenic peptide; DWIVA, amino acid sequence from osteogenic peptide; 10F, 10F medium [Low-glucose Dulbecco's modified Eagle's medium (DMEM) containing 10 % fetal bovine serum (FBS) and 1 % antibiotic/antimycotic] as the control group.

Article Snippet: The plates were then incubated overnight at 4 °C with 1:1,000 mouse anti-BMPR-IA and anti-BMPR-II antibody (R&D systems), followed by reaction with 1:2000 horseradish peroxidase (HRP)-conjugate rabbit anti-mouse IgG (catalog no. 7076; Cell Signaling Technology Inc., Danvers, MA, USA) for 30 min at room temperature.

Techniques: Transduction, Binding Assay, Enzyme-linked Immunosorbent Assay, Activity Assay, Western Blot, Expressing, Sequencing, Modification, Control

Journal: Nature Communications

Article Title: TNFα drives pulmonary arterial hypertension by suppressing the BMP type-II receptor and altering NOTCH signalling

doi: 10.1038/ncomms14079

Figure Lengend Snippet: ( a , b ) BMPR2 mRNA expression, normalized to ACTB , in human dPASMCs ( a ) and PAECs ( b ) treated with IL-1β (1 ng ml −1 ), IL-6 (25 ng ml −1 ), IL-8 (25 ng ml −1 ) or TNFα (1 ng ml −1 ) for 24 h ( n= 3; Student's t -test). ( c , d ) Representative immunoblots of BMPR-II expression in human dPASMCs ( c ) and PAECs ( d ) treated with TNFα (1 ng ml −1 ) for 1, 4, 8 or 24 h. Reprobed for α-tubulin to ensure equal loading. The data shown are representative of three experiments. ( e ) Representative confocal images of immunohistochemical staining for TNFα (turquoise) and αSMA (magenta) in lung sections from control, idiopathic and heritable PAH subjects. Nuclei were counterstained with DAPI (blue). Scale bars, 100 μm. ( f ) Assessment of right ventricular systolic pressure (RVSP) from Bmpr2 +/+ , SP-C/Tnf/Bmpr2 +/+ , Bmpr2 +/− and SP-C/Tnf/Bmpr2 +/− ( n= 4 per group) mice. ( g ) Representative immunoblots of BMPR-II expression in lungs isolated from 8 week old Bmpr2 +/+ and SP-C/Tnf/Bmpr2 +/+ transgenic mice. Reprobed for β-actin to ensure equal loading ( n= 4). ( h ) Representative immunoblots of BMPR-II, ADAM10 and ADAM17 in human dPASMCs transfected with DharmaFECT1 alone (DH1), si ADAM10 , si ADAM17 , combined si ADAM10 +si ADAM17 (siADAM10/17) or non-targeting siRNA control (siCP) with or without TNFα (1 ng ml −1 ) treatment for 24 h. Reprobed for α-tubulin to ensure equal loading. The data shown are representative of three experiments. ( i ) ELISA measurement of soluble BMPR-II in conditioned media from human dPASMCs transfected with wild-type and mutant 5′-myc-tagged BMPR-II constructs and treated with TNFα (1 ng ml −1 ) for 24 h ( n= 3). One-way analysis of variance with post hoc Tukey's for multiple comparisons used in f and i . * P ≤0.05, *** P ≤0.001. Error bars represent mean±s.e.m. Lower molecular mass BMPR-II is indicated by an arrow.

Article Snippet: Briefly, flat-bottom high binding 96-well ELISA plates (Greiner, South Lanarkshire, UK) were coated with 1 μg ml −1 mouse monoclonal anti-human BMPR-II antibody (R&D Systems) for 2 h at room temperature.

Techniques: Expressing, Western Blot, Immunohistochemical staining, Staining, Control, Isolation, Transgenic Assay, Transfection, Enzyme-linked Immunosorbent Assay, Mutagenesis, Construct

Journal: Nature Communications

Article Title: TNFα drives pulmonary arterial hypertension by suppressing the BMP type-II receptor and altering NOTCH signalling

doi: 10.1038/ncomms14079

Figure Lengend Snippet: ( a ) BMPR2 mRNA expression, normalized to ACTB , in human dPASMCs from disease-free controls and HPAH patients stimulated with TNFα (1 ng ml −1 ) for 24 h ( n= 3; Student's t -test). ( b , c ) Pre-treatment of human dPASMCs from disease-free controls and HPAH patients with or without TNFα (1 ng ml −1 ) for 23 h before 1 h stimulation with BMP2 (10 ng ml −1 ) or BMP6 (10 ng ml −1 ). ( b ) Representative immunoblots of phospho-Smad1/5, total Smad1 and ID1 expression. Reprobed for α-tubulin to ensure equal loading. The data shown are representative of three control and HPAH cell lines. ( c ) ID1 mRNA expression, normalized to ACTB ( n= 3). ( d ) Proliferation of human dPASMCs from disease-free controls and HPAH patients after 6 days treatment. Cells treated every 48 h as indicated with BMP2 (10 ng ml −1 ), BMP4 (10 ng ml −1 ) or BMP6 (10, 25 or 50 ng ml −1 ; n= 3). ( e ) Proliferation of human dPASMCs from disease-free controls and HPAH patients after 6 days treatment. Cells treated every 48 h as indicated with TNFα (1 ng ml −1 ) and/or BMP6 (10 or 50 ng ml −1 ) ( n= 3 control and HPAH cell lines). ( f ) Proliferation of human control dPASMCs on day 6 after transfection with DharmaFECT1 alone (DH1), si BMPR2 or non-targeting siRNA control (siCP) and treatment every 48 h with TNFα (1 ng ml −1 ) and/or BMP6 (50 ng ml −1 ) as indicated ( n= 3). ( g ) Proliferation of human HPAH dPASMCs on day 6 following transduction with adenovirus expressing full-length wild-type and kinase-dead D485G mutant BMPR-II cDNA and treatment every 48 h with TNFα (1 ng ml −1 ) and/or BMP6 (50 ng ml −1 ) as indicated ( n= 3). One-way analysis of variance with post hoc Tukey's for multiple comparisons used in ( c – g ). * P ≤0.05, ** P ≤0.01, *** P ≤0.001. Error bars represent mean±s.e.m.

Article Snippet: Briefly, flat-bottom high binding 96-well ELISA plates (Greiner, South Lanarkshire, UK) were coated with 1 μg ml −1 mouse monoclonal anti-human BMPR-II antibody (R&D Systems) for 2 h at room temperature.

Techniques: Expressing, Western Blot, Control, Transfection, Transduction, Mutagenesis

Journal: Nature Communications

Article Title: TNFα drives pulmonary arterial hypertension by suppressing the BMP type-II receptor and altering NOTCH signalling

doi: 10.1038/ncomms14079

Figure Lengend Snippet: Rats were given vehicle injections and maintained in normoxia (Control, n =6) or challenged with SU-5416 (20 mg kg −1 , s.c.) and 3 weeks of hypoxia (10% O2) before 5 weeks of normoxia and 3 weeks of biweekly treatment with saline vehicle (S/H, n =9) or etanercept (S/H+Etan, n =9; 2.5 mg kg −1 , i.p.). ( a , b ) Assessment of RVSP ( a ) and right ventricular hypertrophy (Fulton index (RV/LV+S)) ( b ). ( c ) Quantification of non-, partially and fully muscularized arteries as a percentage of total alveolar wall and duct arteries ( n =6 for control, n =9 for all other groups; Student's t -test for non-muscularized vessels). ( d ) BMPR2 , ACVR2A and ALK2 mRNA expression, normalized to Actb , in lungs isolated from control, S/H and S/H+Etan rats ( n= 6). ( e ) Representative immunoblots of BMPR-II, phospho-Smad1/5, total Smad1, Notch2, Notch3, cleaved Caspase3, total Caspase3 and αSMA expression in lungs isolated from of control, S/H and S/H+Etan rats. Reprobed for β-actin to ensure equal loading ( n= 3). ( f , g ) Notch2 and Notch3 mRNA expression in lungs isolated from lungs of control, S/H and S/H+Etan rats. Expression was normalized to Actb ( n= 6). ( h ) Representative images of immunohistochemical staining for Notch2, Notch3 and αSMA in lung sections from control and S/H rats. Scale bars, 100 μm. One-way analysis of variance with post hoc Tukey's for multiple comparisons used in a , b , d , f and g . */ # P ≤0.05, **/ ## P ≤0.01, *** P ≤0.001. Error bars represent mean±s.e.m.

Article Snippet: Briefly, flat-bottom high binding 96-well ELISA plates (Greiner, South Lanarkshire, UK) were coated with 1 μg ml −1 mouse monoclonal anti-human BMPR-II antibody (R&D Systems) for 2 h at room temperature.

Techniques: Control, Saline, Expressing, Isolation, Western Blot, Immunohistochemical staining, Staining