Journal: BMC Research Notes

Article Title: Identification of a bone morphogenetic protein type 2 receptor neutralizing antibody

doi: 10.1186/s13104-019-4367-0

Figure Lengend Snippet: Antibody 3F6 reduces BMPR2-ECD ligand-binding activity in a modified immunoprecipitation assay. Neutralizing the ligand-binding activity of BMPR2-ECD using various amounts of 3F6. Results are quantified by ELISA and expressed as mean ± SEM relative to the ligand-binding activity of BMPR2-ECD in the absence of 3F6. n ≥ 3 per condition. Asterisk indicates p < 0.05 by paired t test

Article Snippet: BMPR2-ECD/Fc fusion (Sino Biologicals 10551-H03H) was mixed with 5 µL Protein G-coupled Dynabeads (Invitrogen 1003D) at room temperature for 30 min in 200 µL total volume with gentle rocking.

Techniques: Ligand Binding Assay, Activity Assay, Modification, Immunoprecipitation, Enzyme-linked Immunosorbent Assay

Journal: BMC Research Notes

Article Title: Identification of a bone morphogenetic protein type 2 receptor neutralizing antibody

doi: 10.1186/s13104-019-4367-0

Figure Lengend Snippet: Antibody 3F6 reduces activation of BMP pathway in HEK293T cells. a Expression of endogenous BMPR2 by HEK293T cells compared to β-actin loading control. Approximate molecular weights are indicated. b , c BMP2 induces phosphorylation of SMAD1, 5, and 8 (pSMAD1,5,8) in HEK293T cells and this response is blunted by pre-treatment with 3F6. Approximate molecular weights are indicated in b . Results are quantified in c and expressed as mean ± SEM ratio of phosphorylated SMAD1,5,8: total SMAD1 relative to BMP2 treatment alone. No effect on BMP2-responsiveness was observed with pre-treatment using control ascites (Ctrl Asc.). n = 3 per condition. Asterisk indicates p < 0.05 by paired t test

Article Snippet: BMPR2-ECD/Fc fusion (Sino Biologicals 10551-H03H) was mixed with 5 µL Protein G-coupled Dynabeads (Invitrogen 1003D) at room temperature for 30 min in 200 µL total volume with gentle rocking.

Techniques: Activation Assay, Expressing

Journal: BMC Research Notes

Article Title: Identification of a bone morphogenetic protein type 2 receptor neutralizing antibody

doi: 10.1186/s13104-019-4367-0

Figure Lengend Snippet: Antibody 3F6 has no effect on BMP-responsiveness in BMPR2 knock-down HEK293T cells. a Expression levels of endogenous BMPR2 in scramble control HEK293T cells (Control) and HEK293T cells carrying anti- BMPR2 shRNA ( BMPR2 -KD) compared to HPRT1 loading control. Data are expressed as normalized to scramble control (Control) using the 2 −∆∆Ct method. n = 6 per condition. Asterisk indicates p < 0.05 by unpaired t test. b , c Expression of endogenous BMPR2 in scramble control HEK293T cells (Control) and BMPR2 -KD HEK293T cells compared to β-actin loading control. Approximate molecular weights are indicated in b . Representative immunoblot is shown in b and results from three independent runs are quantified in c (data are expressed as mean ± SEM ratio of BMPR2: β-actin normalized to scramble control (Relative Expression)). Asterisk indicates p < 0.05 by paired t test. d , e BMP2 induces phosphorylation of SMAD1, 5, and 8 (pSMAD1,5,8) in BMPR2 - KD HEK293T cells. No effect on BMP2-responsiveness of BMPR2 - KD HEK293T cells was observed with pre-treatment using control ascites (Ctrl Asc.) or 3F6. Approximate molecular weights are indicated in d . Results are quantified in E and expressed as mean ± SEM ratio of phosphorylated SMAD1,5,8: total SMAD1 relative to BMP2 treatment alone (Relative pS1:S1). n = 4 per condition

Article Snippet: BMPR2-ECD/Fc fusion (Sino Biologicals 10551-H03H) was mixed with 5 µL Protein G-coupled Dynabeads (Invitrogen 1003D) at room temperature for 30 min in 200 µL total volume with gentle rocking.

Techniques: Expressing, shRNA, Western Blot

Journal: Scientific Reports

Article Title: miR-889-3p targeting BMPR2 promotes the development of retinoblastoma via JNK/MAPK/ERK signaling

doi: 10.1038/s41598-023-49994-2

Figure Lengend Snippet: Real-time PCR primer sequences.

Article Snippet: The miR-889-3p mimics/inhibitor, negative control (NC) mimics/inhibitors, si-BMPR2, and si-NC were developed and acquired from GenePharma Co., Ltd. (Shanghai, China).

Techniques: Real-time Polymerase Chain Reaction, Sequencing

Journal: Scientific Reports

Article Title: miR-889-3p targeting BMPR2 promotes the development of retinoblastoma via JNK/MAPK/ERK signaling

doi: 10.1038/s41598-023-49994-2

Figure Lengend Snippet: miR‐889-3p directly targets BMPR2. ( A ) Ninety-three genes overlapped with miRDB and GSE97508 according to the Venny 2.1. ( B ) Gene Ontology (GO) enrichment of 93 genes using STRING software. ( C ) The relative enrichment of PLCB1, MEF2C, and BMPR2 in Rb cells transfected with Bio-NC or Bio-miR-889-3p was assessed by an RNA pull-down assay. ** P < 0.001 versus Bio-NC. ( D ) The binding site in the 3′UTR of BMPR2 for miR‐889-3p was predicted by TargetScan. ( E ) Luciferase activity was evaluated in HXO-RB44 and SO-RB50 cells co-transfected with miR‐889-3p mimics or miRNA negative control and wild type (WT) or mutant (MUT) 3′UTR of BMPR2, ** P < 0.001 versus miR-NC. ( F ) BMPR2 expression was determined by qRT-PCR in Rb tissues and matched with nearby normal tissues. ( G ) In two Rb cell lines and a normal retinal epithelial cell line (ARPE-19), BMPR2 expression was determined by qRT-PCR (HXO-RB44 and SO-RB50); ** P < 0.001 versus ARPE-19 cells. ( H ) MiR-889-3p and BMPR2 were used in Pearson’s association analysis.

Article Snippet: The miR-889-3p mimics/inhibitor, negative control (NC) mimics/inhibitors, si-BMPR2, and si-NC were developed and acquired from GenePharma Co., Ltd. (Shanghai, China).

Techniques: Software, Transfection, Pull Down Assay, Binding Assay, Luciferase, Activity Assay, Negative Control, Mutagenesis, Expressing, Quantitative RT-PCR

Journal: Scientific Reports

Article Title: miR-889-3p targeting BMPR2 promotes the development of retinoblastoma via JNK/MAPK/ERK signaling

doi: 10.1038/s41598-023-49994-2

Figure Lengend Snippet: BMPR2 inhibited the biological effects of miR-889-3p in Rb cells. ( A ) BMPR2 expression level was determined using western blotting in HXO-RB44 and SO-RB50 cells transfected with si-NC, inhibitor-NC, si-BMPR2, miR-889-3p inhibitor or co-transfection of si-BMPR2 and miR-889-3p inhibitors. ( B – D ) A CCK-8 assay ( B ), scratch test ( C ), as well as western blotting ( D ) were utilized to compare the effect of co-transfection of miR-889-3p inhibitor and si-BMPR2 on cell proliferation, migration, and expression of key molecules of the JNK/MAPK/ERK signaling pathway in HXO-RB44, as well as in SO-RB50 cells transfected with the miR-889-3p inhibitor or si-BMPR2. ## P < 0.001 versus inhibitor-NC; * P < 0.05, ** P < 0.001 versus si-NC; & P < 0.05, && P < 0.001 versus si-BMPR2 + miR-889-3p inhibitor.

Article Snippet: The miR-889-3p mimics/inhibitor, negative control (NC) mimics/inhibitors, si-BMPR2, and si-NC were developed and acquired from GenePharma Co., Ltd. (Shanghai, China).

Techniques: Expressing, Western Blot, Transfection, Cotransfection, CCK-8 Assay, Migration

Journal: Frontiers in immunology

Article Title: Comprehensive Urinomic Identification of Protein Alternatives to Creatinine Normalization for Diagnostic Assessment of Lupus Nephritis.

doi: 10.3389/fimmu.2022.853778

Figure Lengend Snippet: FIGURE 3 | Urine proteins that correlate best with urine creatinine. The aptamer-based screen identified five urine proteins that correlated best with urine creatinine but not differ between subject groups. As described in Figure 1, the levels of 1129 urine proteins were assayed in 7 active LN, 8 inactive LN, 8 healthy controls using an aptamer-based proteomic screen. Of these, urine BMPRII, Dectin-1, HVEM, RELT, and SPINT2 (A–E) exhibited the highest Pearson R correlation with creatinine but did not differ significantly between the subject groups.

Article Snippet: Kits were purchased for HVEM (Cat. No. EK1226, Boster Biological Technology, Pleasanton, CA, USA), BMPRII (Cat. No. ELH-BMPR2-1, RayBiotech, Inc., Norcross, GA, USA), Dectin-1 (Cat. No. ELH-DECTIN1-1, RayBiotech, Inc., Norcross, GA, USA), SPINT2 (Cat. No. DY1106, R&D Systems, Inc., Minneapolis, MN, USA), and RELT (Cat. No. SEK10530, Sino Biological Inc., Beijing, China).

Techniques:

Journal: International Journal of Molecular Sciences

Article Title: BMP4 Enhances Foam Cell Formation by BMPR-2/Smad1/5/8 Signaling

doi: 10.3390/ijms15045536

Figure Lengend Snippet: BMPR-2 is responsible for BMP4 inhibition of ABCA1 and ABCG1 expression. Unless otherwise specified, all macrophages below were cultured in DMEM with oxLDL. After treatment with PBS (control) or BMP4, macrophages were cultured for 48 h. Then the BMPR-2 expression was detected using RT-PCR ( A ), Western blots ( B ) and its quantification with relative densitometry of BMPR2/β-actin ( C ). After treatment with PBS (control), mock, or BMPR-2 siRNA, the expression of BMPR-2 in macrophages were evaluated using Western blots ( D ) and its quantification with relative densitometry of BMPR2/β-actin ( E ), then the ABCA1 and ABCG1 expression were using RT-PCR ( F ), Western blots ( G ) and its relative quantification over β-actin ( H ). * p < 0.05.

Article Snippet: The membranes were subsequently probed individually with 1:150 polyclonal primary ABCA1 antibody, ABCG1 antibody, SR-A antibody, CD36 antibody, or BMPR-2 antibody (BD Transduction Laboratories, San Jose, CA, USA) or 1:1000 SMAD1/5/8 antibody (Cell Signaling Technology, Beverly, MA, USA).

Techniques: Inhibition, Expressing, Cell Culture, Reverse Transcription Polymerase Chain Reaction, Western Blot

Journal: International Journal of Molecular Sciences

Article Title: BMP4 Enhances Foam Cell Formation by BMPR-2/Smad1/5/8 Signaling

doi: 10.3390/ijms15045536

Figure Lengend Snippet: BMP4 induces foam cell formation by BMPR-2/Smad1/5/8 signaling. Unless otherwise specified, macrophages of murine RAW264.7 cell line below were cultured in DMEM with oxLDL. After treatment with PBS (control) or BMP4, macrophages were cultured for 48 h. Then the expression of p-Smad1/5/8 and Smad1 were evaluated using Western blot ( A ) and quantified with relative densitometry over β-actin ( B ). Then, macrophages were treated with PBS (control), mock, or BMPR-2 siRNA, the expression of BMPR-2 in macrophages were evaluated using Western blots ( C ) and quantified with relative densitometry ( D ). In addition that the macrophages were treated with PBS (control), DMSO, or Smad siRNA, p-Smad1/5/8 and Smad1 were evaluated using Western blot ( E ) and quantified with relative densitometry over β-actin ( F ), then the ABCA1 and ABCG1 expression were evaluated using Western blots ( G ) and its relative quantification over β-actin ( H ) and RT-PCR ( I ). * p < 0.05.

Article Snippet: The membranes were subsequently probed individually with 1:150 polyclonal primary ABCA1 antibody, ABCG1 antibody, SR-A antibody, CD36 antibody, or BMPR-2 antibody (BD Transduction Laboratories, San Jose, CA, USA) or 1:1000 SMAD1/5/8 antibody (Cell Signaling Technology, Beverly, MA, USA).

Techniques: Cell Culture, Expressing, Western Blot, Reverse Transcription Polymerase Chain Reaction