Investigating a microRNA-499-5p network during cardiac development

Wittig, Johannes (2019) Investigating a microRNA-499-5p network during cardiac development. Doctoral thesis, University of East Anglia.

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Cardiovascular disease is a major cause of death world-wide, which makes it important to study cardiac development. Research into the heart, the first functioning organ in the body, will provide new knowledge to understand potential causes of different heart conditions. Cardiac maturation is a complex process depending on several different signalling cascades, which are fine-tuned by microRNAs. Alterations in NOTCH1 and ETS1 signalling have been linked to congenital heart defects and diseases. NOTCH1 is vital for cardiac septation and trabeculation, whereas ETS1 is necessary for neural crest cell coordination and proper maturation of the endocardial cushions. Further, both of them are important for inducing epithelial to mesenchymal transition in different tissues. Next generation sequencing of small RNAs has identified several microRNAs (miR-126, miR-499 & miR- 451) upregulated during cushion formation (HH17-20 chicken). This was confirmed by RT-qPCR. Interestingly, one of these microRNAs (microRNA-499-5p) targets a site present in the ETS1 and NOTCH1 3’UTR and microRNA-499-5p mimics repressed expression of a luciferase reporter gene. A cardiac injection procedure was developed in the context of this project to asses differential expression and phenotypes after AntagomiR mediated microRNA-499-5p knockdown in vivo. RNAseq revealed strong modulation of extracellular matrix genes, which are connected to ETS1 and upregulation of a NOTCH signalling cascade that affected cardiac Troponin. Along with these gene expression changes we have observed increased ECM deposition at the sites of endocardial cushions by sectioning hearts and subsequent 3D-reconstruction and volume measurements. Moreover, a reduced heart rate in knockdown animals has been observed which was linked to cardiac contraction defects due to reduced TNNT2 expression which may have caused Ca2+ desensitization. TNNT2 reduction came with reduced TBX3 expression which potentially links the observed defects to the conduction system. Future experiments should strengthen the observed links and specify TNNT2s and TBX3s relation to cardiac conduction.

Item Type: Thesis (Doctoral)
Faculty \ School: Faculty of Science > School of Biological Sciences
Depositing User: Users 11011 not found.
Date Deposited: 12 Nov 2019 14:27
Last Modified: 07 May 2021 00:40


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