title: Modeling and rescue of the vascular phenotype of Williams-Beuren syndrome in patient induced pluripotent stem cells.
abstract: Elastin haploinsufficiency in Williams-Beuren syndrome (WBS) leads to increased vascular smooth muscle cell (SMC) proliferation and stenoses. Our objective was to generate a human induced pluripotent stem (hiPS) cell model for in vitro assessment of the WBS phenotype and to test the ability of candidate agents to rescue the phenotype. hiPS cells were reprogrammed from skin fibroblasts of a WBS patient with aortic and pulmonary stenosis and healthy control BJ fibroblasts using four-factor retrovirus reprogramming and were differentiated into SMCs. Differentiated SMCs were treated with synthetic elastin-binding protein ligand 2 (EBPL2) (20 μg/ml) or the antiproliferative drug rapamycin (100 nM) for 5 days. We generated four WBS induced pluripotent stem (iPS) cell lines that expressed pluripotency genes and differentiated into all three germ layers. Directed differentiation of BJ iPS cells yielded an 85%-92% pure SMC population that expressed differentiated SMC markers, were functionally contractile, and formed tube-like structures on three-dimensional gel assay. Unlike BJ iPS cells, WBS iPS cells generated immature SMCs that were highly proliferative, showed lower expression of differentiated SMC markers, reduced response to the vasoactive agonists, carbachol and endothelin-1, impaired vascular tube formation, and reduced calcium flux. EBPL2 partially rescued and rapamycin fully rescued the abnormal SMC phenotype by decreasing the smooth muscle proliferation rate and enhancing differentiation and tube formation. WBS iPS cell-derived SMCs demonstrate an immature proliferative phenotype with reduced functional and contractile properties, thereby recapitulating the human disease phenotype. The ability of rapamycin to rescue the phenotype provides an attractive therapeutic candidate for patients with WBS and vascular stenoses.
authors.affiliation: Department of Pediatrics, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Caroline
authors.initials: C
authors.lastname: Kinnear
authors.forename: Wing Y
authors.initials: WY
authors.lastname: Chang
authors.forename: Shahryar
authors.initials: S
authors.lastname: Khattak
authors.forename: Aleksander
authors.initials: A
authors.lastname: Hinek
authors.forename: Tadeo
authors.initials: T
authors.lastname: Thompson
authors.forename: Deivid
authors.initials: D
authors.lastname: de Carvalho Rodrigues
authors.forename: Karen
authors.initials: K
authors.lastname: Kennedy
authors.forename: Naila
authors.initials: N
authors.lastname: Mahmut
authors.forename: Peter
authors.initials: P
authors.lastname: Pasceri
authors.forename: William L
authors.initials: WL
authors.lastname: Stanford
authors.forename: James
authors.initials: J
authors.lastname: Ellis
authors.forename: Seema
authors.initials: S
authors.lastname: Mital
mesh_terms: D000943:Antigens, Differentiation / Q000235:genetics / Q000378:metabolism; D020013:Calcium Signaling; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D005347:Fibroblasts / Q000378:metabolism / Q000502:physiology; D057898:Hemizygote; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000378:metabolism / Q000502:physiology*; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000096:biosynthesis; D008297:Male; D009119:Muscle Contraction; D009131:Muscle, Smooth, Vascular / Q000473:pathology*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D050814:Octamer Transcription Factor-3 / Q000096:biosynthesis; D010446:Peptide Fragments / Q000494:pharmacology; D010641:Phenotype; D016271:Proto-Oncogene Proteins c-myc / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis; D055748:SOXB1 Transcription Factors / Q000096:biosynthesis; D017422:Sequence Analysis, DNA; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors; D059467:Transcriptome / Q000187:drug effects; D018980:Williams Syndrome / Q000235:genetics / Q000473:pathology*
chemical_list: D000943:Antigens, Differentiation; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; C489427:MYC protein, human; D050814:Octamer Transcription Factor-3; C494173:POU5F1 protein, human; D010446:Peptide Fragments; D016271:Proto-Oncogene Proteins c-myc; D011994:Recombinant Proteins; C527782:SOX2 protein, human; D055748:SOXB1 Transcription Factors; C546842:MTOR protein, human; D058570:TOR Serine-Threonine Kinases; D020123:Sirolimus