Characterisation and augmentation of intra-cardiac migration of iPSC-derived cardiomyocytes; Towards the optimisation of myocardial regeneration therapy
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PhD Thesis
Embargoed until: 2025-04-29
Reason: Author Request
Embargoed until: 2025-04-29
Reason: Author Request
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The transplantaRon of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) is an emerging treatment for heart failure. However, current cell delivery methods offer poor donor cell engraNment and pose risks such as arrhythmia occurrence. Our recently developed epicardial placement technology enables safer and more effecRve cell delivery to the heart. Yet, its success may depend on the migraRon of transplanted hiPSC-CMs from the epicardium to the damaged myocardium, with their subsequent integraRon contribuRng to cardiac contracRle performance. This doctoral thesis primarily aimed to characterise the intra-cardiac migraRon of hiPSC-CMs in vitro. Firstly, hiPSC culture and cardiomyogenic differenRaRon were extensively opRmised using four hiPSC lines from different donors. Generated hiPSC-CMs displayed typical characterisRcs including spontaneous contracRon and CM-specific gene and protein expression. The differenRaRon protocol published by Burridge et al. (2015) emerged as the most effecRve, yielding sufficient CMs across all lines with an average of 77-83% cTnT posiRvity of hiPSC-CMs used in further studies. Secondly, we invesRgated the intrinsic migraRon potenRal of hiPSC-CMs within a myocardial infarcRon (MI) environment. Scratch wound assays demonstrated that the migraRon speed of hiPSC-CMs almost doubled aNer exposure to LV homogenate (15.6 ± 3.6 μm/h), which was produced from a rat leN ventricular myocardial Rssue following leN coronary artery ligaRon, compared to cell medium (8.0 ± 4.0 μm/h). Furthermore, a transwell migraRon model revealed significant concentraRon-dependent chemotacRc responses of hiPSC-CMs to LV homogenate, showing up to an 8-fold increase in migraRon. We explored the underlying mechanisms driving LV homogenate-induced hiPSC-CM migraRon through invesRgaRng gene expression differences between migraRng and non-migraRng hiPSC-CMs using RNA sequencing. Obtained results revealed potenRal pathways involved in cell movement and migraRon, including TGF-β signalling. Indeed, inhibiRon of TGF-β through the use of a chemical inhibitor and anRbody neutralisaRon significantly diminished the migraRon of hiPSC-CMs towards LV homogenate. Overall, we have discovered novel informaRon on the inherent migratory capacity of hiPSC-CMs as well as insight into the intricate mechanisms governing their migraRon in an MI environment, holding promise for improving cardiac regeneraRon therapies.
Authors
Deelen, LCollections
- Theses [4201]