Tags

Type your tag names separated by a space and hit enter

The ACVR1 R206H mutation found in fibrodysplasia ossificans progressiva increases human induced pluripotent stem cell-derived endothelial cell formation and collagen production through BMP-mediated SMAD1/5/8 signaling.
Stem Cell Res Ther. 2016 08 17; 7(1):115.SC

Abstract

BACKGROUND

The Activin A and bone morphogenetic protein (BMP) pathways are critical regulators of the immune system and of bone formation. Inappropriate activation of these pathways, as in conditions of congenital heterotopic ossification, are thought to activate an osteogenic program in endothelial cells. However, if and how this occurs in human endothelial cells remains unclear.

METHODS

We used a new directed differentiation protocol to create human induced pluripotent stem cell (hiPSC)-derived endothelial cells (iECs) from patients with fibrodysplasia ossificans progressiva (FOP), a congenital disease of heterotopic ossification caused by an activating R206H mutation in the Activin A type I receptor (ACVR1). This strategy allowed the direct assay of the cell-autonomous effects of ACVR1 R206H in the endogenous locus without the use of transgenic expression. These cells were challenged with BMP or Activin A ligand, and tested for their ability to activate osteogenesis, extracellular matrix production, and differential downstream signaling in the BMP/Activin A pathways.

RESULTS

We found that FOP iECs could form in conditions with low or absent BMP4. These conditions are not normally permissive in control cells. FOP iECs cultured in mineralization media showed increased alkaline phosphatase staining, suggesting formation of immature osteoblasts, but failed to show mature osteoblastic features. However, FOP iECs expressed more fibroblastic genes and Collagen 1/2 compared to control iECs, suggesting a mechanism for the tissue fibrosis seen in early heterotopic lesions. Finally, FOP iECs showed increased SMAD1/5/8 signaling upon BMP4 stimulation. Contrary to FOP hiPSCs, FOP iECs did not show a significant increase in SMAD1/5/8 phosphorylation upon Activin A stimulation, suggesting that the ACVR1 R206H mutation has a cell type-specific effect. In addition, we found that the expression of ACVR1 and type II receptors were different in hiPSCs and iECs, which could explain the cell type-specific SMAD signaling.

CONCLUSIONS

Our results suggest that the ACVR1 R206H mutation may not directly increase the formation of mature chondrogenic or osteogenic cells by FOP iECs. Our results also show that BMP can induce endothelial cell dysfunction, increase expression of fibrogenic matrix proteins, and cause differential downstream signaling of the ACVR1 R206H mutation. This iPSC model provides new insight into how human endothelial cells may contribute to the pathogenesis of heterotopic ossification.

Authors+Show Affiliations

Institute for Human Genetics and the Division of Endocrinology and Metabolism, University of California, 513 Parnassus Avenue, HSE901G, San Francisco, CA, 94143-0794, USA.Institute for Human Genetics and the Division of Endocrinology and Metabolism, University of California, 513 Parnassus Avenue, HSE901G, San Francisco, CA, 94143-0794, USA.Institute for Human Genetics and the Division of Endocrinology and Metabolism, University of California, 513 Parnassus Avenue, HSE901G, San Francisco, CA, 94143-0794, USA.Gladstone Institute of Cardiovascular Disease, 1650 Owens Street, San Francisco, CA, 94158, USA.Gladstone Institute of Cardiovascular Disease, 1650 Owens Street, San Francisco, CA, 94158, USA.Institute for Human Genetics and the Division of Endocrinology and Metabolism, University of California, 513 Parnassus Avenue, HSE901G, San Francisco, CA, 94143-0794, USA.Institute for Human Genetics and the Division of Endocrinology and Metabolism, University of California, 513 Parnassus Avenue, HSE901G, San Francisco, CA, 94143-0794, USA.School of Dentistry, Oral and Craniofacial Sciences Program, University of California, 707 Parnassus Avenue, San Francisco, CA, 94143, USA.Gladstone Institute of Cardiovascular Disease, 1650 Owens Street, San Francisco, CA, 94158, USA.Institute for Human Genetics and the Division of Endocrinology and Metabolism, University of California, 513 Parnassus Avenue, HSE901G, San Francisco, CA, 94143-0794, USA. Edward.Hsiao@ucsf.edu. Department of Endocrinology, Diabetes, and Metabolism, Institute for Human Genetics, University of California, 513 Parnassus Avenue, HSE901G, UCSF Box 0794, San Francisco, CA, 94143-0794, USA. Edward.Hsiao@ucsf.edu.

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

27530160

Citation

Barruet, Emilie, et al. "The ACVR1 R206H Mutation Found in Fibrodysplasia Ossificans Progressiva Increases Human Induced Pluripotent Stem Cell-derived Endothelial Cell Formation and Collagen Production Through BMP-mediated SMAD1/5/8 Signaling." Stem Cell Research & Therapy, vol. 7, no. 1, 2016, p. 115.
Barruet E, Morales BM, Lwin W, et al. The ACVR1 R206H mutation found in fibrodysplasia ossificans progressiva increases human induced pluripotent stem cell-derived endothelial cell formation and collagen production through BMP-mediated SMAD1/5/8 signaling. Stem Cell Res Ther. 2016;7(1):115.
Barruet, E., Morales, B. M., Lwin, W., White, M. P., Theodoris, C. V., Kim, H., Urrutia, A., Wong, S. A., Srivastava, D., & Hsiao, E. C. (2016). The ACVR1 R206H mutation found in fibrodysplasia ossificans progressiva increases human induced pluripotent stem cell-derived endothelial cell formation and collagen production through BMP-mediated SMAD1/5/8 signaling. Stem Cell Research & Therapy, 7(1), 115. https://doi.org/10.1186/s13287-016-0372-6
Barruet E, et al. The ACVR1 R206H Mutation Found in Fibrodysplasia Ossificans Progressiva Increases Human Induced Pluripotent Stem Cell-derived Endothelial Cell Formation and Collagen Production Through BMP-mediated SMAD1/5/8 Signaling. Stem Cell Res Ther. 2016 08 17;7(1):115. PubMed PMID: 27530160.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - The ACVR1 R206H mutation found in fibrodysplasia ossificans progressiva increases human induced pluripotent stem cell-derived endothelial cell formation and collagen production through BMP-mediated SMAD1/5/8 signaling. AU - Barruet,Emilie, AU - Morales,Blanca M, AU - Lwin,Wint, AU - White,Mark P, AU - Theodoris,Christina V, AU - Kim,Hannah, AU - Urrutia,Ashley, AU - Wong,Sarah Anne, AU - Srivastava,Deepak, AU - Hsiao,Edward C, Y1 - 2016/08/17/ PY - 2016/03/22/received PY - 2016/07/21/accepted PY - 2016/8/18/entrez PY - 2016/8/18/pubmed PY - 2017/11/29/medline KW - ACVR1 KW - Activin A signaling KW - BMP KW - Fibrodysplasia ossificans progressiva KW - Tissue fibrosis KW - hiPS-derived endothelial cells SP - 115 EP - 115 JF - Stem cell research & therapy JO - Stem Cell Res Ther VL - 7 IS - 1 N2 - BACKGROUND: The Activin A and bone morphogenetic protein (BMP) pathways are critical regulators of the immune system and of bone formation. Inappropriate activation of these pathways, as in conditions of congenital heterotopic ossification, are thought to activate an osteogenic program in endothelial cells. However, if and how this occurs in human endothelial cells remains unclear. METHODS: We used a new directed differentiation protocol to create human induced pluripotent stem cell (hiPSC)-derived endothelial cells (iECs) from patients with fibrodysplasia ossificans progressiva (FOP), a congenital disease of heterotopic ossification caused by an activating R206H mutation in the Activin A type I receptor (ACVR1). This strategy allowed the direct assay of the cell-autonomous effects of ACVR1 R206H in the endogenous locus without the use of transgenic expression. These cells were challenged with BMP or Activin A ligand, and tested for their ability to activate osteogenesis, extracellular matrix production, and differential downstream signaling in the BMP/Activin A pathways. RESULTS: We found that FOP iECs could form in conditions with low or absent BMP4. These conditions are not normally permissive in control cells. FOP iECs cultured in mineralization media showed increased alkaline phosphatase staining, suggesting formation of immature osteoblasts, but failed to show mature osteoblastic features. However, FOP iECs expressed more fibroblastic genes and Collagen 1/2 compared to control iECs, suggesting a mechanism for the tissue fibrosis seen in early heterotopic lesions. Finally, FOP iECs showed increased SMAD1/5/8 signaling upon BMP4 stimulation. Contrary to FOP hiPSCs, FOP iECs did not show a significant increase in SMAD1/5/8 phosphorylation upon Activin A stimulation, suggesting that the ACVR1 R206H mutation has a cell type-specific effect. In addition, we found that the expression of ACVR1 and type II receptors were different in hiPSCs and iECs, which could explain the cell type-specific SMAD signaling. CONCLUSIONS: Our results suggest that the ACVR1 R206H mutation may not directly increase the formation of mature chondrogenic or osteogenic cells by FOP iECs. Our results also show that BMP can induce endothelial cell dysfunction, increase expression of fibrogenic matrix proteins, and cause differential downstream signaling of the ACVR1 R206H mutation. This iPSC model provides new insight into how human endothelial cells may contribute to the pathogenesis of heterotopic ossification. SN - 1757-6512 UR - https://www.unboundmedicine.com/medline/citation/27530160/The_ACVR1_R206H_mutation_found_in_fibrodysplasia_ossificans_progressiva_increases_human_induced_pluripotent_stem_cell_derived_endothelial_cell_formation_and_collagen_production_through_BMP_mediated_SMAD1/5/8_signaling_ L2 - https://stemcellres.biomedcentral.com/articles/10.1186/s13287-016-0372-6 DB - PRIME DP - Unbound Medicine ER -