Satu Kärkkäinen

Genetics of dilated cardiomyopathy

Dilated cardiomyopathy (DCM) is a myocardial disease characterized by dilatation and impaired systolic function of the left or both ventricles. The etiology of DCM is multifactorial, and many different clinical conditions can lead to the phenotype of DCM. During recent years it has become evident that genetic factors play an important role in the etiology and pathogenesis of idiopathic DCM. The genetics of DCM have been under intensive investigation lately, and thereby the knowledge on the genetic basis of DCM has increased rapidly. The genetic background of the disease seems to be relatively heterogeneous, and the disease‐associated mutations concern mostly single families and only few affected patients. Disease‐associated mutations have been detected e.g. in genes encoding sarcomere, cytoskeletal, and nuclear proteins, as well as proteins involved with regulation of Ca2+ metabolism. The mechanisms, by which mutations eventually result in clinical heart failure, are complex and not yet totally resolved. DCM causes considerable morbidity and mortality. Better knowledge of the genetic background and disease‐causing mechanisms would probably help us in focusing early treatment on right subjects and potentially also developing new treatment modalities and improving cardiac outcome in the affected patients. This review deals with DCM of genetic origin.

Identification of preferred protein interactions by phage‐display of the human Src homology‐3 proteome

We have determined the human genome to contain 296 different Src homology‐3 (SH3) domains and cloned them into a phage‐display vector. This provided a powerful and unbiased system for simultaneous assaying of the complete human SH3 proteome for the strongest binding to target proteins of interest, without the limitations posed by short linear peptide ligands or confounding variables of more indirect methods for protein interaction screening. Studies involving three ligand proteins, human immunodeficiency virus‐1 Nef, p21‐activated kinase (PAK)2 and ADAM15, showed previously reported as well as novel SH3 partners with nanomolar affinities specific for them. This argues that SH3 domains may have a more dominant role in directing cellular protein interactions than has been assumed. Besides showing potentially important new SH3‐directed interactions, these studies also led to the discovery of novel signalling proteins, such as the PAK2‐binding adaptor protein POSH2 and the ADAM15‐binding sorting nexin family member SNX30.

Mutations in the cardiac myosin-binding protein C gene are the predominant cause of familial hypertrophic cardiomyopathy in eastern Finland

Hypertrophic cardiomyopathy (HCM) is a genetic disorder characterized by cardiac hypertrophy caused by mutations in genes encoding sarcomere proteins. This study screened all patients with HCM from the Kuopio University Hospital region in eastern Finland for variants in the cardiac myosin-binding protein C gene ( MYBPC3). All 35 exons of MYBPC3 were screened by the single-strand conformation polymorphism method in 37 unrelated patients with HCM. In MYBPC3 we identified seven novel (Gln1061X, IVS5-2A→C, IVS14-13G→A, Ex25ΔLys, Pro147Leu, Ser236Gly, and Arg1138His) and two previously reported (Arg326Gln, Val896Met) variants, all of which are predicted to affect the structure of the encoded protein. Four of the nine variants, a nonsense mutation Gln1061X, a splice acceptor mutation (IVS5-2A→C), a novel substitution in intron 14 (IVS14-13G→A), and a novel 3-bp deletion in exon 25 (Ex25ΔLys) were concluded to be disease-causing mutations because they cosegregated with the HCM phenotype or were absent in more than 200 normal chromosomes, or both. The mutation Gln1061X was found most frequently, being present in 6 families (23 subjects) while the other three mutations were found in single families each. Haplotype analysis indicated a likely founder effect among the families carrying the Gln1061X mutation. We found four novel mutations in MYBPC3, accounting for approx. 38% of familial and 24% of all cases of HCM. In our previous and unpublished studies no more frequent cause of HCM has been found in genetic analyses of other eight sarcomeric proteins. Consequently MYBPC3 is the predominant gene for HCM in eastern Finland. In addition, several amino acid substitutions in MYBPC3 suspected to be not associated with HCM were identified, indicating that some of the missense variants found in MYBPC3 are possibly not disease-causing mutations.

Late gadolinium enhanced cardiovascular magnetic resonance of lamin A/C gene mutation related dilated cardiomyopathy

BackgroundThe purpose of this study was to identify early features of lamin A/C gene mutation related dilated cardiomyopathy (DCM) with cardiovascular magnetic resonance (CMR). We characterise myocardial and functional findings in carriers of lamin A/C mutation to facilitate the recognition of these patients using this method. We also investigated the connection between myocardial fibrosis and conduction abnormalities.MethodsSeventeen lamin A/C mutation carriers underwent CMR. Late gadolinium enhancement (LGE) and cine images were performed to evaluate myocardial fibrosis, regional wall motion, longitudinal myocardial function, global function and volumetry of both ventricles. The location, pattern and extent of enhancement in the left ventricle (LV) myocardium were visually estimated.ResultsPatients had LV myocardial fibrosis in 88% of cases. Segmental wall motion abnormalities correlated strongly with the degree of enhancement. Myocardial enhancement was associated with conduction abnormalities. Sixty-nine percent of our asymptomatic or mildly symptomatic patients showed mild ventricular dilatation, systolic failure or both in global ventricular analysis. Decreased longitudinal systolic LV function was observed in 53% of patients.ConclusionsCardiac conduction abnormalities, mildly dilated LV and depressed systolic dysfunction are common in DCM caused by a lamin A/C gene mutation. However, other cardiac diseases may produce similar symptoms. CMR is an accurate tool to determine the typical cardiac involvement in lamin A/C cardiomyopathy and may help to initiate early treatment in this malignant familiar form of DCM.

A novel mutation, Arg71Thr, in the δ-sarcoglycan gene is associated with dilated cardiomyopathy

Approximately 20–35% of cases of idiopathic dilated cardiomyopathy are familial. DCM-associated mutations have been reported in 13 genes including the desmin, δ-sarcoglycan, and metavinculin genes. This study screened for variants in these genes in Finnish patients with DCM. All coding regions of the desmin and δ-sarcoglycan genes and the metavinculin-specific exon of the vinculin gene were screened in 52 DCM patients from eastern Finland by PCR-SSCP. We detected a novel mutation, Arg71Thr, in the δ-sarcoglycan gene in two members of a small DCM family. One of the mutation carriers fulfills diagnostic criteria for DCM and is also symptomatic. The other mutation carrier has slightly dilated left ventricle and well preserved systolic function. Therefore carriers of the Arg71Thr mutation had a relatively mild phenotype and a late onset of the disease. Disease-associated mutations were not found in the desmin gene or the metavinculin-specific exon of the vinculin gene. We conclude that the desmin and δ-sarcoglycan genes are not predominant disease-causing genes in patients with DCM in eastern Finland.

Two novel mutations in the β-myosin heavy chain gene associated with dilated cardiomyopathy

Dilated cardiomyopathy (DCM) is familial in approximately 20–35% of cases of idiopathic DCM. Several mutations in the different sarcomere protein genes have been reported to cause DCM.

Serum Lipidomics Meets Cardiac Magnetic Resonance Imaging: Profiling of Subjects at Risk of Dilated Cardiomyopathy

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.

Early familial dilated cardiomyopathy: identification with determination of disease state parameter from cine MR image data.

PURPOSE To characterize early changes in cardiac anatomy and function for lamin A/C gene (LMNA) mutation carriers by using magnetic resonance (MR) imaging and to develop tools to analyze and visualize the findings. MATERIALS AND METHODS The ethical review board of the institution approved the study, and informed written consent was obtained. The patient group consisted of 12 subjects, seven women (mean age, 36 years; age range, 18-54 years) and five men (mean age, 28 years; age range, 18-39 years) of Finnish origin, who were each heterozygotes with one LMNA mutation that may cause familial dilated cardiomyopathy (DCM). All the subjects were judged to be healthy with transthoracic echocardiography. The control group consisted of 14 healthy subjects, 11 women (mean age, 41 years; range, 23-54 years) and three men (mean age, 45 years; range, 34-57 years), of Finnish origin. Cine steady state free precession MR imaging was performed with a 1.5-T system. The volumes, wall thickness, and wall motion of both left ventricle (LV) and right ventricle were assessed. A method combining multiple MR image parameters was used to generate a global cardiac function index, the disease state parameter (DSP). A visual fingerprint was generated to assess the severity of familial DCM. RESULTS The mean DSP of the patient group (0.69 +/- 0.15 [standard deviation]) was significantly higher than that of the control group (0.32 +/- 0.13) (P = .00002). One subject had an enlarged LV. CONCLUSION Subclinical familial DCM was identified by determination of the DSP with MR imaging, and this method might be used to recognize familial DCM at an early stage.

Left Atrial Appendage Volume Increased in More Than Half of Patients with Cryptogenic Stroke

Background Ischemic strokes without a well-defined etiology are labeled as cryptogenic, and account for 30–40% of strokes in stroke registries. The left atrial appendage (LAA) is the most typical origin for intracardiac thrombus formation when associated with atrial fibrillation. Here, we examined whether increased LAA volume detected with cardiac computed tomography (cCT) constitutes a risk factor in cryptogenic stroke patients. Methods This study included 82 stroke/TIA patients (57 males; mean age, 58 years) with a diagnosis of cryptogenic stroke after extensive radiological and cardiological investigations. Cases were classified using the TOAST criteria modified by European Association of Echocardiography recommendations for defining cardiac sources of embolism. Forty age- and gender-matched control subjects without cardiovascular diseases were selected for pair-wise comparisons (21 males; mean age, 54 years). LAA volume adjusted for body surface area was measured three dimensionally by tracing the LAA borders on electrocardiogram-gated CT slices. Results In control subjects, mean LAA volume was 3.4±1.1 mL/m2. Mean+2SD, which was considered the upper limit for normal LAA volume was 5.6 mL/m2. In paired Student t-test between the patient group and matched controls, LAA volume was 67% larger in cryptogenic stroke/TIA patients (5.7±2.0 mL/m2 vs. 3.4±1.1 mL/m2; P<0.001). Forty-five (55%) patients with cryptogenic stroke/TIA had enlarged LAA. Conclusion LAA is significantly enlarged in more than half of patients with cryptogenic stroke/TIA. LAA thrombosis may contribute to the pathogenesis of stroke in patients considered to have cryptogenic stroke after conventional evaluation.

Arachidonic acid increases matrix metalloproteinase 9 secretion and expression in human monocytic MonoMac 6 cells

BackgroundDietary fatty acids may modulate inflammation in macrophages of the atherosclerotic plaque, affecting its stability. The n-6 polyunsaturated fatty acid (PUFA) arachidonic acid (AA) generally promotes inflammation, while the PUFAs of the n-3 series eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA) and docosahexaenoic acid (DHA) are considered anti-inflammatory. We determined how these PUFAs influence MMP-9 expression and secretion by the human monocytic cell line (MonoMac 6) at baseline and after 24-hour exposure. MMP-9 protein was measured by zymography and relative levels of MMP-9 mRNA were determined using quantitative real time PCR.ResultsSupplementation with AA (but not the n-3 fatty acids) increased, in a dose-dependent manner, expression of MMP-9 protein. This stimulation was regulated at the mRNA level. MMP-9 secretion started after 1 h of incubation and could not be prevented by simultaneous presence of n-3 series fatty acids. Finally, the secretion could be attenuated by LY 294002, a specific phosphatidylinositol-3-kinase (PI3K) inhibitor and by SH-5, a selective Akt inhibitor, suggesting that activation of PI3K by AA leads to augmented and sustained MMP-9 production.ConclusionThis study shows that of the PUFA studied, AA alone influences the expression of MMP-9, which might have implications in MMP-9 induced plaque rupture.