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Modelling Premature Cardiac Aging with Induced Pluripotent Stem Cells From a Hutchinson-Gilford Progeria Syndrome Patient

ABSTRACT: Hutchinson-Gilford Progeria Syndrome (HGPS) is a rare genetic disorder that causes accelerated aging and a high risk of cardiovascular complications. However, the underlying mechanisms of cardiac complications of this syndrome are not fully understood. This study modeled HGPS using cardiomyocytes (CM) derived from induced pluripotent stem cells (iPSC) derived from a patient with HGPS and characterized the biophysical, morphological, and molecular changes found in these CM compared to CM derived from a healthy donor. Electrophysiological recordings revealed that the HGPS CM was functional and had normal electrophysiological properties. Electron tomography showed nuclear morphology alteration and 3D reconstruction of electron tomography images indicated structural abnormalities in HGPS CM mitochondria. High-resolution respirometry with isolated CM derived from three distinct cellular differentiations suggests that HGPS-CM had a tendency of lower oxygen consumption capacity. However, there was no difference in mitochondrial content as measured by Mitotracker. Telomere length was measured using qRT-PCR, and no difference was found in either the iPSCs or the CM from HGPS when compared to the control. Proteomic analysis was carried out in a high-resolution system using Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS). The proteomics data show distinct group separations and protein expression differences between HGPS and control-CM, highlighting changes in ribosomal, TCA cycle, and amino acid biosynthesis, among other modifications. Our findings show that iPSC-derived cardiomyocytes from a Progeria Syndrome patient have significant changes in mitochondrial morphology and protein expression, implying novel mechanisms underlying premature cardiac aging

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Stem Cell, Cell Culture

DISEASE(S): Progeria

SUBMITTER: Gustavo Monnerat

LAB HEAD: Gustavo Monnerat

PROVIDER: PXD036557 | Pride | 2022-11-10

REPOSITORIES: Pride

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Project description:Calciprotein particles (CPPs), assembled as a result of molecular interactions between fetuin-A and nascent calcium phosphate clusters, are indispensable scavengers of excessive Ca2+ and PO43– ions, thus representing an elegant mechanism which regulates mineral homeostasis. Generation of CPPs represents an evolutionary mechanism to prevent blood supersaturation with Ca2+ and PO43– ions (e.g., as a result of bone resorption) and to thwart extraskeletal calcification, a pathological condition which is frequent in patients with chronic kidney disease. It is believed that formation of CPPs accompanied evolution from the appearance of bony fish; the existence of CPPs in more ancient animals remains uncertain. Shortly after emergence, amorphous, spherical, and submicrometer-sized (30-100 nm) primary CPPs (CPP-P) evolve into crystalline, spindle- or needle-shaped, and micrometer-sized (100-300 nm) secondary CPPs (CPP-S). Upon executing their function of aggregating Ca2+ and PO43– ions, CPPs are removed from the circulation by endothelial cells (ECs), monocytes, and liver or spleen macrophages. Internalisation and digestion of CPPs by ECs induce a chain of detrimental events including an increase in cytosolic Ca2+, mitochondrial and endoplasmic reticulum stress, nuclear factor (NF)-κB-mediated transcriptional response and cytokine release, ultimately contributing to the development of pro-inflammatory endothelial activation, chronic low-grade inflammation, and inflammaging. Pathological permeability of dysfunctional endothelium promotes lipid retention and leukocyte extravasation, together contributing to the development of vascular inflammation, proteolytic environment, degradation of basement membrane and internal elastic lamina, and contractile-to-synthetic switch of vascular smooth muscle cells. Such an ensemble of molecular events ultimately induces intimal hyperplasia, vascular remodeling, and atherosclerotic progression. Hence, CPPs act as a double-edged sword which rescues the human body from an acute life-threatening disease (i.e., extraskeletal calcification) at the cost of promoting long-lasting conditions (i.e., endothelial dysfunction and atherosclerosis). Among the several forms of calcium transfer (free Ca2+ ions, colloidal calciprotein monomers, and particulate CPPs), CPPs are the most efficient in delivering calcium stress to ECs because of their deposition and dissolution in lysosomes, followed by a lysosomal membrane permeabilisation and an excessive Ca2+ migration into the cytosol. The molecular pattern of CPP-induced calcium stress within the ECs is well defined and includes elevated expression of cell adhesion molecules (VCAM1, ICAM1, and E-selectin) and endothelial-to-mesenchymal transition transcription factors (SNAI1, SNAI2, TWIST1, and ZEB1) in combination with an endothelial nitric oxide synthase (eNOS) uncoupling. However, our knowledge on extracellular signatures of such endothelial response remains limited to an augmented release of pro-inflammatory cytokines (IL-6, IL-8, MCP-1/CCL2, MIF, CXCL1, and MIP-3α/CCL20) and pro- or anti-thrombotic molecules (serpin E1/PAI-1 and uPAR). As EC-secreted bioactive factors (termed angiokines) control vascular tone, maintain hemostasis, and regulate instructive signaling governing local homeostasis within the most organs, decryption and interpretation of endothelial secretome in physiological and pathological conditions is of utmost importance. Discovery of a circulating biomarkers specific for endothelial dysfunction, which can be measured by a routine enzyme-linked immunosorbent assay, might inform on specific disease states and prompt to the respective pharmacological interventions. Here, we aimed at deciphering the secretome of ECs treated with either CPP-P or CPP-S, employing ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS/MS) for the analysis of serum-free cell culture supernatant. For the objective comparison of heterogeneous arterial ECs, we utilised human coronary artery endothelial cells (HCAEC) and human internal thoracic artery endothelial cells (HITAEC) which comprise an innermost lining in atheroprone and atheroresistant blood vessels (coronary artery and internal thoracic artery, respectively).

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Modelling Premature Cardiac Aging with Induced Pluripotent Stem Cells From a Hutchinson-Gilford Progeria Syndrome Patient

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