Dual function of hPSC-derived pericyte-like cells in vascularization and fibrosis-related cardiac tissue remodeling in vitro
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE146150 | GEO | 2020/11/30
REPOSITORIES: GEO
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Project description:Vascularization and maturation options for cardiac tissue engineered structures are currently intensively investigated. Therefore, the generation and characterisation of all cardiovascular cell types from human pluripotent stem cells (hPSC; either induced -iPSC- or embryonic -hESC) are of particular interest. In our group, differentiation and selection methods were described for obtaining highly pure hPSC-derived cardiomyocytes (CM; selected for αMHC), endothelial cells (EC; selected for CD31) and PDGFRβ expressing cardiac pericyte-like cells (PC). With the purpose of identifying cell type-related mechanisms in co-culture and tissue formation, gene expression profile of hPSC-derived CM, ECs, and PCs was compared to their undifferentiated progeny (hPSC) as well as to primary pericytes (hPC-PL) and fibroblasts (HFF).
2020-11-30 | GSE145957 | GEO
Project description:Rationale: Human pluripotent stem cells-derived cardiomyocytes (hPSC-CMs) exhibit the properties of fetal CMs, which limit their applications. Various methods have been used to promote maturation of hPSC-CMs; however, there is a lack of an unbiased and comprehensive method for accurate benchmarking of hPSC-CM maturation.
Objective: We aim to develop an unbiased proteomics method integrating high-throughput top-down targeted proteomics and bottom-up global proteomics for accurate and comprehensive assessment of hPSC-CM maturation.
Methods and Results: Utilizing hPSC-CMs from early- and late-stage two-dimensional monolayer culture and three-dimensional engineered cardiac tissue, we demonstrated high reproducibility and reliability of the top-down proteomics method, which enabled simultaneous quantification of contractile protein isoform expressions and their PTMs. This method allowed for the detection of known maturation-associated contractile protein alterations, and for the first time, identified contractile protein PTMs as promising new markers of maturation. By employing a global proteomics strategy, we identified candidate maturation markers important for sarcomere organization, cardiac excitability, and Ca2+ homeostasis; and validated these markers in the developing mouse cardiac ventricles.
Conclusions: We established an unbiased proteomics method that can provide accurate and specific benchmarking of hPSC-CM maturation, and identified new markers of maturation. Furthermore, this integrated proteomics strategy laid a strong foundation for uncovering molecular basis underlying cardiac development and disease using hPSC-CMs.
| MSV000082985 | MassIVE
Project description:In search for Matrigel substitutes, which is composed of proteins and glycans from mouse sarcoma, we disclose new dextran-derived functionalised hydrogels of different molecular weight suited as matrices in bioartificial cardiac tissue (BCT) cultivation. Moreover, the biopeptide RGD was coupled to the dextran polysaccharide backbone in order to modulate the bioactivity of the new matrices. Dextrans in combination with human collagen I, supported cardiac tissue formation with induced pluripotent stem cell-derived cardiomyocytes and fibroblasts. The strength and frequency of the formed heart muscles were monitored over time in a bioreactor. It was shown that covalently in situ cross linked dextrans are promising matrix systems for tissue formation in the field of regenerative medicine.
2021-03-22 | GSE147122 | GEO
Project description:Technological advancements have enabled the design of increasingly complex engineered tissue constructs, which better mimic native tissue cellularity. Therefore, dissecting the bi-directional interactions between distinct cell types in 3D is necessary to understand how heterotypic interactions at the single-cell level impact tissue-level properties. We systematically interrogated the interactions between cardiomyocytes (CMs) and cardiac non-myocytes in 3D self-assembled tissue constructs in an effort to determine the phenotypic and functional contributions of cardiac fibroblasts (CFs) and endothelial cells (ECs) to cardiac tissue properties. One week after tissue formation, cardiac microtissues containing CFs exhibited improved calcium handling function compared to microtissues comprised of CMs alone or CMs mixed with ECs, and CMs cultured with CFs exhibited distinct transcriptional profiles, with increased expression of cytoskeletal and ECM-associated genes. However, one month after tissue formation, functional and phenotypic differences between heterotypic tissues were mitigated, indicating diminishing impacts of non-myocytes on CM phenotype and function over time. The combination of single-cell RNA-sequencing and calcium imaging enabled the determination of reciprocal transcriptomic changes accompanying tissue-level functional properties in engineered heterotypic cardiac microtissues.
2020-09-21 | GSE155226 | GEO
Project description:Pluripotent stem cell-derived cardiomyocytes (PSC-CMs) provide an unprecedented opportunity to study human heart development and disease. A major caveat however is that they remain functionally and structurally immature in culture, limiting their potential for disease modeling and regenerative approaches. Here we address the question of how different metabolic pathways can be modulated in order to induce efficient cardiac maturation of hPSC-CMs. We show that PPAR signaling acts in an isoform-specific manner to balance the glycolysis and fatty acid oxidation (FAO) pathways. PPARd activation or inhibition results in efficient respective up- or down-regulation of the gene regulatory networks underlying FAO in hPSC-CMs. PPARd induction further increases mitochondrial and peroxisome content, enhances mitochondrial cristae formation and augments FAO flux. Lastly PPARd activation results in enhanced myofibril organization and increased numbers of bi-nucleated hPSC-CMs. Transient lactate exposure, commonly used in hPSC-CM purification protocols, induces an independent program of cardiac maturation, but when combined with PPARd activation equally results in a metabolic switch to FAO. In summary, we identify multiple axes of metabolic modifications of hPSC-CMs including a role for PPARdelta signaling in inducing the metabolic switch to FAO in hPSC-CMs. Our findings provide new opportunities to generate and use metabolically mature hPSC-CMs including for disease modeling and regenerative therapy.
2022-04-09 | GSE178984 | GEO
Project description:Pluripotent stem cell-derived cardiomyocytes (PSC-CMs) provide an unprecedented opportunity to study human heart development and disease. A major caveat however is that they remain functionally and structurally immature in culture, limiting their potential for disease modeling and regenerative approaches. Here we address the question of how different metabolic pathways can be modulated in order to induce efficient cardiac maturation of hPSC-CMs. We show that PPAR signaling acts in an isoform-specific manner to balance the glycolysis and fatty acid oxidation (FAO) pathways. PPARd activation or inhibition results in efficient respective up- or down-regulation of the gene regulatory networks underlying FAO in hPSC-CMs. PPARd induction further increases mitochondrial and peroxisome content, enhances mitochondrial cristae formation and augments FAO flux. Lastly PPARd activation results in enhanced myofibril organization and increased numbers of bi-nucleated hPSC-CMs. Transient lactate exposure, commonly used in hPSC-CM purification protocols, induces an independent program of cardiac maturation, but when combined with PPARd activation equally results in a metabolic switch to FAO. In summary, we identify multiple axes of metabolic modifications of hPSC-CMs including a role for PPARdelta signaling in inducing the metabolic switch to FAO in hPSC-CMs. Our findings provide new opportunities to generate and use metabolically mature hPSC-CMs including for disease modeling and regenerative therapy.
2021-11-01 | GSE160987 | GEO
Project description:Vascularization and efficient perfusion are long-standing challenges in cardiac tissue engineering. Here, we engineer perfusable microvascular constructs, wherein human embryonic stem cell-derived endothelial cells (hESC-ECs) are seeded both into patterned microchannels and the surrounding collagen matrix. In vitro, the hESC-ECs lining the luminal walls readily sprout and anastomose with de novo-formed endothelial tubes in the matrix under flow. When implanted on infarcted rat hearts, the perfusable microvessel grafts integrate with coronary vasculature to a greater degree than non-perfusable self-assembled constructs at 5 days post-implantation. Optical microangiography imaging reveal that perfusable grafts have 6-fold greater vascular density, 2.5-fold higher vascular velocities and >20-fold higher volumetric perfusion rates. Implantation of perfusable grafts containing additional hESC-derived cardiomyocytes show higher cardiomyocyte and vascular density. Thus, pre-patterned vascular networks enhance vascular remodeling and accelerate coronary perfusion, potentially supporting cardiac tissues after implantation. These findings should facilitate the next generation of cardiac tissue engineering design.
2018-12-24 | GSE124314 | GEO
Project description:Coronavirus disease 2019 (COVID-19) is a global pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 patients accompany with high frequencies of cardiac complications, which prominently contributed to the overall SARS-CoV-2-caused mortality. SARS-CoV-2 genome was reported to encode up to 14 genes. Currently, molecular mechanisms underlying SARS-CoV-2 viral gene-induced cardiac manifestations still remain elusive. Here, we overexpressed Orf9c, a SARS-CoV-2 encoded gene, in human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs). Multiple genome-wide analyses were performed to investigate the global impacts of Orf9c on hPSC-CMs. The mRNA-seq indicated stress-related signaling pathways, including cell death, immune and inflammation responses, activated by Orf9c. High-throughput proteomics and Co-immunoprecipitation mass spectrometry studies revealed the global influence of Orf9c on the proteome of hPSC-CMs and Orf9c-interactive protein network in hPSC-CMs. Orf9c overexpression elevated protein expressions of key factors essential for apoptosis while suppressing protein factors crucial for ATP synthesis. Further experimental validations confirmed that Orf9c overexpression induced prominent cell death, abnormal calcium handling and electrical properties, and significantly reduced cellular ATP level in hPSC-CMs. Finally, administration of two FDA-approved drugs, Ivermectin and Meclizine, were proven by our study to restore ATP level to prominently ameliorate Orf9c-induced cell death and electrical abnormalities of hPSC-CMs. Overall, we comprehensively manifest global responses of host human heart muscle cells to SARS-CoV-2 gene Orf9c, characterized molecular mechanisms underlying Orf9c-induced cardiac abnormalities, and explored potentially therapeutic approaches to ameliorate cardiac dysfunctions in COVID-19 patients.
2022-02-23 | GSE171370 | GEO
Project description:Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) show immense promise for patient-specific disease modeling, cardiotoxicity screening, and regenerative therapy development. However, hPSC-CMs in culture have not recapitulated the structural or functional properties of adult CMs in vivo thus far. To gain global insight into hPSC-CM biology, we established a multi-omics method for analyzing the hPSC-CM metabolome and proteome from the same cell culture, creating multi-dimensional profiles of hPSC-CMs. Specifically, we developed a sequential extraction to capture metabolites and proteins from the same hPSC-CM monolayer cultures, and analyzed these extracts using high-resolution mass spectrometry (MS).
2021-08-22 | MSV000088010 | MassIVE
Project description:Perlecan (HSPG2), a multifunctional heparan sulphate proteoglycan (HSPG), is a key player in extracellular matrix maturation and stabilisation. Structurally perlecan is similar to agrin, another HSPG known to contribute to cardiomyocyte phenotype switching by reverting hypertrophy. Although perlecan is crucial for cardiac development, its role in cardiomyocyte phenotypic switching is unknown. Here we show perlecan expression increases over time during the differentiation of pluripotent stem cells to cardiomyocytes (hPSC-CMs). Haploinusuffient hPSCs (HSPG2+/-) differentiate efficiently towards cardiomyocytes with minimal transcriptomic differences seen early in differentiation, but wide-ranging changes seen in late-stage cardiomyocytes. These differences are associated with structural, contractile, metabolic, and ECM genes. In keeping, late-stageHSPG2+/-hPSC-CMs display reduced maximal metabolic capacity and increased extracellular acidification rate, characteristics of reduced maturity. Moreover, engineered heart tissues produced withHSPG2+/-hPSC-CMs exhibit increased ECM remodelling, reduced tissue thickness and force generation. Wild-type hPSC-CMs grown on a substrate coated with a perlecan peptide showed increased cardiomyocyte nucleation typical of hypertrophic growth, suggesting that perlecan plays the opposite role of agrin by promoting cellular maturation rather than hyperplasia and proliferation. These data show that perlecan promotes cardiomyocyte maturity, possibly through hypertrophic growth. Targeting perlecan-dependent signalling may, therefore, reverse the phenotypic switch common to many forms of heart failure, by improving cardiomyocyte functionality.
2024-01-23 | GSE229975 | GEO