1. Faculty of Health Sciences, University of Macau, Taipa, Macau, China;
2. Department of Genetics and Genome Sciences, University of Connecticut Health Center, Farmington, Connecticut, USA;
3. Agricultural Genomes Institute at Shenzhen, Chinese Academy of Agricultural Sciences, Shenzhen, China;
4. Guangdong Province Key Laboratory of Pharmacodynamic Constituents of TCM and New Drugs Research, College of Pharmacy, Jinan University, Guangzhou, Guangdong, China;
5. School of Ophthalmology and Optometry, Eye Hospital, Wenzhou Medical University, State Key Laboratory Cultivation Base and Key Laboratory of Vision Science, Ministry of Health and Zhejiang Provincial Key Laboratory of Ophthalmology and Optometry, Wenzhou, Zhejiang, China;
6. Yunnan Key Laboratory of Primate Biomedical Research, Institute of Primate Translational Medicine, Kunming University of Science and Technology, Kunming, Yunnan, China;
7. Stowers Institute for Medical Research, Kansas City, Missouri, USA;
8. Department of Cell Biology, University of Connecticut Health Center, Farmington, Connecticut, USA.
* The first two authors contributed equally to this work.
Marfan syndrome (MFS) is a connective tissue disorder caused by mutations in FBN1 gene, which encodes a key extracellular matrix protein FIBRILLIN-1. The haplosufficiency of FBN1 has been implicated in pathogenesis of MFS with manifestations primarily in cardiovascular, muscular, and ocular tissues. Due to limitations in animal models to study the late-onset diseases, human pluripotent stem cells (PSCs) offer a homogeneic tool for dissection of cellular and molecular pathogenic mechanism for MFS in vitro. Here, we first derived induced PSCs (iPSCs) from a MFS patient with a FBN1 mutation and corrected the mutation, thereby generating an isogenic “gain-of-function” control cells for the parental MFS iPSCs. Reversely, we knocked out FBN1 in both alleles in a wild-type (WT) human embryonic stem cell (ESC) line, which served as a loss-of-function model for MFS with the WT cells as an isogenic control. Mesenchymal stem cells derived from both FBN1-mutant iPSCs and -ESCs demonstrated reduced osteogenic differentiation and microfibril formation. We further demonstrated that vascular smooth muscle cells derived from FBN1-mutant iPSCs showed less sensitivity to carbachol as demonstrated by contractility and Ca2+ influx assay, compared to the isogenic controls cells. These findings were further supported by transcriptomic anaylsis of the cells. Therefore, this study based on both gain- and loss-of-function approaches confirmed the pathogenetic role of FBN1 mutations in these MFS-related phenotypic changes.
Keywords: Marfan syndrome, human pluripotent stem cells, disease modeling, genome editing, osteogenesis, smooth muscle.