Abstract 58: Modeling Pathogenesis in Familial Hypertrophic Cardiomyopathy Using Patient-Specific Induced Pluripotent Stem Cells

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
Feng lan ◽  
Andrew Lee ◽  
Ping Liang ◽  
Enrique Navarrete ◽  
Li Wang ◽  
...  
Keyword(s):  
Hypertrophic Cardiomyopathy ◽  
Molecular Mechanisms ◽  
Nuclear Translocation ◽  
Molecular Genetic ◽  
Dermal Fibroblasts ◽  
Familial Hypertrophic Cardiomyopathy ◽  
Patient Specific ◽  
Activated T Cells ◽  
Cellular Hypertrophy ◽  
Induced Pluripotent

Background: Hypertrophic cardiomyopathy (HCM) is a prevalent familial cardiac disorder linked to development of heart failure, arrhythmia, and sudden cardiac death. Molecular genetic studies have demonstrated HCM is caused by mutations in genes encoding for the cardiac sarcomere. However, the pathways by which sarcomeric mutations result in myocyte hypertrophy and contractile abnormalities are not well understood. Methods: We aimed to elucidate the molecular mechanisms underlying the development of HCM through the generation of induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) from dermal fibroblasts of a 10 member family, five of whom carry a hereditary HCM missense mutation (Arg663His) in the MYH7 gene. Results: As compared to control iPSC-CMs derived from healthy family members, HCM iPSC-CMs exhibited enlarged cell size, increased atrial natriuretic factor (ANF) expression, nuclear translocation of nuclear factor of activated T-cells (NFAT), and aggravated contractile dysfunction in response to stimulation by β-adrenergic agonists. Interestingly, both video analysis of beating cells and whole cell patch clamping revealed arrhythmia in a significant portion of diseased iPSC-CMs at the single cell level. Ca 2+ imaging demonstrated elevated cytoplasmic Ca 2+ content and irregular transients in HCM iPSC-CMs prior to the onset of cellular hypertrophy, suggesting the HCM phenotype is triggered by dysfunction in Ca 2+ cycling. Treatment of irregular Ca 2+ homeostasis by the Ca 2+ channel blocker verapamil prevented development of cellular hypertrophy and arrhythmia. Conclusions: We hypothesize the cellular abnormalities observed in HCM iPSC-CMs are caused by deficiencies in Ca 2+ regulation. We anticipate our findings will elucidate the mechanisms underlying HCM development and identify novel targets for treatment of the disease.

Abstract 366: Disease Phenotype Assessment Across a Library of iPSC-Derived Cardiomyocytes From Patient Cohorts Carrying Distinct Mutations for Familial Hypertrophic Cardiomyopathy

2015 ◽  
Vol 117 (suppl_1) ◽  
Author(s):  
Jason Tsai ◽  
Jason Lam ◽  
Veronica Sanchez-Freire ◽  
Rishali Gadkari ◽  
Maya Agarwal ◽  
...  
Keyword(s):  
Hypertrophic Cardiomyopathy ◽  
Induced Pluripotent Stem Cell ◽  
Familial Hypertrophic Cardiomyopathy ◽  
Patient Specific ◽  
Multielectrode Array ◽  
Disease Phenotype ◽  
Disease Phenotypes ◽  
Distinct Disease ◽  
Induced Pluripotent

Familial hypertrophic cardiomyopathy (HCM) is the leading cause of sudden cardiac death in the young, and is the most common inherited heart defect affecting 1 in 500 individuals worldwide. Induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) have been demonstrated to model aspects of HCM, but only one iPSC model has been reported for a single HCM mutation in one gene. Here we compare disease phenotypes across a library of patient-specific HCM iPSC-CMs carrying distinct mutations to assess the range of phenotypes that may present in iPSC-CMs derived from different patient cohorts. iPSCs were generated from three patient cohorts carrying known hereditary mutations for HCM in TNNI3, TNNT2, and MYH7 and family-matched controls. Disease phenotypes in patient-specific iPSC-CMs were modeled using immunostaining, Ca2+ imaging, multielectrode array, and video analysis of contractile motion. HCM iPSC-CMs displayed a range of disease phenotypes as assessed by cell size, Ca2+ homeostasis, electrophysiology, and contractile arrhythmia. Different HCM mutations resulted in distinct disease phenotype presentation. Importantly, identical mutations demonstrated similar readouts across multiple lines and clones whereas distinct mutations exhibited differential disease phenotypes. These findings indicate disease-specific iPSC-CMs present with a range of phenotypes for HCM that vary by specific mutation and that iPSC libraries are important for cellular characterization of diseases such as HCM. Figure 1. Derivation and disease phenotype modeling of iPSC-CMs generated from patients carrying distinct familial HCM mutations and family-matched controls.

Abstract P491: Myosin-7 E848G Mutation Induces P53-associated Cell Death That Leads To Impaired Tissue Contractility In Patient-derived Induced Pluripotent Stem Cell Model Of Hypertrophic Cardiomyopathy

2021 ◽  
Vol 129 (Suppl_1) ◽  
Author(s):  
Alexander Loiben ◽  
Clayton Friedman ◽  
Wei-Ming Chen ◽  
Benjamin Chung ◽  
Kai-chun Yang
Keyword(s):  
Cell Death ◽  
Hypertrophic Cardiomyopathy ◽  
Cell Model ◽  
Systolic Function ◽  
Cell Number ◽  
Familial Hypertrophic Cardiomyopathy ◽  
Patient Specific ◽  
Dependent Manner ◽  
Twitch Force ◽  
Induced Pluripotent

Introduction: Familial hypertrophic cardiomyopathy (HCM), affecting 1 in 500 adults, is characterized by idiopathic thickening of the heart and occasional impaired systolic function. Mechanisms through which cardiac sarcomeric mutations manifest in HCM are poorly understood. Hypothesis: We previously identified a novel MYH7 E848G mutation associated with HCM. We hypothesize E848G induces cell death that results in impaired tissue contractility in a dose-dependent manner. Methods: We created MYH7 expressing CMs with WT/WT, E848G/WT, or E848G/E848G alleles by CRISPR/Cas9 gene-editing patient-specific induced pluripotent stem cells (hiPSCs). hiPSC-derived cardiomyocytes were metabolically purified and cocultured with stromal cells on PDMS posts to create 3D engineered heart tissues (EHTs) or cultured as a monolayer. Results: Day 65 monolayer E848G/E848G CMs had 48.5% effective cell number relative to WT/WT. p53 (2.80 ± .11-fold), p21 (7.24 ± .18-fold), and BAX (1.64 ± 0.14-fold) mRNA transcripts were upregulated in day 60 monolayer E848G/E848G relative to WT/WT. E848G/E848G EHTs (n = 12) exhibited lower maximum active twitch force (104.6 ± 18.2 μN) and smaller 2D projected area (5.31 ± 0.22 mm 2 ) at day 14 relative to WT/WT (n = 15; 238.0 ± 20.4 μN; 6.87 ± 0.26 mm 2 ). E848G/WT EHTs (n = 7) had intermediate twitch force (168.7 ± 12.2 μN) and 2D area (6.18 ± 0.36 mm 2 ). Conclusion: These results suggest the MYH7 E848G mutation induces p53-associated cell death that leads to reduced tissue contractility. Ongoing studies will elucidate the molecular mechanism through which E848G activates cell death pathways. Figure: Representative EHTs. L-R: WT/WT, E848G/WT, E848G/E848G.

scholarly journals Facilitated Cross-Bridge Interactions with Thin Filaments by Familial Hypertrophic Cardiomyopathy Mutations inα-Tropomyosin

2011 ◽  
Vol 2011 ◽  
pp. 1-12 ◽  
Author(s):  
Fang Wang ◽  
Nicolas M. Brunet ◽  
Justin R. Grubich ◽  
Ewa A. Bienkiewicz ◽  
Thomas M. Asbury ◽  
...  
Keyword(s):  
Hypertrophic Cardiomyopathy ◽  
Molecular Mechanisms ◽  
Helical Structure ◽  
Familial Hypertrophic Cardiomyopathy ◽  
Maximum Speed ◽  
Sliding Speed ◽  
Thin Filaments ◽  
In Vitro Motility ◽  
Cardiac Sarcomeres

Familial hypertrophic cardiomyopathy (FHC) is a disease of cardiac sarcomeres. To identify molecular mechanisms underlying FHC pathology, functional and structural differences in three FHC-related mutations in recombinantα-Tm (V95A, D175N, and E180G) were characterized using both conventional and modified in vitro motility assays and circular dichroism spectroscopy. Mutant Tm's exhibited reducedα-helical structure and increased unordered structure. When thin filaments were fully occupied by regulatory proteins, little or no motion was detected at pCa 9, and maximum speed (pCa 5) was similar for all tropomyosins. Ca2+-responsiveness of filament sliding speed was increased either by increasedpCa50(V95A), reduced cooperativityn(D175N), or both (E180G). When temperature was increased, thin filaments with E180G exhibited dysregulation at temperatures ~10°C lower, and much closer to body temperature, than WT. When HMM density was reduced, thin filaments with D175N required fewer motors to initiate sliding or achieve maximum sliding speed.

scholarly journals LRRK2 mediates microglial neurotoxicity via NFATc2 in rodent models of synucleinopathies

2020 ◽  
Vol 12 (565) ◽  
pp. eaay0399
Author(s):  
Changyoun Kim ◽  
Alexandria Beilina ◽  
Nathan Smith ◽  
Yan Li ◽  
Minhyung Kim ◽  
...  
Keyword(s):  
Molecular Mechanisms ◽  
Nuclear Translocation ◽  
Critical Role ◽  
Lewy Bodies ◽  
Neuronal Loss ◽  
Activated T Cells ◽  
Behavioral Deficits ◽  
Downstream Signaling ◽  
Toll Like Receptor 2 ◽  
Factor Α

Synucleinopathies are neurodegenerative disorders characterized by abnormal α-synuclein deposition that include Parkinson’s disease, dementia with Lewy bodies, and multiple system atrophy. The pathology of these conditions also includes neuronal loss and neuroinflammation. Neuron-released α-synuclein has been shown to induce neurotoxic, proinflammatory microglial responses through Toll-like receptor 2, but the molecular mechanisms involved are poorly understood. Here, we show that leucine-rich repeat kinase 2 (LRRK2) plays a critical role in the activation of microglia by extracellular α-synuclein. Exposure to α-synuclein was found to enhance LRRK2 phosphorylation and activity in mouse primary microglia. Furthermore, genetic and pharmacological inhibition of LRRK2 markedly diminished α-synuclein–mediated microglial neurotoxicity via lowering of tumor necrosis factor–α and interleukin-6 expression in mouse cultures. We determined that LRRK2 promoted a neuroinflammatory cascade by selectively phosphorylating and inducing nuclear translocation of the immune transcription factor nuclear factor of activated T cells, cytoplasmic 2 (NFATc2). NFATc2 activation was seen in patients with synucleinopathies and in a mouse model of synucleinopathy, where administration of an LRRK2 pharmacological inhibitor restored motor behavioral deficits. Our results suggest that modulation of LRRK2 and its downstream signaling mediator NFATc2 might be therapeutic targets for treating synucleinopathies.

scholarly journals Modeling Parkinson’s Disease Using Induced Pluripotent Stem Cells

2020 ◽  
Vol 2020 ◽  
pp. 1-15
Author(s):  
Xinchao Hu ◽  
Chengyuan Mao ◽  
Liyuan Fan ◽  
Haiyang Luo ◽  
Zhengwei Hu ◽  
...  
Keyword(s):  
Parkinson’S Disease ◽  
Stem Cells ◽  
Parkinson's Disease ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Molecular Mechanisms ◽  
Cellular Level ◽  
Patient Specific ◽  
Ubiquitin Protein Ligase ◽  
Induced Pluripotent

Parkinson’s disease (PD) is the second most common neurodegenerative disease. The molecular mechanisms of PD at the cellular level involve oxidative stress, mitochondrial dysfunction, autophagy, axonal transport, and neuroinflammation. Induced pluripotent stem cells (iPSCs) with patient-specific genetic background are capable of directed differentiation into dopaminergic neurons. Cell models based on iPSCs are powerful tools for studying the molecular mechanisms of PD. The iPSCs used for PD studies were mainly from patients carrying mutations in synuclein alpha (SNCA), leucine-rich repeat kinase 2 (LRRK2), PTEN-induced putative kinase 1 (PINK1), parkin RBR E3 ubiquitin protein ligase (PARK2), cytoplasmic protein sorting 35 (VPS35), and variants in glucosidase beta acid (GBA). In this review, we summarized the advances in molecular mechanisms of Parkinson’s disease using iPSC models.

scholarly journals Antiphotoaging Effect of 3,5-Dicaffeoyl-epi-quinic Acid against UVA-Induced Skin Damage by Protecting Human Dermal Fibroblasts In Vitro

2020 ◽  
Vol 21 (20) ◽  
pp. 7756
Author(s):  
Jung Hwan Oh ◽  
Fatih Karadeniz ◽  
Chang-Suk Kong ◽  
Youngwan Seo
Keyword(s):  
Molecular Mechanisms ◽  
Type I Collagen ◽  
Nuclear Translocation ◽  
Quinic Acid ◽  
Dermal Fibroblasts ◽  
Type I ◽  
Human Dermal Fibroblasts ◽  
Skin Damage ◽  
Chronological Aging

Cutaneous aging is divided into intrinsic and exogenous aging correspondingly contributing to the complex biological phenomenon in skin. Intrinsic aging is also termed chronological aging, which is the accumulation of inevitable changes over time and is largely genetically determined. Superimposed on this intrinsic process, exogenous aging is associated with environmental exposure, mainly to ultraviolet (UV) radiation and more commonly termed as photoaging. UV-induced skin aging induces increased expression of matrix metalloproteinases (MMPs) which in turn causes the collagen degradation. Therefore, MMP inhibitors of natural origin are regarded as a primary approach to prevent or treat photoaging. This study investigated the effects of 3,5-dicaffeoyl-epi-quinic acid (DEQA) on photoaging and elucidated its molecular mechanisms in UVA-irradiated human dermal fibroblasts (HDFs). The results show that treatment with DEQA decreases MMP-1 production and increases type I collagen production in UVA-damaged HDFs. In addition, treatment of UVA-irradiated HDFs with DEQA downregulates MMP-1, MMP-3 and MMP-9 expression via blocking MAPK-cascade-regulated AP-1 transcriptional activity in UVA-irradiated HDFs. Furthermore, DEQA relieves the UVA-mediated suppression of type I procollagen and collagen expression through stimulating TGF-β/Smad signaling, leading to activation of the Smad 2/3 and Smad 4 nuclear translocation. These results suggest that DEQA could be a potential cosmetic agent for prevention and treatment of skin photoaging.

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