University of Groningen Endothelial plasticity in fibrosis and epigenetics as a therapeutic target Hulshoff, Melanie

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University of Groningen

Endothelial plasticity in fibrosis and epigenetics as a therapeutic target

Hulshoff, Melanie

DOI:

10.33612/diss.146265795

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Publication date: 2020

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Citation for published version (APA):

Hulshoff, M. (2020). Endothelial plasticity in fibrosis and epigenetics as a therapeutic target. University of Groningen. https://doi.org/10.33612/diss.146265795

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546222-L-bw-Hulshoff 546222-L-bw-Hulshoff 546222-L-bw-Hulshoff 546222-L-bw-Hulshoff Processed on: 27-10-2020 Processed on: 27-10-2020 Processed on: 27-10-2020

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RESEARCH SUMMARY

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(EPI)GENETIC REGULATION OF ENDOTHELIAL PLASTICITY

Endothelial-to-mesenchymal transition (EndMT) is the hallmark of endothelial plasticity and is the main focus of the first part of this thesis. EndMT can be induced via several signaling pathways, including TGF-β, and modulators, including hypoxia (lack of oxygen) and high glucose concentrations. These signaling pathways and modulators play a crucial role in the activation of epigenetic modifiers which in part regulate EndMT. In Chapter 2 we summarize epigenetic changes that are associated with EndMT in the context of chronic heart disease. We identified histone methylation and (de)acetylation, DNA methylation as well as noncoding RNAs as important regulators and thus potential epigenetic targets of EndMT. An example of this is demonstrated in Chapter 3 where hypoxia induces TGF-β signaling which in turn causes DNMT3a-mediated promoter methylation of Rasal1, thereby contributing to EndMT.

The primary cause of morbidity and mortality in diabetes patients is cardiovascular disease. Diabetes patients exhibit accelerated aortic stiffening which precedes hypertension development and is therefore believed to be an early contributor to cardiovascular disease. In Chapter 4 we identified robust EndMT in aortas of type 2 diabetic mice and patients which might contribute to aortic stiffening in type 2 diabetes. We also identified miR-132-3p as well as KLF7 as potential novel regulators in this context. This shows that EndMT does not only play a role in the context of cardiac fibrosis, but is also associated with other pathologies.

Therefore, we continued by summarizing the non-coding RNAs which are known to regulate EndMT in different contexts such as atherosclerosis, myocardial fibrosis but also of kidney disease in Chapter 5. We also highlight non-coding RNAs which have different effects on EndMT in development and pathology. In all, we could show that a remarkable number of non-coding RNAs is associated to the EndMT regulatory program.

In Chapter 6 we performed stage-specific mapping of EndMT to get insights in the genetic regulatory mechanisms that facilitate the active stage of EndMT. We identified that the active stage of EndMT is characterized by changes in the Rap1 signaling pathway, and that Rap1 inhibition in part alleviates EndMT.

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Altogether, the first part of this thesis highlights the interplay between the different signaling pathways, modulators as well as epigenetic modifiers in regulating EndMT.

(EPI)GENETICS AS A THERAPEUTIC TARGET IN FIBROSIS

In the second part of this thesis, we focus on (epi)genetics as a therapeutic target in fibrosis. In Chapter 7 we show that Serelaxin, a recombinant form of the human hormone Relaxin-2, inhibits EndMT and ameliorates cardiac fibrosis. Serelaxin reversed the TGF-β-mediated histone modifications at the RXFP1 promoter thereby restoring the expression of RXFP1 which enabled RXFP1-mediated inhibition of EndMT.

Cardiovascular disease and heart failure are the primary cause of morbidity and mortality in patients with chronic kidney disease. In Chapter 8 we review the current knowledge of how kidney injury causes heart failure and fibrosis. Different components such as inflammation but also EndMT and non-coding RNAs are potential regulators which facilitate chronic kidney disease-induced cardiac fibrosis.

In Chapter 9 we summarize genomic editing tools including the recent discovered CRISPR/Cas9 system. We describe distinct functions of CRISPR/Cas9 derivatives including tracking, transcriptional activation and repression, and base editing. We also discuss the in vivo applications and potential of CRISPR/Cas9 derivatives which results in changes in (disease) phenotype. In Chapter 10 we make use of the CRISPR/Cas system by deactivating the Cas9 nuclease and fusing it to the catalytic domain of TET3. This enabled gene-specific hydroxymethylation and thus reactivation which ameliorates kidney fibrosis. In Chapter 11 we use split-intein mediated AAV delivery of the established system (deactivated Cas9 nuclease fused to the catalytic domain of TET3) to perform gene-specific reactivation of Rasal1 expression which ameliorates cardiac fibrosis. This shows the possibilities of targeting epigenetic modifications thereby ameliorating organ fibrosis.

In conclusion, in this thesis we have shown that endothelial plasticity is highly regulated by epigenetic modifications. Besides, we have shown that epigenetic

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RESEARCH SUMMARY

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modifications can be targeted in vivo which results in the amelioration of organ

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