Periodic Reporting for period 1 - ACCURACY (Human cardiac microtissues for studying lamin cardiomyopathy)
Période du rapport: 2023-06-01 au 2025-05-31
Heart disease places a significant burden on multiple aspects of society—primarily by shortening the lifespan of those affected, increasing healthcare costs, and reducing the quality of life for both patients and their caregivers. Familial dilated cardiomyopathy is a particular form of heart disease that is genetically driven.The ACCURACY project was focused on the study of pathological variants of the LMNA gene, which encodes for the Lamin AC protein, involved in maintaining the integrity of the nucleus. Within this project, a human model that mimics dilated cardiomyopathy traits was generated in the laboratory starting from pluripotent stem cells. It was possible to generate different cell types of the heart, including cardiomyocytes (the cells that develop the force of contraction), cells of the vessels, and the supporting cells. The study highlighted that all of the different heart cells were affected by the disease. By aggregating the distinct cardiac cell types, a three-dimensional model of cardiomyopathy was generated. Here, the pathological features previously observed in cardiomyocytes alone were enhanced by their culture in a three-dimensional environment. These results were important for better understanding the disease mechanisms and the link between the genetic defect and the abnormal cardiac function. By providing a novel model and shedding light into the cellular and molecular mechanisms of dilated cardiomyopathy caused by mutations in Lamin AC, the ACCURACY project can contribute to defining new ideas for developing new treatments. Ultimately this could result in better disease management for patients and their families.
With the support of collaborators (Monzino hospital, Milan, Italy), we obtained the biopsy material of a patient affected by lamin cardiomyopathy and created stem cells which can be directed to develop into heart-specific cells. We also used a control line from a healthy donor. To generate the cardiac microtissues we first created individual cellular types that are most frequent in the heart, namely cardiomyocytes, fibroblasts and endothelial cells. First, we found that there was a strong reduction in Lamin AC in the nucleus of the patient cells, compared to the healthy control cells. We also noticed that the nuclei of different heart cell types were less round than in control conditions, suggesting that structural integrity is compromised in diseased conditions. In addition, the DNA, that is normally tightly packed around the nuclear borders helped by its anchoring to the Lamin AC protein, was instead in less compact state in the diseased cardiac cells. This effect is particularly evident in cardiomyocytes, which are the contracting cells in the heart and therefore subjected to mechanical stress. We also examined calcium cycling within the cell and nucleus, as calcium plays a central role in contraction. Our analysis revealed changes in calcium dynamics that suggest contractility is impaired in the mutated cells. To recreate the native environment of the heart, that is a three-dimensional (3D) organ composed of many different cell types, we built microtissues with the three cardiac cell types and saw that the traits previously observed in cardiomyocytes alone hold true also in cardiomyocytes within the 3D environment and are even augmented. The work done so far has laid the foundations to perform additional assessments related to the lamin mutations phenotype. To explore further the link between the changes in the DNA regions that are linked to the Lamin AC protein we will analyze the cardiac microtissues at the single cell level. Altogether, this work has created the lay ground to generate a disease platform that can be used not only for better understanding cardiac laminopathies, but also to identify novel therapeutic approaches.
We have generated the cellular building blocks for the assembly of an advanced microtissue model of the heart to study cardiac laminopathy. This multicellular model is able to recapitulate the disease traits seen in patients carrying mutations in the LMNA gene. Moreover, this model can be applied beyond the current state of the art in multiscale settings for identifying disease phenotypes and for drug testing, in collaboration with biotech and pharmaceutical companies.