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Generation of GLA-Knockout Human Embryonic Stem Cell Lines to Model Autophagic Dysfunction and Exosome Secretion in Fabry Disease-Associated Hypertrophic Cardiomyopathy

1
Institute of Pharmacology, National Yang-Ming University, Taipei 11221, Taiwan
2
Department of Medical Research, Taipei Veterans General Hospital, Taipei 11217, Taiwan
3
School of Medicine, National Yang-Ming University, Taipei 11221, Taiwan
4
School of Pharmaceutical Sciences, National Yang-Ming University, Taipei 11221, Taiwan
5
Department of Emergent Medicine, Taipei Veterans General Hospital, Taipei 11217, Taiwan
6
Division of Cardiology & Department of Medicine, Taipei Veterans General Hospital, Taipei 11217, Taiwan
7
Department of Pharmacy, Taipei Veterans General Hospital, Taipei 11217, Taiwan
8
Genomics Research Center, Academia Sinica, Taipei 11574, Taiwan
*
Authors to whom correspondence should be addressed.
Cells 2019, 8(4), 327; https://doi.org/10.3390/cells8040327
Received: 25 February 2019 / Revised: 1 April 2019 / Accepted: 4 April 2019 / Published: 8 April 2019
(This article belongs to the Special Issue Stem Cell Research on Cardiology)
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PDF [3163 KB, uploaded 17 April 2019]
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Abstract

Fabry disease (FD) is a rare inherited disorder characterized by a wide range of systemic symptoms; it is particularly associated with cardiovascular and renal problems. Enzyme replacement therapy and pharmacological chaperone migalastat are the only approved and effective treatment strategies for FD patients. It is well documented that alpha-galactosidase A (GLA) enzyme activity deficiency causes globotriaosylceramide (Gb3) accumulation, which plays a crucial role in the etiology of FD. However, the detailed mechanisms remain unclear, and the lack of a reliable and powerful disease model is an obstacle. In this study, we created such a model by using CRISPR/Cas9-mediated editing of GLA gene to knockout its expression in human embryonic stem cells (hESCs). The cardiomyocytes differentiated from these hESCs (GLA-null CMs) were characterized by the accumulation of Gb3 and significant increases of cell surface area, the landmarks of FD-associated cardiomyopathy. Furthermore, we used mass spectrometry to compare the proteomes of GLA-null CMs and parental wild type CMs and found that the Rab GTPases involved in exocytotic vesicle release were significantly downregulated. This caused impairment of autophagic flux and protein turnover, resulting in an increase of reactive oxygen species and apoptosis. To summarize, we established a FD model which can be used as a promising tool to study human hypertrophic cardiomyopathy in a physiologically and pathologically relevant manner and to develop new therapies by targeting Rab GTPases signaling-related exosomal vesicles transportation. View Full-Text
Keywords: Fabry disease; human embryonic stem cells; CRISPR/Cas9 genomic editing; Mass spectrometry proteomic analysis; hypertrophic cardiomyopathy; disease model Fabry disease; human embryonic stem cells; CRISPR/Cas9 genomic editing; Mass spectrometry proteomic analysis; hypertrophic cardiomyopathy; disease model
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited (CC BY 4.0).

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Song, H.-Y.; Chien, C.-S.; Yarmishyn, A.A.; Chou, S.-J.; Yang, Y.-P.; Wang, M.-L.; Wang, C.-Y.; Leu, H.-B.; Yu, W.-C.; Chang, Y.-L.; Chiou, S.-H. Generation of GLA-Knockout Human Embryonic Stem Cell Lines to Model Autophagic Dysfunction and Exosome Secretion in Fabry Disease-Associated Hypertrophic Cardiomyopathy. Cells 2019, 8, 327.

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