Biology subjects

Hamledari, H.

Publications and source records attributed to Hamledari, H..

3 recordsLinked to original sources

bioRxiv2025

Molecular rewiring of human induced pluripotent stem cell-derived cardiomyocytes during metabolic maturation

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are widely used to model cardiac development and inherited cardiomyopathies, yet their immature metabolic state limits interpretation of disease-associated molecular programs. While multiple strategies promote structural and functional maturation, less is known about the molecular regulation of metabolic maturation as a distinct developmental transition. Here, we examine how metabolic maturation reshapes molecular and metabolic states in wild-type and TNNT2-linked hypertrophic cardiomyopathy (HCM) hiPSC-CMs. Using integrated transcriptomic, chromatin accessibility, proteomic, and metabolic profiling, we define the molecular trajectory associated with metabolic maturation in wild-type hiPSC-CMs, characterized by coordinated transcriptional and epigenetic remodeling, enhanced mitochondrial oxidative metabolism, and progressive suppression of mTORC1 signaling. In contrast, hiPSC-CMs carrying TNNT2 HCM variants (I79N+/- and R278C+/-) exhibit variant-specific deviations from this metabolic maturation trajectory. While early CM differentiation is largely preserved, metabolic maturation reveals defects in mitochondrial respiration and chromatin organization, with the more clinically severe I79N+/- variant showing sustained metabolic impairment. We further find that mTORC1 activity is temporally misregulated during metabolic maturation in HCM hiPSC-CMs, with reduced signaling at early stages and normalization at later time points. Pharmacological inhibition of mTORC1 with rapamycin partially improves disease-associated protein expression signatures in the I79N+/- variant, particularly when applied during early differentiation. Together, these findings demonstrate that TNNT2-linked HCM involves disrupted metabolic maturation programs coupled to altered gene regulatory states, highlighting metabolic maturation as a critical context for studying cardiomyopathy-associated molecular phenotypes in hiPSC-CMs.

molecular biology↗

bioRxiv2023

Molecular and metabolomic characterization of hiPSC-derived cardiac fibroblasts transitioning to myofibroblasts

1.Mechanical stress and pathological signaling trigger the activation of fibroblasts to myofibroblasts, which impacts extracellular matrixcomposition, disrupts normal wound healing,andcan generate deleterious fibrosis (Bohl et al., 2008; Sutton and Sharpe, 2000). Myocardial fibrosis independently promotes cardiac arrhythmias, sudden cardiac arrest, and contributes to the severity of heart failure (Frangogiannis, 2021). Fibrosis can also alter cell-to-cell communication and increase myocardial stiffness which eventually may lead to lusitropic and inotropic cardiac dysfunction (PMID: 33135058). Human induced pluripotent stem cell derived cardiac fibroblasts (hiPSC-CFs) have the potential to enhance clinical relevance in precision disease modeling by facilitating the study of patient-specific phenotypes. However, it is unclear whether hiPSC-CFs can be activated to become myofibroblasts akin to primary cells, and the key signaling mechanisms in this process remain unidentified. We hypothesize that the passaging of hiPSC-CFs, like primary cardiac fibroblasts, induces specific genes required for myofibroblast activation and increased mitochondrial metabolism. Passaging of hiPSC-CFs from passage 0 to 3 (P0 to P3) and treatment of P0 with TGF{beta}1 was associated with a gradual induction of genes to initiate the activation of these cells to myofibroblasts, including collagen, periostin, fibronectin, and collagen fiber processing enzymes with concomitant downregulation of cellular proliferation markers. Most importantly, canonical TGF{beta}1 and Hippo signaling component genes including TAZ were influenced by passaging hiPSC-CFs. Seahorse assay revealed that passaging and TGF{beta}1 treatment increased mitochondrial respiration, consistent with fibroblast activation requiring increased energy production, whereas treatment with the glutaminolysis inhibitor BPTES completely attenuated this process. Based on these data, the hiPSC-CF passaging enhanced fibroblast activation, activated fibrotic signaling pathways, and enhanced mitochondrial metabolism approximating what has been reported in primary cardiac fibroblasts. Thus, hiPSC-CFs may provide an accurate in vitro preclinical model for the cardiac fibrotic condition, which may facilitate the identification of putative anti-fibrotic therapies, including patient-specific approaches. HighlightsO_LIPassaging promotes the activation of fibroblasts to myofibroblasts. C_LIO_LITGF{beta}1 treatment activates the fibroblasts, but their expression profile was uniquely different from myofibroblasts. C_LIO_LIHigh energy requiring fibroblast activation is dependent on glutaminase-based mitochondrial metabolism. C_LIO_LIPassaging induces TGF{beta}1 and Hippo signaling pathways in activated fibroblasts and myofibroblasts. C_LI Graphical Abstract CaptionProbing the activation of fibroblasts to myofibroblasts is key in ECM remodeling processes to avoid fibrosis-related adverse complications, and to better understand disease pathology. Here we report that passaging of hiPSC-derived cardiac fibroblasts promotes fibroblast activation along with a gradual shift in gene expression and metabolic changes towards myofibroblasts. TGF{beta}1 treatment activates non-passaged fibroblasts, but they are dissimilar to myofibroblasts. The energy-intensive fibroblast to myofibroblast activation process is dependent on glutaminase-mediated mitochondrial metabolism and is prevented by treatment with GLS-1 inhibitor BPTES. Our work demonstrates that hiPSC-CFs can offer a preclinical model analogous to primary cardiac fibroblasts that is comparable with passage-mediated myofibroblast activation and increased mitochondrial metabolism. hiPSC-CFs may also facilitate patient-specific novel anti-fibrosis drug screening and disease management. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=154 SRC="FIGDIR/small/561455v2_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@139f3a9org.highwire.dtl.DTLVardef@1abea35org.highwire.dtl.DTLVardef@19d7bfborg.highwire.dtl.DTLVardef@369e9e_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2023

Mechanisms of Pathogenicity of Hypertrophic Cardiomyopathy-Associated Troponin T (TNNT2) Variant R278C+/- During Development

Hypertrophic cardiomyopathy (HCM) is one of the most common heritable cardiovascular diseases and variants of TNNT2 (cardiac troponin T) are linked to increased risk of sudden cardiac arrest despite causing limited hypertrophy. In this study, a TNNT2 variant, R278C+/-, was generated in both human cardiac recombinant/reconstituted thin filaments (hcRTF) and human-induced pluripotent stem cells (hiPSCs) to investigate the mechanisms by which the R278C+/- variant affects cardiomyocytes at the proteomic and functional levels. The results of proteomics analysis showed a significant upregulation of markers of cardiac hypertrophy and remodeling in R278C+/- vs. the isogenic control. Functional measurements showed that R278C+/- variant enhances the myofilament sensitivity to Ca2+, increases the kinetics of contraction, and causes arrhythmia at frequencies >75 bpm. This study uniquely shows the profound impact of the TNNT2 R278C+/- variant on the cardiomyocyte proteomic profile, cardiac electrical and contractile function in the early stages of cardiac development. Translational PerspectiveHypertrophic cardiomyopathy (HCM) is the leading known cause of sudden cardiac arrest in the young. Thin-variant associated HCM variants make up to 15% of familial HCM yet their molecular mechanisms remain less clear relative to thick filament variants. Here, we employ computational modeling, human cardiac recombinant/reconstituted thin filaments (hcRTF), and hiPSC-CMs to study the thin filament TNNT2 R278C+/- variant, revealing its extensive pathogenicity and potential mechanisms by which it can lead to HCM and sudden death. Mavacamten, the recently FDA-approved treatment, was effective in alleviating contractile dysfunction in TNNT2 R278C+/- hiPSC-CMs, positing it as a potential therapy for thin filament HCM. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=117 HEIGHT=200 SRC="FIGDIR/small/542948v2_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@102a8b0org.highwire.dtl.DTLVardef@190e356org.highwire.dtl.DTLVardef@13d3f9aorg.highwire.dtl.DTLVardef@1a9c33_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗