Cardiomyocyte-specific Activation of the Insulin-like Growth Factor Binding Protein 5 (IGFBP5) in the Postnatal Heart Via CRISPR/Cas9

by Janek Fischer
Doctoral thesis
Date of Examination:2025-06-25
Date of issue:2025-06-12
Advisor:Prof. Dr. Laura Zelarayán-Behrend
Referee:Prof. Dr. Elisabeth Zeisberg
Referee:Prof. Dr. Margarete Schön
Persistent Address: http://dx.doi.org/10.53846/goediss-11318

 

 

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Abstract

English

IGFBP5 is one of six proteins that bind insulin-like growth factors (IGFs) and is known to regulate processes such as cell growth, aging, and metabolism. Previous studies have shown that its transcription is elevated in various cardiovascular diseases, including heart failure, but its exact role in the heart remains unclear. A CRISPR activation system was used to investigate the mechanistic role of endogenous Igfbp5 activation in the myocardium. This system consists of an inactivated dCas9 variant fused to transcriptional activators (VP64, p65 and RTA) and gene-targeting gRNAs. For in vivo application, a transgenic mouse line expressing the dCas9VPR construct exclusively in cardiomyocytes was used. In combination with the systemic administration of previously validated gRNAs, the construct can hybridize to form a cardiomyocyte-specific, programmable transcription factor for the endogenous activation of Igfbp5 and its subsequent secretion. Heart failure was induced by ligation of the ascending aorta, resulting in hypertrophy and reduced cardiac output. Further evaluation included functional parameters and transcriptomic changes resulting from endogenous Igfbp5 activation. Observations of standard echocardiographic parameters, such as fractional area shortening, did not show significant changes upon Igfbp5 activation in the healthy heart. Moreover, Igfbp5 activation did not improve heart function in heart failure-induced mice, nor was there a substantial improvement in overall survival. Activation of Igfbp5 in the adult heart revealed strongly activated pathways involved in vascular formation. These results suggest an undefined paracrine mechanism of action of IGFBP5 in the myocardium, potentially linked to vascular biology-related pathways.
Keywords: Heart failure; Endogenous gene activation; IGFBP5; CRISPRa