human alk-1 antibody Search Results


anti-human alk1  (R&D Systems)
90
R&D Systems anti-human alk1
Anti Human Alk1, supplied by R&D Systems, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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anti psmad1 5  (R&D Systems)
94
R&D Systems anti psmad1 5
Anti Psmad1 5, supplied by R&D Systems, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 94 stars, based on 1 article reviews
anti psmad1 5 - by Bioz Stars, 2026-03
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human alk1 inhibitory antibody  (R&D Systems)
92
R&D Systems human alk1 inhibitory antibody
Hepsin regulates EGFR signaling in a TGFβ‐dependent manner. (A) Western blot comparison of total‐EGFR (tEGFR) levels in control MCF10A pL6‐Ctrl ( N = 3) and MCF10A pL6‐TGFβ1V5, V5‐tagged TGFβ1 overexpressing cells ( N = 3), which secrete around 80–100 pg·mL −1 of TGFβ1. Data are presented as mean ± SD. Significance was tested using the student's t‐test. (B) Western blot analysis of tEGFR in control doxycycline‐ (DOX − ) ( N = 3) and hepsin overexpressing (DOX + ; N = 3) MCF10A‐ pIND20‐HPN pL6‐ TGFβ1V5 cells. Data are presented as mean ± SD. Significance was tested with the student's t ‐test. (C) Western blot analysis of tEGFR, phospho‐SMAD2/3 (pSMAD2/3), and total‐SMAD2 (tSMAD2) in MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 cells. The cells were treated with ALK5 inhibitor (i) (10 μ m RepSox, N = 4) and ALK4/5/7 inhibitor (5 μ m A‐83‐01, N = 3) or DMSO (control, N = 4) for 48 h with (DOX + ; 1 μg·mL −1 ) or without (DOX − , control) hepsin overexpression (the numbers below the tEGFR blot indicate loading normalized values for tEGFR band intensity fold leftmost control lane). The histogram data are presented as mean ± SD. Significance was tested using unpaired t ‐test. (D) Western blot analysis of phospho‐EGFR (pEGFR), tEGFR, total‐MAPK (tMAPK) and phospho‐MAPK (pMAPK), and vinculin (loading control) in MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 cells. The cells were treated with the <t>ALK1/2/3/6</t> inhibitor (i) (10 μ m K02288), EGFR inhibitor (i) (10 μ m Erlotinib), and DMSO (control) for 48 h. Data are presented as mean ± SD. Significance was tested with the student's t ‐test ( N = 3). (E) Phase‐contrast microscopy images of MCF10A‐pIND20‐HPN cells and MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 (overexpressing TGFβ1) cells grown in 3D culture for 2 weeks with (DOX + ; 1 μg·mL −1 ) or without (DOX − ) induction of hepsin overexpression. Cells were treated with 10 μ m EGFR or ALK1/2/3/6 inhibitor for 2 weeks as indicated in the figure. Scale bar represents 100 μm. (F) Phase‐contrast microscopy images of MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 cells grown in 3D culture for 2 weeks in the presence of ALK1 <t>inhibitory</t> antibody (25 μg·mL −1 ). Scale bar represents 50 μm. For E and F, the experiments were repeated three times, with at least 100 epithelial structures counted per group in each repeat (one dot represents one structure), and black lines denote the mean values of each group. Significance was tested using the student's t ‐test. (G) Western blot analysis of the mitotic marker phospho histone H3 (pH3 S10) MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 cell line grown in 3D culture in the presence of the EGFRi (1 μ m , 24 h) or DMSO. The quantification of blots of three independent experiments is shown in the graph (on the right), where the black lines denote the mean of each group. Significance was tested using the student's t ‐test. ns, not significant.
Human Alk1 Inhibitory Antibody, supplied by R&D Systems, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/human alk1 inhibitory antibody/product/R&D Systems
Average 92 stars, based on 1 article reviews
human alk1 inhibitory antibody - by Bioz Stars, 2026-03
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goat anti human alk1  (R&D Systems)
93
R&D Systems goat anti human alk1
GATA6 expression is induced via <t>BMP10-BMPR2/ALK1</t> axis. ( A–F ) HPAECs were transfected with ALK1, BMPR2, Endoglin, or control scr siRNA for 48 h, and then treated with 10 ng/ml BMP10 or vehicle for 6 h for RNA isolation and 24 h for protein isolation. ( A,C,E ): GATA6 mRNA measured by qPCR. Data are means ± SE; each experiment was repeated at least three times. ( B,D,F ): GATA6 protein levels were measured by immunoblot analysis. Data are means ± SE, each experiment was repeated at least three times. Representative blots are shown. *p < 0.05, **p < 0.01, ***p < 0.001 by Kruskal–Wallis test with post hoc Dunn’s test for multiple comparisons. ( G–I ) Human PAH PASMC were treated with 10 ng/ml BMP10 or vehicle ( − ) for 48 h and immunoblot analysis to detect indicated proteins was performed. Data are means ± SE from n = 4 subjects/group. *p < 0.05 by Mann Whitney U test. ( J,K ) Equal amounts of human PAH HPAEC and PASMC were plated at 6-well plates and treated with 10 ng/ml BMP10 or vehicle ( − ). 48 h later cell counts were performed. Data are means ± SE from n = 3 subjects/group, 3 technical repetitions/subject. *p < 0.05 by Mann Whitney U test. ( L,M ) HPAECs were transfected with SMAD1 siRNA, and then treated with BMP10 for 6 h. GATA6 and SMAD1 mRNA levels were measured by qPCR. Data shown as means ± SE. Each experiment was repeated at least three times. *p < 0.05, **p < 0.01 by Kruskal–Wallis test with post-hoc correction for multiple comparisons. ( N,O ) HPAEC were treated for 30 min with diluent ( − ), 5 µM ERK1/2 inhibitor SCH772984 (ERKi), or 5 µM GSK3 inhibitor CHIR99021 (GSK3i) and then stimulated with BMP10 (10 ng/ml) or vehicle for 24 h. Representative immunoblots ( N ) and statistical analysis ( O ) are shown. ( O ): Data represent GATA6/β-actin ratio. Data are means ± SE from five independent experiments. *p < 0.05 by Kruskal–Wallis test with post-hoc Dunn’s correction for multiple comparisons. ( E ) HPAECs were treated with BMP10 in the presence or absence of 10 µM ERK1/2 inhibitor SCH772984 for 24 h. GATA6 mRNA levels were measured by qPCR. Data shown as means ± SE. Each experiment was repeated six times. *p < 0.05 by Kruskal–Wallis test with post-hoc Dunn’s correction for multiple comparisons. The original blots are presented in Supplementary Fig. .
Goat Anti Human Alk1, supplied by R&D Systems, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/goat anti human alk1/product/R&D Systems
Average 93 stars, based on 1 article reviews
goat anti human alk1 - by Bioz Stars, 2026-03
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anti human tgf 1  (R&D Systems)
86
R&D Systems anti human tgf 1
GATA6 expression is induced via <t>BMP10-BMPR2/ALK1</t> axis. ( A–F ) HPAECs were transfected with ALK1, BMPR2, Endoglin, or control scr siRNA for 48 h, and then treated with 10 ng/ml BMP10 or vehicle for 6 h for RNA isolation and 24 h for protein isolation. ( A,C,E ): GATA6 mRNA measured by qPCR. Data are means ± SE; each experiment was repeated at least three times. ( B,D,F ): GATA6 protein levels were measured by immunoblot analysis. Data are means ± SE, each experiment was repeated at least three times. Representative blots are shown. *p < 0.05, **p < 0.01, ***p < 0.001 by Kruskal–Wallis test with post hoc Dunn’s test for multiple comparisons. ( G–I ) Human PAH PASMC were treated with 10 ng/ml BMP10 or vehicle ( − ) for 48 h and immunoblot analysis to detect indicated proteins was performed. Data are means ± SE from n = 4 subjects/group. *p < 0.05 by Mann Whitney U test. ( J,K ) Equal amounts of human PAH HPAEC and PASMC were plated at 6-well plates and treated with 10 ng/ml BMP10 or vehicle ( − ). 48 h later cell counts were performed. Data are means ± SE from n = 3 subjects/group, 3 technical repetitions/subject. *p < 0.05 by Mann Whitney U test. ( L,M ) HPAECs were transfected with SMAD1 siRNA, and then treated with BMP10 for 6 h. GATA6 and SMAD1 mRNA levels were measured by qPCR. Data shown as means ± SE. Each experiment was repeated at least three times. *p < 0.05, **p < 0.01 by Kruskal–Wallis test with post-hoc correction for multiple comparisons. ( N,O ) HPAEC were treated for 30 min with diluent ( − ), 5 µM ERK1/2 inhibitor SCH772984 (ERKi), or 5 µM GSK3 inhibitor CHIR99021 (GSK3i) and then stimulated with BMP10 (10 ng/ml) or vehicle for 24 h. Representative immunoblots ( N ) and statistical analysis ( O ) are shown. ( O ): Data represent GATA6/β-actin ratio. Data are means ± SE from five independent experiments. *p < 0.05 by Kruskal–Wallis test with post-hoc Dunn’s correction for multiple comparisons. ( E ) HPAECs were treated with BMP10 in the presence or absence of 10 µM ERK1/2 inhibitor SCH772984 for 24 h. GATA6 mRNA levels were measured by qPCR. Data shown as means ± SE. Each experiment was repeated six times. *p < 0.05 by Kruskal–Wallis test with post-hoc Dunn’s correction for multiple comparisons. The original blots are presented in Supplementary Fig. .
Anti Human Tgf 1, supplied by R&D Systems, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/anti human tgf 1/product/R&D Systems
Average 86 stars, based on 1 article reviews
anti human tgf 1 - by Bioz Stars, 2026-03
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mouse anti-human alk1 monoclonal antibody  (DaAn Gene)
90
DaAn Gene mouse anti-human alk1 monoclonal antibody
GATA6 expression is induced via <t>BMP10-BMPR2/ALK1</t> axis. ( A–F ) HPAECs were transfected with ALK1, BMPR2, Endoglin, or control scr siRNA for 48 h, and then treated with 10 ng/ml BMP10 or vehicle for 6 h for RNA isolation and 24 h for protein isolation. ( A,C,E ): GATA6 mRNA measured by qPCR. Data are means ± SE; each experiment was repeated at least three times. ( B,D,F ): GATA6 protein levels were measured by immunoblot analysis. Data are means ± SE, each experiment was repeated at least three times. Representative blots are shown. *p < 0.05, **p < 0.01, ***p < 0.001 by Kruskal–Wallis test with post hoc Dunn’s test for multiple comparisons. ( G–I ) Human PAH PASMC were treated with 10 ng/ml BMP10 or vehicle ( − ) for 48 h and immunoblot analysis to detect indicated proteins was performed. Data are means ± SE from n = 4 subjects/group. *p < 0.05 by Mann Whitney U test. ( J,K ) Equal amounts of human PAH HPAEC and PASMC were plated at 6-well plates and treated with 10 ng/ml BMP10 or vehicle ( − ). 48 h later cell counts were performed. Data are means ± SE from n = 3 subjects/group, 3 technical repetitions/subject. *p < 0.05 by Mann Whitney U test. ( L,M ) HPAECs were transfected with SMAD1 siRNA, and then treated with BMP10 for 6 h. GATA6 and SMAD1 mRNA levels were measured by qPCR. Data shown as means ± SE. Each experiment was repeated at least three times. *p < 0.05, **p < 0.01 by Kruskal–Wallis test with post-hoc correction for multiple comparisons. ( N,O ) HPAEC were treated for 30 min with diluent ( − ), 5 µM ERK1/2 inhibitor SCH772984 (ERKi), or 5 µM GSK3 inhibitor CHIR99021 (GSK3i) and then stimulated with BMP10 (10 ng/ml) or vehicle for 24 h. Representative immunoblots ( N ) and statistical analysis ( O ) are shown. ( O ): Data represent GATA6/β-actin ratio. Data are means ± SE from five independent experiments. *p < 0.05 by Kruskal–Wallis test with post-hoc Dunn’s correction for multiple comparisons. ( E ) HPAECs were treated with BMP10 in the presence or absence of 10 µM ERK1/2 inhibitor SCH772984 for 24 h. GATA6 mRNA levels were measured by qPCR. Data shown as means ± SE. Each experiment was repeated six times. *p < 0.05 by Kruskal–Wallis test with post-hoc Dunn’s correction for multiple comparisons. The original blots are presented in Supplementary Fig. .
Mouse Anti Human Alk1 Monoclonal Antibody, supplied by DaAn Gene, used in various techniques. Bioz Stars score: 90/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/mouse anti-human alk1 monoclonal antibody/product/DaAn Gene
Average 90 stars, based on 1 article reviews
mouse anti-human alk1 monoclonal antibody - by Bioz Stars, 2026-03
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Image Search Results


Hepsin regulates EGFR signaling in a TGFβ‐dependent manner. (A) Western blot comparison of total‐EGFR (tEGFR) levels in control MCF10A pL6‐Ctrl ( N = 3) and MCF10A pL6‐TGFβ1V5, V5‐tagged TGFβ1 overexpressing cells ( N = 3), which secrete around 80–100 pg·mL −1 of TGFβ1. Data are presented as mean ± SD. Significance was tested using the student's t‐test. (B) Western blot analysis of tEGFR in control doxycycline‐ (DOX − ) ( N = 3) and hepsin overexpressing (DOX + ; N = 3) MCF10A‐ pIND20‐HPN pL6‐ TGFβ1V5 cells. Data are presented as mean ± SD. Significance was tested with the student's t ‐test. (C) Western blot analysis of tEGFR, phospho‐SMAD2/3 (pSMAD2/3), and total‐SMAD2 (tSMAD2) in MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 cells. The cells were treated with ALK5 inhibitor (i) (10 μ m RepSox, N = 4) and ALK4/5/7 inhibitor (5 μ m A‐83‐01, N = 3) or DMSO (control, N = 4) for 48 h with (DOX + ; 1 μg·mL −1 ) or without (DOX − , control) hepsin overexpression (the numbers below the tEGFR blot indicate loading normalized values for tEGFR band intensity fold leftmost control lane). The histogram data are presented as mean ± SD. Significance was tested using unpaired t ‐test. (D) Western blot analysis of phospho‐EGFR (pEGFR), tEGFR, total‐MAPK (tMAPK) and phospho‐MAPK (pMAPK), and vinculin (loading control) in MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 cells. The cells were treated with the ALK1/2/3/6 inhibitor (i) (10 μ m K02288), EGFR inhibitor (i) (10 μ m Erlotinib), and DMSO (control) for 48 h. Data are presented as mean ± SD. Significance was tested with the student's t ‐test ( N = 3). (E) Phase‐contrast microscopy images of MCF10A‐pIND20‐HPN cells and MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 (overexpressing TGFβ1) cells grown in 3D culture for 2 weeks with (DOX + ; 1 μg·mL −1 ) or without (DOX − ) induction of hepsin overexpression. Cells were treated with 10 μ m EGFR or ALK1/2/3/6 inhibitor for 2 weeks as indicated in the figure. Scale bar represents 100 μm. (F) Phase‐contrast microscopy images of MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 cells grown in 3D culture for 2 weeks in the presence of ALK1 inhibitory antibody (25 μg·mL −1 ). Scale bar represents 50 μm. For E and F, the experiments were repeated three times, with at least 100 epithelial structures counted per group in each repeat (one dot represents one structure), and black lines denote the mean values of each group. Significance was tested using the student's t ‐test. (G) Western blot analysis of the mitotic marker phospho histone H3 (pH3 S10) MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 cell line grown in 3D culture in the presence of the EGFRi (1 μ m , 24 h) or DMSO. The quantification of blots of three independent experiments is shown in the graph (on the right), where the black lines denote the mean of each group. Significance was tested using the student's t ‐test. ns, not significant.

Journal: Molecular Oncology

Article Title: Hepsin promotes breast tumor growth signaling via the TGFβ‐EGFR axis

doi: 10.1002/1878-0261.13545

Figure Lengend Snippet: Hepsin regulates EGFR signaling in a TGFβ‐dependent manner. (A) Western blot comparison of total‐EGFR (tEGFR) levels in control MCF10A pL6‐Ctrl ( N = 3) and MCF10A pL6‐TGFβ1V5, V5‐tagged TGFβ1 overexpressing cells ( N = 3), which secrete around 80–100 pg·mL −1 of TGFβ1. Data are presented as mean ± SD. Significance was tested using the student's t‐test. (B) Western blot analysis of tEGFR in control doxycycline‐ (DOX − ) ( N = 3) and hepsin overexpressing (DOX + ; N = 3) MCF10A‐ pIND20‐HPN pL6‐ TGFβ1V5 cells. Data are presented as mean ± SD. Significance was tested with the student's t ‐test. (C) Western blot analysis of tEGFR, phospho‐SMAD2/3 (pSMAD2/3), and total‐SMAD2 (tSMAD2) in MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 cells. The cells were treated with ALK5 inhibitor (i) (10 μ m RepSox, N = 4) and ALK4/5/7 inhibitor (5 μ m A‐83‐01, N = 3) or DMSO (control, N = 4) for 48 h with (DOX + ; 1 μg·mL −1 ) or without (DOX − , control) hepsin overexpression (the numbers below the tEGFR blot indicate loading normalized values for tEGFR band intensity fold leftmost control lane). The histogram data are presented as mean ± SD. Significance was tested using unpaired t ‐test. (D) Western blot analysis of phospho‐EGFR (pEGFR), tEGFR, total‐MAPK (tMAPK) and phospho‐MAPK (pMAPK), and vinculin (loading control) in MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 cells. The cells were treated with the ALK1/2/3/6 inhibitor (i) (10 μ m K02288), EGFR inhibitor (i) (10 μ m Erlotinib), and DMSO (control) for 48 h. Data are presented as mean ± SD. Significance was tested with the student's t ‐test ( N = 3). (E) Phase‐contrast microscopy images of MCF10A‐pIND20‐HPN cells and MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 (overexpressing TGFβ1) cells grown in 3D culture for 2 weeks with (DOX + ; 1 μg·mL −1 ) or without (DOX − ) induction of hepsin overexpression. Cells were treated with 10 μ m EGFR or ALK1/2/3/6 inhibitor for 2 weeks as indicated in the figure. Scale bar represents 100 μm. (F) Phase‐contrast microscopy images of MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 cells grown in 3D culture for 2 weeks in the presence of ALK1 inhibitory antibody (25 μg·mL −1 ). Scale bar represents 50 μm. For E and F, the experiments were repeated three times, with at least 100 epithelial structures counted per group in each repeat (one dot represents one structure), and black lines denote the mean values of each group. Significance was tested using the student's t ‐test. (G) Western blot analysis of the mitotic marker phospho histone H3 (pH3 S10) MCF10A‐pIND20‐HPN pL6‐TGFβ1V5 cell line grown in 3D culture in the presence of the EGFRi (1 μ m , 24 h) or DMSO. The quantification of blots of three independent experiments is shown in the graph (on the right), where the black lines denote the mean of each group. Significance was tested using the student's t ‐test. ns, not significant.

Article Snippet: The inhibitors used in this study were K02288 (Selleckchem, S7359, Houston, TX, USA), Galunisertib/LY2157299 (Selleckchem, S2230), RepSox (Sigma‐Aldrich/Merck, R0158), A‐83‐01 (MedChem Express, HY‐10432, Monmouth Junction, NJ, USA), Erlotinib (EGFRi, Selleckchem, S1023), and human ALK1 inhibitory antibody (R&D Systems, MAB3701).

Techniques: Western Blot, Comparison, Control, Over Expression, Microscopy, Marker

GATA6 expression is induced via BMP10-BMPR2/ALK1 axis. ( A–F ) HPAECs were transfected with ALK1, BMPR2, Endoglin, or control scr siRNA for 48 h, and then treated with 10 ng/ml BMP10 or vehicle for 6 h for RNA isolation and 24 h for protein isolation. ( A,C,E ): GATA6 mRNA measured by qPCR. Data are means ± SE; each experiment was repeated at least three times. ( B,D,F ): GATA6 protein levels were measured by immunoblot analysis. Data are means ± SE, each experiment was repeated at least three times. Representative blots are shown. *p < 0.05, **p < 0.01, ***p < 0.001 by Kruskal–Wallis test with post hoc Dunn’s test for multiple comparisons. ( G–I ) Human PAH PASMC were treated with 10 ng/ml BMP10 or vehicle ( − ) for 48 h and immunoblot analysis to detect indicated proteins was performed. Data are means ± SE from n = 4 subjects/group. *p < 0.05 by Mann Whitney U test. ( J,K ) Equal amounts of human PAH HPAEC and PASMC were plated at 6-well plates and treated with 10 ng/ml BMP10 or vehicle ( − ). 48 h later cell counts were performed. Data are means ± SE from n = 3 subjects/group, 3 technical repetitions/subject. *p < 0.05 by Mann Whitney U test. ( L,M ) HPAECs were transfected with SMAD1 siRNA, and then treated with BMP10 for 6 h. GATA6 and SMAD1 mRNA levels were measured by qPCR. Data shown as means ± SE. Each experiment was repeated at least three times. *p < 0.05, **p < 0.01 by Kruskal–Wallis test with post-hoc correction for multiple comparisons. ( N,O ) HPAEC were treated for 30 min with diluent ( − ), 5 µM ERK1/2 inhibitor SCH772984 (ERKi), or 5 µM GSK3 inhibitor CHIR99021 (GSK3i) and then stimulated with BMP10 (10 ng/ml) or vehicle for 24 h. Representative immunoblots ( N ) and statistical analysis ( O ) are shown. ( O ): Data represent GATA6/β-actin ratio. Data are means ± SE from five independent experiments. *p < 0.05 by Kruskal–Wallis test with post-hoc Dunn’s correction for multiple comparisons. ( E ) HPAECs were treated with BMP10 in the presence or absence of 10 µM ERK1/2 inhibitor SCH772984 for 24 h. GATA6 mRNA levels were measured by qPCR. Data shown as means ± SE. Each experiment was repeated six times. *p < 0.05 by Kruskal–Wallis test with post-hoc Dunn’s correction for multiple comparisons. The original blots are presented in Supplementary Fig. .

Journal: Scientific Reports

Article Title: GATA6 coordinates cross-talk between BMP10 and oxidative stress axis in pulmonary arterial hypertension

doi: 10.1038/s41598-023-33779-8

Figure Lengend Snippet: GATA6 expression is induced via BMP10-BMPR2/ALK1 axis. ( A–F ) HPAECs were transfected with ALK1, BMPR2, Endoglin, or control scr siRNA for 48 h, and then treated with 10 ng/ml BMP10 or vehicle for 6 h for RNA isolation and 24 h for protein isolation. ( A,C,E ): GATA6 mRNA measured by qPCR. Data are means ± SE; each experiment was repeated at least three times. ( B,D,F ): GATA6 protein levels were measured by immunoblot analysis. Data are means ± SE, each experiment was repeated at least three times. Representative blots are shown. *p < 0.05, **p < 0.01, ***p < 0.001 by Kruskal–Wallis test with post hoc Dunn’s test for multiple comparisons. ( G–I ) Human PAH PASMC were treated with 10 ng/ml BMP10 or vehicle ( − ) for 48 h and immunoblot analysis to detect indicated proteins was performed. Data are means ± SE from n = 4 subjects/group. *p < 0.05 by Mann Whitney U test. ( J,K ) Equal amounts of human PAH HPAEC and PASMC were plated at 6-well plates and treated with 10 ng/ml BMP10 or vehicle ( − ). 48 h later cell counts were performed. Data are means ± SE from n = 3 subjects/group, 3 technical repetitions/subject. *p < 0.05 by Mann Whitney U test. ( L,M ) HPAECs were transfected with SMAD1 siRNA, and then treated with BMP10 for 6 h. GATA6 and SMAD1 mRNA levels were measured by qPCR. Data shown as means ± SE. Each experiment was repeated at least three times. *p < 0.05, **p < 0.01 by Kruskal–Wallis test with post-hoc correction for multiple comparisons. ( N,O ) HPAEC were treated for 30 min with diluent ( − ), 5 µM ERK1/2 inhibitor SCH772984 (ERKi), or 5 µM GSK3 inhibitor CHIR99021 (GSK3i) and then stimulated with BMP10 (10 ng/ml) or vehicle for 24 h. Representative immunoblots ( N ) and statistical analysis ( O ) are shown. ( O ): Data represent GATA6/β-actin ratio. Data are means ± SE from five independent experiments. *p < 0.05 by Kruskal–Wallis test with post-hoc Dunn’s correction for multiple comparisons. ( E ) HPAECs were treated with BMP10 in the presence or absence of 10 µM ERK1/2 inhibitor SCH772984 for 24 h. GATA6 mRNA levels were measured by qPCR. Data shown as means ± SE. Each experiment was repeated six times. *p < 0.05 by Kruskal–Wallis test with post-hoc Dunn’s correction for multiple comparisons. The original blots are presented in Supplementary Fig. .

Article Snippet: Antibodies were as follows: Goat anti-human GATA6 (R&D; AF1700, 1:500), mouse anti-human GATA6 (Santa-Cruz, 517554, 1:500), Rabbit anti-human/mouse SOD2 (Cell signaling technology; 13141, 1:1000; 13145, 1:5000), Goat anti-human ALK1 (R&D; AF370, 1:500), Rabbit anti-mouse ALK1 (ABGENT AP7807a, 1:1000), rabbit anti-human/mouse ActR2B (LS bio; LS-B7781, 1:1000), mouse anti-human BMPR2 (Novus; NBP2-37624 Clone 3F6; 1:1000), rabbit anti-mouse BMPR2 (Proteintech; 19087–1-AP; 1:1000), mouse anti-mouse/human/rat BMPR2 (Abcam, catalog #130206 1:500), rabbit anti-human/mouse Endoglin (Proteintech; 10862–1-AP; 1:1000), rabbit anti-mouse BiP (Abcam; ab53068; 1:1000), rabbit anti-mouse CHOP (Novus; NBP2-13172; 1:1000), mouse anti-beta-actin (Sigma; A1978; 1:2000), mouse anti-human PRPF4 (Abcam, 69878, 1:1000), rabbit anti-human MYEOV (Invitrogen, PA5-48938 1:1000), rabbit anti-human STING (Invitrogen, PA5-26751, 1:1000), rabbit anti-human/mouse/rat EAF1 (Invitrogen, PA5-100493, 1:1000), rabbit anti-human/mouse/rat Histone H3 (Cell Signaling, 9715 1:1000), rabbit anti-human/mouse/rat a-b Tubulin (Cell Signaling, 2148, 1:1000).

Techniques: Expressing, Transfection, Isolation, Western Blot, MANN-WHITNEY

GATA6 deficiency in PAEC and PASMC results in loss of BMP receptors. ( A ) qPCR of HPAECs transfected with GATA6 or control scr siRNA ( − ) to measure indicated mRNA, each experiment was repeated at least three times. Data are means ± SE, n = 4–7. **p < 0.01, ***p < 0.001 by Mann Whitney U test. ( B ) Chromatin immune precipitation (ChiP) assay in HPAECs, n = 8–10. Representative gel images and data quantification are shown. Data are means ± SE, ***p < 0.001, ****p < 0.0001 by Mann Whitney U test. ( C,D ) Immunoblot analysis of control human PASMC transfected with siRNA GATA6 or control scr siRNA for 48 h. Data are means ± SE, 3 subjects/group, *p < 0.05 by Mann Whitney U test. Please see Fig. F,G for GATA6 immunoblots. ( E,F ) Expression of BmpR2, Alk1 , ActRIIB, and endoglin measured by qPCR in PAEC ( F ) and whole lungs ( G ) from WT and Gata6 CKO mice. Data are means ± SE; E: n = 4–5/group; F: n = 6–11 mice/group. Male and female mice responded similarly. *p < 0.05, **p < 0.01 by Mann Whitney U test (( E,F ) BmpR2, Alk1 , and ActRIIB ) and unpaired τ test (F Endoglin ). ( G,H ) Control HPAECs transfected with siGATA6 or control scr siRNA ( − ) were assayed by immunoblot analysis to detect indicated BMP receptors. Values are means ± SE of the relative protein levels by densitometry, n = 4–7.*p < 0.05, ***p < 0.001 by Mann Whitney U test. ( I,J ) Immunoblot analysis of whole lung tissue from Gata6 CKO and WT mice. Values are means ± SE of the relative protein levels by densitometry, n = 3–5/group. Male and female mice responded similarly. *p < 0.05, **p < 0.01 by Mann Whitney U test. The original blots are presented in Supplementary Fig. .

Journal: Scientific Reports

Article Title: GATA6 coordinates cross-talk between BMP10 and oxidative stress axis in pulmonary arterial hypertension

doi: 10.1038/s41598-023-33779-8

Figure Lengend Snippet: GATA6 deficiency in PAEC and PASMC results in loss of BMP receptors. ( A ) qPCR of HPAECs transfected with GATA6 or control scr siRNA ( − ) to measure indicated mRNA, each experiment was repeated at least three times. Data are means ± SE, n = 4–7. **p < 0.01, ***p < 0.001 by Mann Whitney U test. ( B ) Chromatin immune precipitation (ChiP) assay in HPAECs, n = 8–10. Representative gel images and data quantification are shown. Data are means ± SE, ***p < 0.001, ****p < 0.0001 by Mann Whitney U test. ( C,D ) Immunoblot analysis of control human PASMC transfected with siRNA GATA6 or control scr siRNA for 48 h. Data are means ± SE, 3 subjects/group, *p < 0.05 by Mann Whitney U test. Please see Fig. F,G for GATA6 immunoblots. ( E,F ) Expression of BmpR2, Alk1 , ActRIIB, and endoglin measured by qPCR in PAEC ( F ) and whole lungs ( G ) from WT and Gata6 CKO mice. Data are means ± SE; E: n = 4–5/group; F: n = 6–11 mice/group. Male and female mice responded similarly. *p < 0.05, **p < 0.01 by Mann Whitney U test (( E,F ) BmpR2, Alk1 , and ActRIIB ) and unpaired τ test (F Endoglin ). ( G,H ) Control HPAECs transfected with siGATA6 or control scr siRNA ( − ) were assayed by immunoblot analysis to detect indicated BMP receptors. Values are means ± SE of the relative protein levels by densitometry, n = 4–7.*p < 0.05, ***p < 0.001 by Mann Whitney U test. ( I,J ) Immunoblot analysis of whole lung tissue from Gata6 CKO and WT mice. Values are means ± SE of the relative protein levels by densitometry, n = 3–5/group. Male and female mice responded similarly. *p < 0.05, **p < 0.01 by Mann Whitney U test. The original blots are presented in Supplementary Fig. .

Article Snippet: Antibodies were as follows: Goat anti-human GATA6 (R&D; AF1700, 1:500), mouse anti-human GATA6 (Santa-Cruz, 517554, 1:500), Rabbit anti-human/mouse SOD2 (Cell signaling technology; 13141, 1:1000; 13145, 1:5000), Goat anti-human ALK1 (R&D; AF370, 1:500), Rabbit anti-mouse ALK1 (ABGENT AP7807a, 1:1000), rabbit anti-human/mouse ActR2B (LS bio; LS-B7781, 1:1000), mouse anti-human BMPR2 (Novus; NBP2-37624 Clone 3F6; 1:1000), rabbit anti-mouse BMPR2 (Proteintech; 19087–1-AP; 1:1000), mouse anti-mouse/human/rat BMPR2 (Abcam, catalog #130206 1:500), rabbit anti-human/mouse Endoglin (Proteintech; 10862–1-AP; 1:1000), rabbit anti-mouse BiP (Abcam; ab53068; 1:1000), rabbit anti-mouse CHOP (Novus; NBP2-13172; 1:1000), mouse anti-beta-actin (Sigma; A1978; 1:2000), mouse anti-human PRPF4 (Abcam, 69878, 1:1000), rabbit anti-human MYEOV (Invitrogen, PA5-48938 1:1000), rabbit anti-human STING (Invitrogen, PA5-26751, 1:1000), rabbit anti-human/mouse/rat EAF1 (Invitrogen, PA5-100493, 1:1000), rabbit anti-human/mouse/rat Histone H3 (Cell Signaling, 9715 1:1000), rabbit anti-human/mouse/rat a-b Tubulin (Cell Signaling, 2148, 1:1000).

Techniques: Transfection, MANN-WHITNEY, Western Blot, Expressing

Treatment with DMF restores expression of the BMP receptors, reverses oxidative stress and pulmonary hypertension in Gata6 CKO mice. (DMF or vehicle were administered daily via i.p. injection for 3 weeks. ( A ) qPCR analysis of whole lung tissue from WT and Gata6 CKO mice treated with DMF or vehicle to detect expression of indicated BMP receptors. Data are means ± SE, n = 6–12, *p < 0.05. **p < 0.01 by Kruskal–Wallis test followed by Dunn’s multiple comparisons test ( BmpR2, ActRIIB, Alk1 ) and one-way ANOVA followed by post hoc Tukey’s multiple comparison ( Endoglin ). ( B ) qPCR analysis of whole lung tissue from WT and Gata6 CKO mice treated with DMF or vehicle to detect expression of the antioxidant enzymes and eNOS . Data are means ± SE, n = 6–17, *p < 0.05., **p < 0.01, ***p < 0.001 by one-way ANOVA followed by post hoc Tukey’s multiple comparisons test ( SOD2, GPX1, CAT ) and Kruskal–Wallis test with post hoc Dunn’s multiple comparisons test ( eNOS ). ( C,D ) mRNA levels of indicated BMP receptors and antioxidant enzymes measured by qPCR in PAEC from WT and Gata6 CKO mice treated with DMF or vehicle. Data are means ± SE, n = 3–6 mice/group, *p < 0.05, **p < 0.01 by Kruskal–Wallis test with post hoc Dunn’s multiple comparisons test. ( E–G ) RVSP, pulmonary acceleration time as a fraction of ejection time (PAT/ET) and Fulton index (RV/[LV + S]) were evaluated in WT and Gata6 CKO mice in the presence or absence of DMF. Data are means ± SE. n = 5–11 mice/group. *p < 0.05, **p < 0.01. ***p < 0.001 by Kruskal–Wallis test with post hoc Dunn’s multiple comparisons test (RVSP) and one-way ANOVA followed by post hoc Tukey’s multiple comparisons test (PAT/ET and RV/(LV + S).

Journal: Scientific Reports

Article Title: GATA6 coordinates cross-talk between BMP10 and oxidative stress axis in pulmonary arterial hypertension

doi: 10.1038/s41598-023-33779-8

Figure Lengend Snippet: Treatment with DMF restores expression of the BMP receptors, reverses oxidative stress and pulmonary hypertension in Gata6 CKO mice. (DMF or vehicle were administered daily via i.p. injection for 3 weeks. ( A ) qPCR analysis of whole lung tissue from WT and Gata6 CKO mice treated with DMF or vehicle to detect expression of indicated BMP receptors. Data are means ± SE, n = 6–12, *p < 0.05. **p < 0.01 by Kruskal–Wallis test followed by Dunn’s multiple comparisons test ( BmpR2, ActRIIB, Alk1 ) and one-way ANOVA followed by post hoc Tukey’s multiple comparison ( Endoglin ). ( B ) qPCR analysis of whole lung tissue from WT and Gata6 CKO mice treated with DMF or vehicle to detect expression of the antioxidant enzymes and eNOS . Data are means ± SE, n = 6–17, *p < 0.05., **p < 0.01, ***p < 0.001 by one-way ANOVA followed by post hoc Tukey’s multiple comparisons test ( SOD2, GPX1, CAT ) and Kruskal–Wallis test with post hoc Dunn’s multiple comparisons test ( eNOS ). ( C,D ) mRNA levels of indicated BMP receptors and antioxidant enzymes measured by qPCR in PAEC from WT and Gata6 CKO mice treated with DMF or vehicle. Data are means ± SE, n = 3–6 mice/group, *p < 0.05, **p < 0.01 by Kruskal–Wallis test with post hoc Dunn’s multiple comparisons test. ( E–G ) RVSP, pulmonary acceleration time as a fraction of ejection time (PAT/ET) and Fulton index (RV/[LV + S]) were evaluated in WT and Gata6 CKO mice in the presence or absence of DMF. Data are means ± SE. n = 5–11 mice/group. *p < 0.05, **p < 0.01. ***p < 0.001 by Kruskal–Wallis test with post hoc Dunn’s multiple comparisons test (RVSP) and one-way ANOVA followed by post hoc Tukey’s multiple comparisons test (PAT/ET and RV/(LV + S).

Article Snippet: Antibodies were as follows: Goat anti-human GATA6 (R&D; AF1700, 1:500), mouse anti-human GATA6 (Santa-Cruz, 517554, 1:500), Rabbit anti-human/mouse SOD2 (Cell signaling technology; 13141, 1:1000; 13145, 1:5000), Goat anti-human ALK1 (R&D; AF370, 1:500), Rabbit anti-mouse ALK1 (ABGENT AP7807a, 1:1000), rabbit anti-human/mouse ActR2B (LS bio; LS-B7781, 1:1000), mouse anti-human BMPR2 (Novus; NBP2-37624 Clone 3F6; 1:1000), rabbit anti-mouse BMPR2 (Proteintech; 19087–1-AP; 1:1000), mouse anti-mouse/human/rat BMPR2 (Abcam, catalog #130206 1:500), rabbit anti-human/mouse Endoglin (Proteintech; 10862–1-AP; 1:1000), rabbit anti-mouse BiP (Abcam; ab53068; 1:1000), rabbit anti-mouse CHOP (Novus; NBP2-13172; 1:1000), mouse anti-beta-actin (Sigma; A1978; 1:2000), mouse anti-human PRPF4 (Abcam, 69878, 1:1000), rabbit anti-human MYEOV (Invitrogen, PA5-48938 1:1000), rabbit anti-human STING (Invitrogen, PA5-26751, 1:1000), rabbit anti-human/mouse/rat EAF1 (Invitrogen, PA5-100493, 1:1000), rabbit anti-human/mouse/rat Histone H3 (Cell Signaling, 9715 1:1000), rabbit anti-human/mouse/rat a-b Tubulin (Cell Signaling, 2148, 1:1000).

Techniques: Expressing, Injection

Graphical representation of the role of GATA6 in coordinating cross-talk between BMP10 and oxidative stress axis in PAH. GATA6 is an activator of anti-oxidant enzymes and its deficiency in PAEC and PASMC induces oxidative stress and mitochondrial dysfunction. BMP10 induces expression of GATA6 through the ALK1, BMPRII, ENG and ERK pathway. GATA6, in turn, transcriptionally activates BMP receptors in PAEC. Endothelial GATA6 regulates PASMC function via paracrine factors. TGFβ2 secreted by GATA6 deficient PAEC induces PASMC proliferation. Administration of dimethyl fumarate (DMF) to mice with endothelial Gata6 loss restores expression of BMP receptors, resolves oxidative stress, and reverses PH.

Journal: Scientific Reports

Article Title: GATA6 coordinates cross-talk between BMP10 and oxidative stress axis in pulmonary arterial hypertension

doi: 10.1038/s41598-023-33779-8

Figure Lengend Snippet: Graphical representation of the role of GATA6 in coordinating cross-talk between BMP10 and oxidative stress axis in PAH. GATA6 is an activator of anti-oxidant enzymes and its deficiency in PAEC and PASMC induces oxidative stress and mitochondrial dysfunction. BMP10 induces expression of GATA6 through the ALK1, BMPRII, ENG and ERK pathway. GATA6, in turn, transcriptionally activates BMP receptors in PAEC. Endothelial GATA6 regulates PASMC function via paracrine factors. TGFβ2 secreted by GATA6 deficient PAEC induces PASMC proliferation. Administration of dimethyl fumarate (DMF) to mice with endothelial Gata6 loss restores expression of BMP receptors, resolves oxidative stress, and reverses PH.

Article Snippet: Antibodies were as follows: Goat anti-human GATA6 (R&D; AF1700, 1:500), mouse anti-human GATA6 (Santa-Cruz, 517554, 1:500), Rabbit anti-human/mouse SOD2 (Cell signaling technology; 13141, 1:1000; 13145, 1:5000), Goat anti-human ALK1 (R&D; AF370, 1:500), Rabbit anti-mouse ALK1 (ABGENT AP7807a, 1:1000), rabbit anti-human/mouse ActR2B (LS bio; LS-B7781, 1:1000), mouse anti-human BMPR2 (Novus; NBP2-37624 Clone 3F6; 1:1000), rabbit anti-mouse BMPR2 (Proteintech; 19087–1-AP; 1:1000), mouse anti-mouse/human/rat BMPR2 (Abcam, catalog #130206 1:500), rabbit anti-human/mouse Endoglin (Proteintech; 10862–1-AP; 1:1000), rabbit anti-mouse BiP (Abcam; ab53068; 1:1000), rabbit anti-mouse CHOP (Novus; NBP2-13172; 1:1000), mouse anti-beta-actin (Sigma; A1978; 1:2000), mouse anti-human PRPF4 (Abcam, 69878, 1:1000), rabbit anti-human MYEOV (Invitrogen, PA5-48938 1:1000), rabbit anti-human STING (Invitrogen, PA5-26751, 1:1000), rabbit anti-human/mouse/rat EAF1 (Invitrogen, PA5-100493, 1:1000), rabbit anti-human/mouse/rat Histone H3 (Cell Signaling, 9715 1:1000), rabbit anti-human/mouse/rat a-b Tubulin (Cell Signaling, 2148, 1:1000).

Techniques: Expressing