Background Supravalvular aortic stenosis (SVAS) is caused by mutations in the elastin (ELN) gene and is characterized by abnormal proliferation of vascular smooth muscle cells (SMCs) that can lead to narrowing or blockage of the ascending aorta and other arterial vessels. Availability of patient-specific SMCs may facilitate studying disease mechanisms and developing novel therapeutic interventions. Methods and Results Here, we report the development of a human induced pluripotent stem cell (iPSC) line from a patient with SVAS caused by the premature termination in exon 10 of the ELN gene due to an exon 9 4-nucleotide insertion. We showed that SVAS iPSC-derived SMCs (iPSC-SMCs) had significantly fewer organized networks of smooth muscle alpha actin (SM α-actin) filament bundles, a hallmark of mature contractile SMCs, compared to control iPSC-SMCs. Addition of elastin recombinant protein or enhancement of small GTPase RhoA signaling was able to rescue the formation of SM α-actin filament bundles in SVAS iPSC-SMCs. Cell counts and BrdU analysis revealed a significantly higher proliferation rate in SVAS iPSC-SMCs than control iPSC-SMCs. Furthermore, SVAS iPSC-SMCs migrated at a markedly higher rate to the chemotactic agent platelet-derived growth factor (PDGF) in comparison with the control iPSC-SMCs. We also provided evidence that elevated activity of extracellular signal-regulated kinase 1/2 (ERK1/2) is required for hyper-proliferation of SVAS iPSC-SMCs. The phenotype was confirmed in iPSC-SMCs generated from a patient with deletion of elastin due to Williams-Beuren syndrome (WBS). Conclusions Thus, SVAS iPSC-SMCs recapitulate key pathological features of patients with SVAS and may provide a promising strategy to study disease mechanisms and to develop novel therapies.
SummaryThere is an urgent need for an efficient approach to obtain a large-scale and renewable source of functional human vascular smooth muscle cells (VSMCs) to establish robust, patient-specific tissue model systems for studying the pathogenesis of vascular disease, and for developing novel therapeutic interventions. Here, we have derived a large quantity of highly enriched functional VSMCs from human induced pluripotent stem cells (hiPSC-VSMCs). Furthermore, we have engineered 3D tissue rings from hiPSC-VSMCs using a facile one-step cellular self-assembly approach. The tissue rings are mechanically robust and can be used for vascular tissue engineering and disease modeling of supravalvular aortic stenosis syndrome. Our method may serve as a model system, extendable to study other vascular proliferative diseases for drug screening. Thus, this report describes an exciting platform technology with broad utility for manufacturing cell-based tissues and materials for various biomedical applications.
We have developed an engineered heart tissue (EHT) system that uses laser-cut sheets of decellularized myocardium as scaffolds. This material enables formation of thin muscle strips whose biomechanical characteristics are easily measured and manipulated. To create EHTs, sections of porcine myocardium were laser-cut into ribbon-like shapes, decellularized, and mounted in specialized clips for seeding and culture. Scaffolds were first tested by seeding with neonatal rat ventricular myocytes. EHTs beat synchronously by day five and exhibited robust length-dependent activation by day 21. Fiber orientation within the scaffold affected peak twitch stress, demonstrating its ability to guide cells toward physiologic contractile anisotropy. Scaffold anisotropy also made it possible to probe cellular responses to stretch as a function of fiber angle. Stretch that was aligned with the fiber direction increased expression of brain natriuretic peptide, but off-axis stretches (causing fiber shear) did not. The method also produced robust EHTs from cardiomyocytes derived from human embryonic stem cells and induced pluripotent stem cells (hiPSC). hiPSC-EHTs achieved maximum peak stress of 6.5 mN/mm2 and twitch kinetics approaching reported values from adult human trabeculae. We conclude that laser-cut EHTs are a viable platform for novel mechanotransduction experiments and characterizing the biomechanical function of patient-derived cardiomyoctyes.
TH is expressed in a dynamic pattern during the primitive heart tube formation. TH induces cardiac differentiation in vivo and it is a key regulator of the heart patterning, conferring atriogenic identity.
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