Vici syndrome is a rare, autosomal recessive disorder with severe dysfunction of multiple organ systems. Mutations in the EPG5 gene encoding ectopic P-granule autophagy protein 5, which lead to defects in the autophagic flux, are found as the genetic cause of Vici syndrome. In around 80% of cases, development of dilated and hypertrophic forms of cardiomyopathies is observed. However, the mechanisms of cardiomyopathy development and possible therapeutic targets remain to be elucidated. Interestingly, previous work in Caenorhabditis elegans showed that targeting the enzyme N-acetylglucosamine transferase (OGT) in EPG5-deficient nematodes restored fully functional autophagy. However, the effect of targeting OGT in a human Vici disease model is unknown. In order to understand more about the mechanisms of Vici cardiomyopathies and the effect of targeting OGT in Vici syndrome, we established a Vici patient-specific in vitro disease model by generating induced pluripotent stem cell-derived cardiomyocytes (iVici iPSC-CMs) from fibroblasts obtained from a Vici patient. The Vici patient carried a homologous EPG5 mutation (c.4952+1G>A), which results in the aberrant splicing of the EPG5 transcript and the removal of exon 28 and introduces a frameshift and a premature stop codon at exon 29. In the iVici iPSC-CMs model, we observed some of the known Vici-associated cardiomyopathy phenotypes including cellular hypertrophy and sarcomere disarray. The aims of this study were to investigate physiological and functional properties of iVici iPSC-CMs and to provide an understanding of the molecular mechanisms driving cardiomyopathy development in Vici patients by investigating the presence of potential molecular defects in iVici iPSC-CMs. In this study we also tested the effect of pharmacological OGT inhibitors in iVici iPSC-CMs. We showed that iVici iPSC-CMs exhibited defects in the autophagosome and lysosome fusion and displayed slower beating frequency and altered beating kinetics possibly resulting from the alterations in calcium handling in iVici iPSC-CMs. We observed the accumulation of fragmented mitochondria and increase in oxidative stress and protein O-GlcNAcylation in iVici iPSC-CMs, which are possible causes of the Vici Syndrome-associated cardiomyopathies. Moreover, we found that the non-specific OGT inhibitor alloxan and the specific OGT inhibitor ST045849 could enhance the fusion between the autophagosome and the lysosome in iVici iPSC-CMs. Taken together, in the current work, we demonstrated that iVici iPSC-CMs model can recapitulate some of the disease phenotypes of Vici syndrome, and at the same time, provided further insight into the underlying mechanisms of Vici cardiomyopathy and possible pharmacological therapeutics. Future studies should focus on further investigation of the feasibility of using OGT inhibition as a therapeutic approach and of the underlying mechanisms to identify new therapeutic targets using iVici iPSC-CMs.
| Identifer | oai:union.ndltd.org:DRESDEN/oai:qucosa:de:qucosa:72380 |
| Date | 07 October 2020 |
| Creators | Alfar, Ezzaldin Ahmed |
| Contributors | Guan, Kaomei, Antos, Christopher, Technische Universität Dresden |
| Source Sets | Hochschulschriftenserver (HSSS) der SLUB Dresden |
| Language | English |
| Detected Language | English |
| Type | info:eu-repo/semantics/publishedVersion, doc-type:doctoralThesis, info:eu-repo/semantics/doctoralThesis, doc-type:Text |
| Rights | info:eu-repo/semantics/openAccess |
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