Characterizing intracellular lipid signaling in dilated cardiomyopathy using human iPSC-derived cardiac cells

by Nadezda Ignatyeva
Cumulative thesis
Date of Examination:2023-03-29
Date of issue:2023-04-18
Advisor:PD Dr. Antje Ebert
Referee:PD Dr. Antje Ebert
Referee:Prof. Dr. Silvio O. Rizzoli
Persistent Address: http://dx.doi.org/10.53846/goediss-9835

 

 

Files in this item

Name:PhD thesis - Nadezda Ignatyeva.pdf
Size:10.3Mb
Format:PDF

This file will be freely accessible after 2024-03-28.

Abstract

English

Sarcomere protein misalignment is a primary consequence of inherited mutations causing dilated cardiomyopathy (DCM) and results in severe molecular patho-phenotypes in cardiomyocytes from patients. In this study, human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) carrying mutations in troponin T (TnT-R173W, TnT-R141W) and tropomyosin in combination with CRISPR/Cas9 genome editing were used as a model to investigate novel disease mechanisms in DCM. We revealed attenuated interactions between the sarcomere and other organelles in DCM iPSC-CMs and resulting downstream patho-phenotypes, such as defective lipid signaling. Using super-resolution stimulated emission depletion (STED) imaging, we described molecular disorganization and defective endosome distribution in patient-derived or CRISPR/Cas9 mutation-introduced iPSC-CMs. Moreover, we applied a quantitative assessment to recapitulate vesicle distribution in DCM iPSC-CMs as well as WT controls. Treatment of DCM iPSC-CMs with a small molecule chemical, a RhoA activating peptide, or CRISPR/Cas9-based genomic mutation-correction, rescued these molecular dysfunctions. Vice versa, applying a lipid signaling modifier to WT iPSC-CMs introduced the molecular patho-phenotype of abnormal protein-protein interactions and signaling observed previously in DCM iPSC-CMs into the WT controls. We corroborated our findings using assessment of left ventricular tissues from patients with end-stage heart failure due to DCM or other conditions compared to tissues from healthy donors. Tissues of failing hearts were found to display the molecular signaling patho-phenotype including altered protein expression. This suggested defective organization of organellar microdomains and subsequent pathological lipid signaling as well as dysregulated metabolite homeostasis. This molecular disease mechanism may potentially be present also in systolic heart failure due to other reasons than DCM, although further studies are required to clarify this point. Altogether, we revealed a novel patho-mechanistic crosstalk of dysregulated protein-protein interactions and defective lipid signaling in human models of DCM. Our research may contribute to potential translational and therapeutic directions in cardiac disease.
Keywords: sarcomere protein mutations; iPSC-CMs; dilated cardiomyopathy DCM; heart failure; druggable targets; endoplasmic reticulum; endosome distribution