Abstract 9109: Patient-Specific Ipsc-Derived Cardiac Fibroblasts Phenotypes in Lmna Cardiomyopathy

Abstract

Introduction: Mutations in LMNA (encoding lamin A/C) are an important cause of familial Dilated Cardiomyopathy (DCM), characterized by systolic ventricular dysfunction and prominent dysrhythmias. However, little is known about how diverse mutations in LMNA can vary in molecular pathways leading to DCM other than affecting cardiomyocytes (CMs). Induced pluripotent stem cells (iPSCs) retain the genetic identity of the individual donor and enable the generation of patient-specific cells to enhance Precision Medicine. We generated CMs and cardiac fibroblasts (CFs) from patient-specific iPSCs harboring various LMNA mutations to compare their cellular phenotypes. Hypothesis: We hypothesize that different mutation locations within LMNA lead to DCM by affecting both CMs and CFs. Methods: iPSCs from seven DCM patients with heterozygous mutations in LMNA (M1I, R216C/R399H, R216C#1, R216C#2, R335Q, R377H and R541C) were differentiated into CMs and CFs. Nuclear morphology, sarcomere structure, molecular signatures, cell migration rate, and RNA-sequencing were analyzed in CMs and CFs. Results: Immunostaining of lamin A/C displayed no discernable nuclear deformity in LMNA -mutated CMs, whereas CFs showed dramatic nuclear morphologic defects of varying degrees between patient cell lines. Staining of troponin T revealed pronounced sarcomere disarray due to LMNA mutation. LMNA -patient lines exhibited dysregulated expression of connexin 43, MMP1, collagen 1 and 3, and fibronectin 1 in CFs assessed by qPCR and immunoblot, and reduced cell migration rate detected by wound healing assay. RNA-sequencing of variant R377H demonstrated differentially expressed genes in both CMs (up:1500, down:2630, padj<0.05) and CFs (up:491, down:783, padj<0.05). LMNA R377H CMs showed altered Gene Ontology terms including MAPK pathway, cardiac muscle contraction and calcium signaling pathway. Enrichment analysis of R377H CFs highlighted extracellular matrix constituent and organization pathway. Conclusions: CFs exhibit mutation-specific molecular and cellular pathophysiology in LMNA related DCM. Further functional studies may uncover novel therapeutic targets in cardiac fibroblasts for the treatment of laminopathies.