Journal: Frontiers in Cell and Developmental Biology

Article Title: TGF-β-Induced Endothelial to Mesenchymal Transition Is Determined by a Balance Between SNAIL and ID Factors

doi: 10.3389/fcell.2021.616610

Figure Lengend Snippet: TGF–β2 induces EndMT whilst BMP9 does not. (A) Assessing cell morphological changes induced by TGF–β2 and BMP9. Brightfield microscopy images of MS–1 (upper panel) and 2H11 (lower panel) cells showing distinct cell morphologies (i.e., cobblestone or fibroblast–like) after TGF–β2 (1 ng ml –1 ) and BMP9 (5 ng ml –1 ) treatments for 3 days. Scale bar: 200 μm. (B–D) RT-qPCR analysis of endothelial and mesenchymal markers in MS-1 cells. MS-1 cells were exposed to medium containing TGF-β2 (1 ng ml –1 ) or BMP9 (5 ng ml –1 ) or medium containing ligand buffer (control) for 3 days. The expression of mesenchymal cell marker genes Acta2 (B) and Tagln (C) and endothelial cell marker gene Kdr (D) was quantified by RT-qPCR. Expression levels were normalized to those of the housekeeping gene Gapdh . Results are expressed as mean ± SD. Representative results from three independent experiments are shown. (E) Fluorescence microscopy analysis of endothelial and mesenchymal markers in MS-1 and 2H11 cells. MS-1 and 2H11 cells were incubated in medium containing TGF-β2 (1 ng ml –1 ) or BMP9 (5 ng ml –1 ) or medium containing ligand buffer (control) for 3 days. The expression of the endothelial cell marker PECAM-1 (green) and mesenchymal cell marker SM22α (red) in nuclei (blue) stained MS-1 cells (upper panel) and mesenchymal cell markers F-actin (green) and SM22α (red) in nuclei (blue) stained 2H11 cells (lower panel) were assessed by using immunofluorescent staining, respectively. Representative results from at least three independent experiments are shown. Scale bar: 50 μm.

Article Snippet: Human BMP9 (3209–BP/CF) and mouse BMP9 (5566–BP) were obtained from R&D systems.

Techniques: Microscopy, Quantitative RT-PCR, Control, Expressing, Marker, Fluorescence, Incubation, Staining

Journal: Frontiers in Cell and Developmental Biology

Article Title: TGF-β-Induced Endothelial to Mesenchymal Transition Is Determined by a Balance Between SNAIL and ID Factors

doi: 10.3389/fcell.2021.616610

Figure Lengend Snippet: Effects of TGF-β2 and BMP9 on SNAIL and SLUG expression. (A) RT-qPCR analysis of the effects of TGF-β2 (1 ng ml –1 ) and BMP9 (5 ng ml –1 ) on Snail mRNA expression after 3, 6, and 24 h treatments in MS-1 cells. (B) Western blot analysis of the effects of TGF-β2 (1 ng ml –1 ) and BMP9 (5 ng ml –1 ) on SNAIL and SLUG protein expression after 6 and 24 h treatments in MS-1 cells. (C) RT-qPCR analysis of the effects of TGF-β2 (1 ng ml –1 ) and BMP9 (5 ng ml –1 ) on Slug mRNA expression after 3, 6, and 24 h treatments in MS-1 cells. (D) RT-qPCR analysis of the effects of TGF-β2 (1 ng ml –1 ) and BMP9 (5 ng ml –1 ) on Snail mRNA expression after 3 days treatments in MS-1 cells. (E) Western blot analysis of the effects of TGF-β2 (1 ng ml –1 ) and BMP9 (5 ng ml –1 ) on SNAIL protein expression after 3 days treatments in MS-1 cells. (F) RT-qPCR analysis of the effects of TGF-β2 (1 ng ml –1 ) and BMP9 (5 ng ml –1 ) on Snail mRNA expression after 3, 6, and 24 h treatments in 2H11 cells. (G) Western blot analysis of the effects of TGF-β2 (1 ng ml –1 ) and BMP9 (5 ng ml –1 ) on SNAIL and SLUG proteins expression after 6 and 24 h treatments in 2H11 cells. (H) RT-qPCR analysis of the effects of TGF-β2 (1 ng ml –1 ) and BMP9 (5 ng ml –1 ) on Slug mRNA expression after 3, 6, and 24 h treatments in 2H11 cells. (I) RT-qPCR analysis of the effects of TGF-β2 (1 ng ml –1 ) and BMP9 (5 ng ml –1 ) on Snail mRNA expression after 3 days treatments in 2H11 cells. (J) Western blot analysis of the effects of TGF-β2 (1 ng ml –1 ) and BMP9 (5 ng ml –1 ) on SNAIL and SLUG proteins expression after 3 days treatments in 2H11 cells. (K) RT-qPCR analysis of the effects of TGF-β2 (1 ng ml –1 ) and BMP9 (5 ng ml –1 ) on Slug mRNA expression after 3 days treatments in 2H11 cells. All the mRNA expression levels were normalized to those of the housekeeping gene Gapdh . Results are expressed as mean ± SD. Representative results from three independent experiments are shown.

Article Snippet: Human BMP9 (3209–BP/CF) and mouse BMP9 (5566–BP) were obtained from R&D systems.

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: Frontiers in Cell and Developmental Biology

Article Title: TGF-β-Induced Endothelial to Mesenchymal Transition Is Determined by a Balance Between SNAIL and ID Factors

doi: 10.3389/fcell.2021.616610

Figure Lengend Snippet: BMP9 induces Id1 , Id2 , and Id3 expression, but not TGF-β2, in both MS-1 and 2H11 cells. (A–C) RT-qPCR analysis of the effects of BMP9 (5 ng ml –1 ) and TGF-β2 (1 ng ml –1 ) on Id1 (A) , Id2 (B) and Id3 (C) mRNA expression after 3, 6, and 24 h treatments in MS-1 cells. (D–F) RT-qPCR analysis of the effects of BMP9 (5 ng ml –1 ) and TGF-β2 (1 ng ml –1 ) on Id1 (D) , Id2 (E) , and Id3 (F) mRNA expression after 3, 6, and 24 h treatments in 2H11 cells. Expression levels were normalized to those of the housekeeping gene Gapdh . Results are expressed as mean ± SD. Representative results from three independent experiments are shown.

Article Snippet: Human BMP9 (3209–BP/CF) and mouse BMP9 (5566–BP) were obtained from R&D systems.

Techniques: Expressing, Quantitative RT-PCR

Journal: Frontiers in Cell and Developmental Biology

Article Title: TGF-β-Induced Endothelial to Mesenchymal Transition Is Determined by a Balance Between SNAIL and ID Factors

doi: 10.3389/fcell.2021.616610

Figure Lengend Snippet: ID proteins are critical in the BMP9-dependent maintenance of endothelial cell phenotypes. (A–C) RT-qPCR analysis of the siRNA-mediated silencing of endogenous and BMP9-induced Id1 (A) , Id2 (B) and Id3 (C) mRNA expression in MS-1 cells. Expression levels were normalized to those of the housekeeping gene Gapdh . Results are expressed as mean ± SD. Representative results from three independent experiments are shown. (D) Fluorescence microscopy analysis of endothelial and mesenchymal markers in MS-1 cells. Non-targeting knockdown, Id1 knockdown, Id2 knockdown and Id3 knockdown MS-1 cells were incubated in medium containing BMP9 (5 ng ml –1 ) or medium containing ligand buffer (control) for 3 days. The expression of endothelial cell marker PECAM-1 (green) and mesenchymal cell marker SM22α (red) in nuclei (blue) stained MS-1 cells were assessed by using immunofluorescent staining. Scale bar: 50 μm. (E–G) RT-qPCR analysis of the siRNA-mediated silencing of endogenous and BMP9-induced Id1 (E) , Id2 (F) , and Id3 (G) mRNA expression in MS-1 cells. The expression levels were normalized to those of the housekeeping gene Gapdh . Results are expressed as mean ± SD. Representative results from three independent experiments are shown. (H) Fluorescence microscopy analysis of mesenchymal marker in 2H11 cells. Non-targeting knockdown, Id1 knockdown, Id2 knockdown and Id3 knockdown 2H11 cells were incubated in medium containing BMP9 (5 ng ml –1 ) or medium containing ligand buffer (control) for 3 days. The expression of mesenchymal cell marker SM22α (red) in nuclei (blue) stained 2H11 cells was assessed by using immunofluorescent staining. Scale bar: 50 μm. (I) Quantified mean fluorescence intensity of SM22α. At least six representative images from three repeated experiments were quantified. Results are expressed as mean ± SD. NS, not significant; ** p < 0.005.

Article Snippet: Human BMP9 (3209–BP/CF) and mouse BMP9 (5566–BP) were obtained from R&D systems.

Techniques: Quantitative RT-PCR, Expressing, Fluorescence, Microscopy, Knockdown, Incubation, Control, Marker, Staining

Journal: Frontiers in Cell and Developmental Biology

Article Title: TGF-β-Induced Endothelial to Mesenchymal Transition Is Determined by a Balance Between SNAIL and ID Factors

doi: 10.3389/fcell.2021.616610

Figure Lengend Snippet: BMP9 does not antagonize TGF-β2-induced EndMT. (A) Fluorescence microscopy analysis of endothelial and mesenchymal markers in MS-1 cells. MS-1 cells were incubated in medium containing ligand buffer (control, C) or TGF-β2 (1 ng ml –1 , T) and BMP9 (5 ng ml –1 , B) for 4 days, or firstly incubated with TGF-β2 (1 ng ml –1 ) for 2 days and then change to BMP9 (5 ng ml –1 ) for 2 days, or firstly incubated with BMP9 (5 ng ml –1 ) for 2 days and then change to TGF-β2 (1 ng ml –1 ) for 2 days. The expression of the endothelial cell marker PECAM-1 (green) and the mesenchymal cell marker SM22α (red) in nuclei (blue) stained MS-1 cells were assessed by using immunofluorescent staining. Scale bar: 50 μm. (B,C) Quantified mean fluorescence intensity of PECAM-1 (B) and SM22α (C) . At least four representative images from three independent experiments were quantified. Results are expressed as mean ± SD. NS, not significant; * p < 0.05, ** p < 0.005. (D) Western blot analysis of the effects of TGF-β2 (1 ng ml –1 ) or/and BMP9 (5 ng ml –1 ) on pSmad1/5, pSMAD2, SNAIL, and ID1 proteins expression after 1 and 4 h treatments in MS-1 cells.

Article Snippet: Human BMP9 (3209–BP/CF) and mouse BMP9 (5566–BP) were obtained from R&D systems.

Techniques: Fluorescence, Microscopy, Incubation, Control, Expressing, Marker, Staining, Western Blot

Journal: Frontiers in Cell and Developmental Biology

Article Title: TGF-β-Induced Endothelial to Mesenchymal Transition Is Determined by a Balance Between SNAIL and ID Factors

doi: 10.3389/fcell.2021.616610

Figure Lengend Snippet: Schematic of a working model by which TGF-β and BMP9 regulate EndMT. TGF-β/SMAD signaling induces EndMT by promoting SNAIL expression. BMP9/SMAD signaling promotes SNAIL and ID1/2/3 expression, and stimulates EndMT only when BMP9-induced expression of Id1/2/3 is silenced.

Article Snippet: Human BMP9 (3209–BP/CF) and mouse BMP9 (5566–BP) were obtained from R&D systems.

Techniques: Expressing

Journal: bioRxiv

Article Title: An angiopoietin-2 vaccine improves arteriovenous malformation pathology in hereditary hemorrhagic telangiectasia mice

doi: 10.1101/2025.10.13.682178

Figure Lengend Snippet: ( A ) Schematic diagram of the vaccination schedule and generation of the BMP9/10ib model. ( B ) Protein sequence alignment of the C-terminal end of human ANG2 (hANG2), mouse ANG2 (mANG2), human ANG1 (hANG1), and mouse ANG1 (mANG1), along with the peptide sequences of ANG2-P3 and ANG1-P3. ( C and D ) Serum antibody titers against ANG2-P3 (C) and ANG1-P3 (D) in ANG2-P3:CRM197-vaccinated females (Vac-1 to Vac-5) and controls [injected with saline (Sal-1 and Sal-2) or CRM197-only (CRM-1 to CRM-5)]. The vaccinated females with the highest anti-ANG2-P3 titers were identified as “best responders” (marked with a red box). OD, optical density.

Article Snippet: The anti-BMP9 and anti-BMP10 antibody ELISAs were performed as follows: 96-well ELISA plates (Maxisorp, Nunc) were coated with 100 μL per well of recombinant BMP9 (3209-BP-010, R&D Systems) or BMP10 (2926-BP-025, R&D Systems) at 1 μg/mL in a coating buffer (15 mM K2HPO4, 25 mM KH2PO4, 0.1 M NaCl, 0.1 mM EDTA, and 7.5 mM NaN3), and incubated overnight at 4°C.

Techniques: Sequencing, Injection, Saline

Journal: bioRxiv

Article Title: An angiopoietin-2 vaccine improves arteriovenous malformation pathology in hereditary hemorrhagic telangiectasia mice

doi: 10.1101/2025.10.13.682178

Figure Lengend Snippet: ( A ) Serum antibody titers against ANG2-P3 in pups and their corresponding dams vaccinated with ANG2-P3:CRM197 (Vac-1 to Vac-3) or injected with saline (Saline). ( B ) Representative immunofluorescence staining with isolectin B4 (IB4, green) and of α-smooth muscle actin (SMA, red) in P6 retinas of pups treated with PBS or BMP9/10ib, from a dam vaccinated with ANG2-P3:CRM197 or injected with saline. a, artery; v, vein. Scale bar, 1.5 mm. ( C and D) AVM count per retina (C) and retinal AVM surface area (D) in BMP9/10ib pups from dams vaccinated with ANG2-P3:CRM197 or injected with saline (Saline). ( E ) Spearman’s rank correlation matrix of the indicated variables. ( F-H ) retinal artery diameter (F), retinal vein diameter (G), and SMA coverage area (H) in BMP9/10ib pups from dams vaccinated with ANG2-P3:CRM197 or injected with saline (Saline). Data are shown as meanLJ±LJs.e.m.; unpaired t-test with Welch’s correction (C), Mann-Whitney test (D), and one-way ANOVA with Tukey’s multiple comparisons test (F-H). ns, not significant; * P < 0.05; *** P ≤ 0.001; **** P < 0.0001.

Article Snippet: The anti-BMP9 and anti-BMP10 antibody ELISAs were performed as follows: 96-well ELISA plates (Maxisorp, Nunc) were coated with 100 μL per well of recombinant BMP9 (3209-BP-010, R&D Systems) or BMP10 (2926-BP-025, R&D Systems) at 1 μg/mL in a coating buffer (15 mM K2HPO4, 25 mM KH2PO4, 0.1 M NaCl, 0.1 mM EDTA, and 7.5 mM NaN3), and incubated overnight at 4°C.

Techniques: Injection, Saline, Immunofluorescence, Staining, MANN-WHITNEY

Journal: Immunity

Article Title: Stellate Cells, Hepatocytes, and Endothelial Cells Imprint the Kupffer Cell Identity on Monocytes Colonizing the Liver Macrophage Niche

doi: 10.1016/j.immuni.2019.08.017

Figure Lengend Snippet:

Article Snippet: Recombinant Human/Mouse/Rat BMP-9 Protein , RnD Systems , Cat#5566-BP-010.

Techniques: Control, Recombinant, Blocking Assay, Irradiation, Enzyme-linked Immunosorbent Assay, cDNA Synthesis, Microarray, Software, Microscopy