The R403Q Myosin Mutation Implicated in Familial Hypertrophic Cardiomyopathy Causes Disorder at the Actomyosin Interface

Mutations in virtually all of the proteins comprising the cardiac muscle sarcomere have been implicated in causing Familial Hypertrophic Cardiomyopathy (FHC). Mutations in the beta-myosin heavy chain (MHC) remain among the most common causes of FHC, with the widely studied R403Q mutation resulting in an especially severe clinical prognosis. In vitro functional studies of cardiac myosin containing the R403Q mutation have revealed significant changes in enzymatic and mechanical properties compared to wild-type myosin. It has been proposed that these molecular changes must trigger events that ultimately lead to the clinical phenotype.Here we examine the structural consequences of the R403Q mutation in a recombinant smooth muscle myosin subfragment (S1), whose kinetic features have much in common with slow beta-MHC. We obtained three-dimensional reconstructions of wild-type and R403Q smooth muscle S1 bound to actin filaments in the presence (ADP) and absence (apo) of nucleotide by electron cryomicroscopy and image analysis. We observed that the mutant S1 was attached to actin at highly variable angles compared to wild-type reconstructions, suggesting a severe disruption of the actin-myosin interaction at the interface.These results provide structural evidence that disarray at the molecular level may be linked to the histopathological myocyte disarray characteristic of the diseased state

3,3′Diindolylmethane Suppresses Vascular Smooth Muscle Cell Phenotypic Modulation and Inhibits Neointima Formation after Carotid Injury

3,3'Diindolylmethane (DIM), a natural phytochemical, has shown inhibitory effects on the growth and migration of a variety of cancer cells; however, whether DIM has similar effects on vascular smooth muscle cells (VSMCs) remains unknown. The purpose of this study was to assess the effects of DIM on the proliferation and migration of cultured VSMCs and neointima formation in a carotid injury model, as well as the related cell signaling mechanisms.DIM dose-dependently inhibited the platelet-derived growth factor (PDGF)-BB-induced proliferation of VSMCs without cell cytotoxicity. This inhibition was caused by a G0/G1 phase cell cycle arrest demonstrated by fluorescence-activated cell-sorting analysis. We also showed that DIM-induced growth inhibition was associated with the inhibition of the expression of cyclin D1 and cyclin-dependent kinase (CDK) 4/6 as well as an increase in p27(Kip1) levels in PDGF-stimulated VSMCs. Moreover, DIM was also found to modulate migration of VSMCs and smooth muscle-specific contractile marker expression. Mechanistically, DIM negatively modulated PDGF-BB-induced phosphorylation of PDGF-recptorβ (PDGF-Rβ) and the activities of downstream signaling molecules including Akt/glycogen synthase kinase(GSK)3β, extracellular signal-regulated kinase1/2 (ERK1/2), and signal transducers and activators of transcription 3 (STAT3). Our in vivo studies using a mouse carotid arterial injury model revealed that treatment with 150 mg/kg DIM resulted in significant reduction of the neointima/media ratio and proliferating cell nuclear antigen (PCNA)-positive cells, without affecting apoptosis of vascular cells and reendothelialization. Infiltration of inflammatory cells was also inhibited by DIM administration.These results demonstrate that DIM can suppress the phenotypic modulation of VSMCs and neointima hyperplasia after vascular injury. These beneficial effects on VSMCs were at least partly mediated by the inhibition of PDGF-Rβ and the activities of downstream signaling pathways. The results suggest that DIM has the potential to be a candidate for the prevention of restenosis

Screening mutations in myosin binding protein C3 gene in a cohort of patients with Hypertrophic Cardiomyopathy

<p>Abstract</p> <p>Background</p> <p><it>MyBPC3 </it>mutations are amongst the most frequent causes of hypertrophic cardiomyopathy, however, its prevalence varies between populations. They have been associated with mild and late onset disease expression. Our objectives were to establish the prevalence of <it>MyBPC3 </it>mutations and determine their associated clinical characteristics in our patients.</p> <p>Methods</p> <p>Screening by Single Strand Conformation Polymorphisms (SSCP) and sequencing of the fragments with abnormal motility of the <it>MyBPC3 </it>gene in 130 unrelated consecutive HCM index cases. Genotype-Phenotype correlation studies were done in positive families.</p> <p>Results</p> <p>16 mutations were found in 20 index cases (15%): 5 novel [D75N, V471E, Q327fs, IVS6+5G>A (homozygous), and IVS11-9G>A] and 11 previously described [A216T, R495W, R502Q (2 families), E542Q (3 families), T957S, R1022P (2 families), E1179K, K504del, K600fs, P955fs and IVS29+5G>A]. Maximum wall thickness and age at time of diagnosis were similar to patients with <it>MYH7 </it>mutations [25(7) vs. 27(8), p = 0.16], [46(16) vs. 44(19), p = 0.9].</p> <p>Conclusions</p> <p>Mutations in <it>MyBPC3 </it>are present in 15% of our hypertrophic cardiomyopathy families. Severe hypertrophy and early expression are compatible with the presence of <it>MyBPC3 </it>mutations. The genetic diagnosis not only allows avoiding clinical follow up of non carriers but it opens new possibilities that includes: to take preventive clinical decisions in mutation carriers than have not developed the disease yet, the establishment of genotype-phenotype relationship, and to establish a genetic diagnosis routine in patients with familial HCM.</p

Transcriptome-scale similarities between mouse and human skeletal muscles with normal and myopathic phenotypes

BACKGROUND: Mouse and human skeletal muscle transcriptome profiles vary by muscle type, raising the question of which mouse muscle groups have the greatest molecular similarities to human skeletal muscle. METHODS: Orthologous (whole, sub-) transcriptome profiles were compared among four mouse-human transcriptome datasets: (M) six muscle groups obtained from three mouse strains (wildtype, mdx, mdx(5cv)); (H1) biopsied human quadriceps from controls and Duchenne muscular dystrophy patients; (H2) four different control human muscle types obtained at autopsy; and (H3) 12 different control human tissues (ten non-muscle). RESULTS: Of the six mouse muscles examined, mouse soleus bore the greatest molecular similarities to human skeletal muscles, independent of the latters' anatomic location/muscle type, disease state, age and sampling method (autopsy versus biopsy). Significant similarity to any one mouse muscle group was not observed for non-muscle human tissues (dataset H3), indicating this finding to be muscle specific. CONCLUSION: This observation may be partly explained by the higher type I fiber content of soleus relative to the other mouse muscles sampled

Myocardial energy depletion and dynamic systolic dysfunction in hypertrophic cardiomyopathy

Evidence indicates that anatomical and physiological phenotypes of hypertrophic cardiomyopathy (HCM) stem from genetically mediated, inefficient cardiomyocyte energy utilization, and subsequent cellular energy depletion. However, HCM often presents clinically with normal left ventricular (LV) systolic function or hyperkinesia. If energy inefficiency is a feature of HCM, why is it not manifest as resting LV systolic dysfunction? In this Perspectives article, we focus on an idiosyncratic form of reversible systolic dysfunction provoked by LV obstruction that we have previously termed the 'lobster claw abnormality' — a mid-systolic drop in LV Doppler ejection velocities. In obstructive HCM, this drop explains the mid-systolic closure of the aortic valve, the bifid aortic pressure trace, and why patients cannot increase stroke volume with exercise. This phenomenon is characteristic of a broader phenomenon in HCM that we have termed dynamic systolic dysfunction. It underlies the development of apical aneurysms, and rare occurrence of cardiogenic shock after obstruction. We posit that dynamic systolic dysfunction is a manifestation of inefficient cardiomyocyte energy utilization. Systolic dysfunction is clinically inapparent at rest; however, it becomes overt through the mechanism of afterload mismatch when LV outflow obstruction is imposed. Energetic insufficiency is also present in nonobstructive HCM. This paradigm might suggest novel therapies. Other pathways that might be central to HCM, such as myofilament Ca2+ hypersensitivity, and enhanced late Na+ current, are discussed