Abstract 3105: Lamin Mutations: Single Nuclei Multiomic Analysis Reveals Endothelial Cell Dysfunction And Its Cardiovascular Implications

Can Genetic Profile of Patients Undergoing Heart Transplantation Alter Clinical Decisions?

Cardiolaminopathies are autosomal dominant genetic diseases caused by mutations in the LMNA gene which encodes the nucleus envelope protein Lamin AC.1 There are now 12 phenotypes associated with LMNA mutations, both autosomal dominant and recessive, the majority of which have cardiac involvement (http://omim.org/entry/150330). These include dilated cardiomyopathy (DCM) associated with conduction disease, atrial and ventricular arrhythmias, and phenotypes mimicking arrhythmogenic right ventricular cardiomyopathy.2 See Article by Hoorntje et al Cardiolaminopathies are characterized by high arrhythmogenic risk3 that may manifest in mildly dilated hearts and may be the first clinical presentation of the disease. Fatal arrhythmias may occur even when left ventricular dysfunction does not reach the cutoff value established by guidelines for implantable cardioverter–defibrillator implantation for the primary prevention of sudden death.4 Clinical cardiologists involved in programs dedicated to familial cardiomyopathies are well aware of the arrhythmogenic risk of cardiolaminopathies. The latest ESC Guidelines for primary prevention of sudden cardiac death recommend (Class IIa; Level of Evidence B) implantable cardioverter–defibrillator in patients with DCM, a confirmed disease-causing LMNA mutation, and the following clinical risk factors: nonsustained ventricular tachycardia during ambulatory electrocardiographic monitoring, left ventricular ejection fraction <45% on initial evaluation, male sex, and nonmissense mutations.5 The genetic diagnosis of laminopathy in DCM patients impacts therapeutic and prognostic strategies. For example, the percutaneous catheter ablation of sustained monomorphic ventricular tachycardia in LMNA cardiomyopathy could be an important consideration. Poor outcomes, including high rates of recurrence of arrhythmia, progression to end-stage heart failure, and high mortality, have been reported in patients with cardiolaminopathies, wherein 91% of patients experienced ≥1 ventricular tachycardia recurrence, 44% received or were awaiting mechanical circulatory support or transplant for end-stage heart failure, and 26% died during a median 7-month follow-up.6 Use of β-blockers in conduction disease should be postponed until after pacemaker …

Phenotypic plasticity of mutations in LMNA, which encodes Lamin A/C, is unsurpassed by any other gene. Mutations in LMNA are responsible for least a dozen distinct phenotype sthat affect various mesenchymal organs and are collectively referred to as lamino - pathies or less frequently envelopathies. Cardiolaminopathies are a subset of lamino - pathies wherein involvement of the heart is the most prominent feature. The typical phenotype of cardiolaminopathies encompasses dilated cardiomyopathy (DCM) and conduction defects. LMNA is probably the most common causal gene for human DCM, being responsible up to 8% of all familial DCM. Several hundred mutations in the LMNA gene have already been described. The p.R644C mutation is the most commonly reported mutation in cardiolaminopathies. The phenotype in cardiolaminopathies is notable for a rapid progression of cardiac failure, conduction defects and arrhythmias, often necessitating implantation of a pacemaker and/or a defibrillator. The molecular pathogenesis of cardiolamino - pathies is poorly understood. Studies in animal models and cultured cells suggest involvement of the Mitogen-Activated Protein Kinase (MAPK) and transforming growth factor –β1 pathways. Comprehensive molecular genetics studies complemented with mechanistic studies are needed to delineate the mechanistic underpinnings of cardiolaminopathies, prerequisite for the ultimate cure of these potentially deadly disorders.

See related article, pages 1032–1039 Hypertension, diabetes, and atherosclerosis are three diseases responsible for the majority of cardiovascular morbidity in humans. Not only are they independent risk factors, but they clearly potentiate and complement each other in the pathogenesis of cardiovascular disease. The first manifestation of end-organ damage in these diseases is the vascular remodeling of small (resistance) and large (conductance), and this precedes the development of cardiac hypertrophy, renal insufficiency or stroke.1 This vascular remodeling is characterized by hyperplasia, hypertrophy, and apoptosis of smooth muscle cells (SMCs) and vascular fibrosis, caused by increased extracellular matrix deposition of total collagen, and changes in ratio of type I/III collagen, fibronectin, and proteoglycans.1 Angiotensin II (Ang II) has recently emerged as a key mediator of vascular fibrosis (sclerosis) both in humans and animal models of hypertension because of its pleiotropic effects of SMCs, fibroblasts, and inflammatory cells. Angiotensin converting enzyme inhibitors (ACE-I) and angiotensin receptor antagonists (ARB) diminish cardiac hypertrophy and vascular fibrosis in animal models of hypertension.1,2 In human clinical trials of hypertension, ARBs reverse vascular remodeling in resistance arteries and new incidence of strokes more potently than any other antihypertensive agents (beta-blockers or calcium channel blockers), despite similar reduction in blood pressure.3,4 This provides strong evidence that targeting the mechanisms involved in arterial remodeling process may provide the key in preventing the long-term consequences of hypertension or atherosclerosis in humans. However, deciphering the intimate mechanisms of in vivo vascular remodeling has become very complex because of enormous reciprocal interactions between Ang II and other growth factors. Ang II exerts its effects not only by activating its specific receptors, but also by transactivating other growth factor receptors such as epidermal growth factor …

Temporal Relationship of Conduction System Disease and Ventricular Dysfunction in LMNA Cardiomyopathy

Lamins are nuclear intermediate filament proteins. They provide mechanical stability, organize chromatin and regulate transcription, replication, nuclear assembly and nuclear positioning. Recent studies provide new insights into the role of lamins in development, differentiation and tissue response to mechanical, reactive oxygen species and thermal stresses. These studies also propose the existence of separate filament networks for A- and B-type lamins and identify new roles for the different networks. Furthermore, they show changes in lamin composition in different cell types, propose explanations for the more than 14 distinct human diseases caused by lamin A and lamin C mutations and propose a role for lamin B1 in these diseases.

BackgroundDilated cardiomyopathy (DCM), which is characterized by enlarged ventricular dimensions and systolic dysfunction, is the most common type of cardiomyopathy. Mutations in the LMNA gene are reported in approximately 10% of familial DCM cases. However, the mechanism of LMNA mutations in human DCM remains unclear.Material/MethodsWe used the GSE36502 and GSE123916 datasets to obtain gene expression profiles from LMNA-related DCM mice and to identify differentially expressed genes (DEGs). Crucial function and pathway enrichment analyses of DEGs were performed. Protein–protein interaction (PPI) network analysis was carried out to identify the top 10 hub genes, which were validated using reverse transcription-polymerase chain reaction (RT-PCR) to find target genes. Weighted gene co-expression network analysis (WGCNA) was used to explore the module relevant to external traits of LMNA-related DCM mice. Transcription factors (TFs) for the selected genes were analyzed using NetworkAnalyst.ResultsA total of 156 common DEGs (co-DEGs) were identified, including 80 up-regulated and 76 down-regulated genes. The enriched biological functions and pathways were oxidative stress, regulation of apoptosis, regulation of fibrosis, and MAPK pathways. Five target genes (Timp1, Hmox1, Spp1, Atf3, and Adipoq) were verified after RT-PCR. Most co-DEGs were discovered to be related to the development of external traits. Three TFs (ELF1, ETS1, and NRF1) showed close interactions with the hub genes.ConclusionsOur study used integrated bioinformatic analyses and revealed some important genes in mice with LMNA-related DCM, which could provide novel insights into the mechanism underlying human LMNA-related DCM.

LMNA gene encodes the ubiquitous protein lamin A/C, which is a component of the fibrous meshwork of intermediate filaments located at the inner surface of the nuclear envelope (Worman 2012). Lamin A/C are splicing variants encoded by the LMNA gene and share the first 566 amino acids. Mutations in LMNA gene are recently known to be responsible for more than 10 different clinical syndromes, most of which show overlapping features. Worman and Bonne (2007) classified the disorders into four major types: diseases of skeletal and cardiac muscle, lipodystrophy syndromes, peripheral neuropathy and premature ageing. Of the skeletal and cardiac muscles types, limb–girdle muscular dystrophy type 1B (LGMD1B), autosomal dominant Emery–Dreifuss muscular dystrophy 2 (EDMD2) and dilated cardiomyopathy 1A (CMD1A) are common (Capell and Collins 2006). Among the muscle type disorders related to LMNA gene mutations, LGMD1B is slowly progressive, with age-related atrioventricular cardiac conduction disturbance, dilated cardiomyopathy, muscle weakness and the absence of early contractures. However, EDMD2 is characterized by early contractures of elbows and Achilles tendons, slowly progressive muscle wasting and weakness, and cardiomyopathywith a variable degree of conduction system disease (Capell and Collins 2006). In this report, we characterized a large Chinese family with atrioventricular block (AVB) as the most prominent feature, with concomitant moderate pelvic limb skeletal muscle defects. Due to the medical history, clinical manifestations and neurological/cardiological examinations, this family was diagnosed with autosomal LGMD1B. Then, a mutation

BackgroundUp to fifty percent of idiopathic dilated cardiomyopathy (DCM) has a familial basis. Variants can occur in over 40 genes, though truncating variants in the sarcomeric gene titin account for...

Characterization of Adiposity and Metabolism in Lmna-Deficient Mice

Mutations in the LMNA gene (encoding lamin A/C proteins) cause several human cardiac diseases, including dilated cardiomyopathies (LMNA-DCM). The main clinical risks in LMNA-DCM patients are sudden cardiac death and progressive left ventricular ejection fraction deterioration, and therefore most human and animal studies have sought to define the mechanisms through which LMNA mutations provoke cardiac alterations, with a particular focus on cardiomyocytes. To investigate if LMNA mutations also cause vascular alterations that might contribute to the etiopathogenesis of LMNA-DCM, we generated and characterized Lmnaflox/floxSM22αCre mice, which constitutively lack lamin A/C in vascular smooth muscle cells (VSMCs), cardiac fibroblasts, and cardiomyocytes. Like mice with whole body or cardiomyocyte-specific lamin A/C ablation, Lmnaflox/floxSM22αCre mice recapitulated the main hallmarks of human LMNA-DCM, including ventricular systolic dysfunction, cardiac conduction defects, cardiac fibrosis, and premature death. These alterations were associated with elevated expression of total and phosphorylated (active) Smad3 and cleaved (active) caspase 3 in the heart. Lmnaflox/floxSM22αCre mice also exhibited perivascular fibrosis in the coronary arteries and a switch of aortic VSMCs from the ‘contractile’ to the ‘synthetic’ phenotype. Ex vivo wire myography in isolated aortic rings revealed impaired maximum contraction capacity and an altered response to vasoconstrictor and vasodilator agents in Lmnaflox/floxSM22αCre mice. To our knowledge, our results provide the first evidence of phenotypic alterations in VSMCs that might contribute significantly to the pathophysiology of some forms of LMNA-DCM. Future work addressing the mechanisms underlying vascular defects in LMNA-DCM may open new therapeutic avenues for these diseases.

Lamins A/C, encoded by LMNA gene, are constituents of the nuclear lamina, a meshwork of proteins underneath the nuclear envelope. Human disorders linked to LMNA mutations are known as laminopathies and include cardiac and muscular dystrophies, lipodystrophies, progeroid syndromes and overlapping phenotypes. Associated with this wide clinical variability, there is also a large allelic heterogeneity that is still expanding. New links between LMNA mutations and human diseases with already known or unknown molecular cause have been established. In other cases, no evidence that a human disease is another laminopathy has been found. Moreover, the recent availability of high‐throughput sequencing technologies has made feasible to uncover the disease‐causing mutations in patients who were originally diagnosed with laminopathies but whose mutational analysis of candidate gene did not reveal any change in LMNA . Key Concepts Mutations in the LMNA gene, encoding A‐type lamin proteins, lead to a wide range of diseases known as laminopathies. The clinical and allelic heterogeneity of laminopathies is still expanding. Studies of the molecular mechanisms underlying laminopathies have led to advances in the understanding of lamin A functions and developing novel therapeutic strategies. A group of progeroid phenotypes is phenocopy of progeroid laminopathies. High‐throughput sequencing technologies have made it possible to uncover the disease‐causing mutations in genes that encode proteins involved in nuclear envelope architecture or in control of genomic stability. Nuclear alterations are linked to human ageing.

Dilated cardiomyopathy (DCM), characterized by left ventricular dilatation and systolic dysfunction, constitutes a significant cause for heart failure, sudden cardiac death or need for heart transplantation. Lamin A/C gene (LMNA) on chromosome 1p12 is the most significant disease gene causing DCM and has been reported to cause 7–9% of DCM leading to cardiac transplantation. We have previously performed cardiac magnetic resonance imaging (MRI) to LMNA carriers to describe the early phenotype. Clinically, early recognition of subjects at risk of developing DCM would be important but is often difficult. Thus we have earlier used the MRI findings of these LMNA carriers for creating a model by which LMNA carriers could be identified from the controls at an asymptomatic stage. Some LMNA mutations may cause lipodystrophy. To characterize possible effects of LMNA mutations on lipid profile, we set out to apply global serum lipidomics using Ultra Performance Liquid Chromatography coupled to mass spectrometry in the same LMNA carriers, DCM patients without LMNA mutation and controls. All DCM patients, with or without LMNA mutation, differed from controls in regard to distinct serum lipidomic profile dominated by diminished odd-chain triglycerides and lipid ratios related to desaturation. Furthermore, we introduce a novel approach to identify associations between the molecular lipids from serum and the MR images from the LMNA carriers. The association analysis using dependency network and regression approaches also helped us to obtain novel insights into how the affected lipids might relate to cardiac shape and volume changes. Our study provides a framework for linking serum derived molecular markers not only with clinical endpoints, but also with the more subtle intermediate phenotypes, as derived from medical imaging, of potential pathophysiological relevance.

We investigated the role of LMNA in adipose tissue by developing a novel mouse model of lipodystrophy. Transgenic mice were generated that express the LMNA mutation that causes familial partial lipodystrophy of the Dunnigan type (FPLD2). The phenotype observed in FPLD-transgenic mice resembles many of the features of human FPLD2, including lack of fat accumulation, insulin resistance, and enlarged, fatty liver. Similar to the human disease, FPLD-transgenic mice appear to develop normally, but after several weeks they are unable to accumulate fat to the same extent as their wild-type littermates. One poorly understood aspect of lipodystrophies is the mechanism of fat loss. To this end, we have examined the effects of the FPLD2 mutation on fat cell function. Contrary to the current literature, which suggests FPLD2 results in a loss of fat, we found that the key mechanism contributing to the lack of fat accumulation involves not a loss, but an apparent inability of the adipose tissue to renew itself. Specifically, preadipocytes are unable to differentiate into mature and fully functional adipocytes. These findings provide insights not only for the treatment of lipodystrophies, but also for the study of adipogenesis, obesity, and insulin resistance.

141st ENMC International Workshop Inaugural Meeting of the EURO-Laminopathies Project Nuclear Envelope-linked Rare Human Diseases: From Molecular Pathophysiology towards Clinical Applications 10–12 March 2006, Naarden, The Netherlands