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Muchir, A.

Publications and source records attributed to Muchir, A..

3 recordsLinked to original sources
bioRxiv2025

The Mutated p.H222P A-type Lamins Drive Loxl2-Mediated Extracellular Matrix Remodeling in Both Patient-Derived Cardiomyocytes and Mouse Models of Dilated Cardiomyopathy

LMNA cardiomyopathy, caused by mutations in the LMNA gene, is a severe form of dilated cardiomyopathy characterized by arrhythmias, contractile dysfunction, and increased myocardial fibrosis, which impairs left ventricular function and predisposes to heart failure. While the disease has been well characterized, a lack of insight into the pathogenesis impeded the development of therapies. We here used patient-derived LMNA p.H222P cardiomyocytes (hiPSC-CMs) and their isogenic controls and a LmnaH222P/H222P mouse model to dissect abnormal cardiac mechanisms leading to the development of the disease. We showed that LMNA p.H222P hiPSC-CMs exhibit elevated diastolic calcium levels and hypocontractility. They displayed nuclear shape abnormalities, a hallmark of LMNA cardiomyopathy, associated with altered chromosome spatial organization and gene expression profiles. Using transcriptomic analysis, we further revealed that genes related to cardiac extracellular matrix (ECM) remodeling, deposition, and components are dysregulated in both LMNA p.H222P hiPSC-CMs and mutated mice, suggesting a conserved pathogenic mechanism across species. Conversely, molecular inhibition of Loxl2, a key component of the ECM establishment, preserved the cardiac function in vivo. Taken together, our findings suggest that targeting Loxl2 could be a promising therapeutic strategy to maintain cardiac function in LMNA cardiomyopathy.

pathology↗
bioRxiv2024

The p.H222P lamin A/C mutation induces heart failure via impaired mitochondrial calcium uptake in human cardiac laminopathy

BackgroundMutations in the LMNA gene, which encodes lamin A/C, cause a variety of diseases known as laminopathies. Some mutations are particularly associated with the occurrence of dilated cardiomyopathy and heart failure, but the genotype-phenotype relationship and underlying mechanisms are unclear. Here, we used induced pluripotent stem cells (hiPSCs) from a patient carrying a LMNA point mutation (c.665A>C, p.His222Pro) to investigate the mechanisms leading to contractile dysfunction. MethodsLMNA p.H222P mutant and a CRISPR/Cas9 corrected isogenic control hiPSCs clones were differentiated into cardiomyocytes (hiPSC-CMs). Immunofluorescence staining was performed on hiPSC-CMs to quantify their sarcomere organization (SarcOrgScore) using a Matlab code. Ring-shaped cardiac 3D organoids were generated to compare the contractile properties of the two clones. Calcium transients in mutant and corrected hiPSC-CMs were measured by live confocal imaging. Mitochondrial respiration parameters were measured by Seahorse. ResultshiPSC-CMs were generated from the LMNA mutant and the corrected hiPSCs with no difference in the differentiation yield (proportion of troponin-positive cells: 95.0% for LMNA p.H222P vs. 95.1% for Ctrl-iso1, p=0.726). hiPSC-CMs displayed well-formed sarcomeres and their organization was similar between the two cell lines. However, cardiac 3D organoids generated with LMNA p.H222P hiPSC-CMs showed an impaired contractility compared to control organoids. Calcium transient recordings in LMNA p.H222P mutant cardiomyocytes showed a significantly higher calcium transient amplitude with a significantly slower calcium re-uptake. Transcriptomic analyses suggested a global mitochondrial dysfunction and in particular an impaired mitochondrial calcium uptake with a significantly decreased expression of the mitochondrial calcium uniporter (MCU). This decrease in MCU expression was confirmed by western blot and was accompanied by an increased MICU1:MCU, as well as an increased PDH Ser232 and PDH Ser300 phosphorylation, indicating a decreased mitochondrial calcium uptake in the LMNA mutant hiPSC-CMs. Measurement of mitochondrial respiration showed lower basal and maximal respiration in LMNA p.H222P hiPSC-CMs. Consistently, the ATP levels were significantly lower in LMNA p.H222P hiPSC-CMs as compared to isogenic controls. ConclusionsLMNA p.H222P mutant hiPSC-CMs exhibit contractile dysfunction associated with mitochondrial dysfunction with impaired MCU complex activity, decreased mitochondrial calcium homeostasis and reduced mitochondrial energy production. NOVELTY AND SIGNIFICANCEO_ST_ABSWhat is known?C_ST_ABS- Mutations in LMNA, which encodes the nuclear lamins A/C, cause a variety of diseases (called laminopathies), which can involve the cardiac muscle leading to dilated cardiomyopathy and systolic heart failure. - The pathological mechanisms linking the nuclear envelope abnormalities induced by LMNA mutations to the development of a reduced cardiac muscle contractility are not well understood. What new information does this article contribute?- LMNA mutant cardiomyocytes have a profound mitochondrial dysfunction with impaired MCU complex activity, decreased mitochondrial calcium homeostasis, and reduced mitochondrial energy production. - Our study uncovers an unappreciated pathophysiological mechanism and opens new possibilities by suggesting MCU activators as a novel therapeutic for patients with LMNA cardiomyopathy.

pathology↗
bioRxiv2021

Bioengineering a Miniaturized In Vitro 3D Myotube Contraction Monitoring Chip For Modelization of Muscular Dystrophies

Quantification of skeletal muscle functional contraction is essential to assess the outcomes of therapeutic procedures for muscular disorders. Muscle three-dimensional "Organ-on-chip" models usually require a substantial amount of biological material, which is problematic in the context of limited patient sample. Here we developed a miniaturized 3D myotube culture chip with contraction monitoring capacity. Optimized micropatterned substrate design enabled to obtain high culture yields in tightly controlled microenvironments. Spontaneous contractions in myotubes derived from primary human myoblasts were observed. Analysis of nuclear morphology confirmed a similar organization between obtained myotubes and in vivo myofibers. LMNA-related Congenital Muscular Dystrophy (L-CMD) was modelled with successful development of mutant 3D myotubes displaying contractile dysfunction. This technology can thus be used to study contraction characteristics and evaluate how diseases affect muscle organization and force generation. Importantly, it requires significantly fewer starting materials than current systems, which should allow to substantially improve drug screening capability.

bioengineering↗