Hepatic multilineage organoids derived from hypoimmunogenic iPSCs for liver regenerative medicine
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE313725 | GEO | 2026/03/26
REPOSITORIES: GEO
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Project description:Hepatic multilineage organoids derived from hypoimmunogenic iPSCs for liver regenerative medicine
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Project description:Induced pluripotent stem cells (iPSC) have huge potential as cell therapy for various diseases, given their potential for unlimited self-renewal and capability to differentiate into a wide range of cell types. Although autologous iPSCs represents the ideal source for patient-tailored regenerative medicine, the high costs of the extensive and time-consuming production process and the impracticability for treating acute conditions hinder their use for broad applications. An allogeneic iPSC-based strategy may overcome these issues, but it carries the risk of triggering an immune response. So far, several approaches based on genome-editing techniques to silence human leukocyte antigen class I (HLA-I) or II (HLA-II) expression have been explored to overcome the immune rejection of allogeneic iPSCs. In this study, we employed the CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR associated protein 9) system to delete the β2-Microglobulin (B2M) and the Class II Major Histocompatibility Complex Transactivator (CIITA) genes, essential for the correct surface expression of HLA-I and HLA-II proteins. The resulting hypoimmunogenic iPSC line has a normal karyotype, expresses the pluripotency stem cell markers, and is capable of differentiating into the three embryonic germ layers. Furthermore, we showed that it specifically retains the ability to differentiate towards different liver cells, such as endothelial-like cells, hepatocyte-like cells, and hepatic stellate-like cells. Our results indicate that hypoimmunogenic iPSCs could give a new cost-effective and off-the-shelf opportunity for cell therapy in liver diseases.
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Project description:Human pluripotent stem cell (hPSC)-derived cardiac therapies hold great promise for heart 41 regeneration but face major translational barriers due to allogeneic immune rejection. Here, we 42 engineered hypoimmunogenic hPSCs using a two-step CRISPR-Cas9 strategy: (1) B2M 43 knockout, eliminating HLA class I surface expression, and (2) knock-in of HLA-E or HLA-G trimer 44 constructs in the AAVS1 safe harbor locus to confer robust immune evasion. Hypoimmunogenic 45 hPSCs maintained pluripotency, efficiently differentiated into cardiac cell types that resisted both 46 T and NK cell-mediated cytotoxicity in vitro, and self-assembled into engineered cardiac 47 organoids. Comprehensive analyses of the hypoimmunogenic cells and organoids revealed 48 preservation of transcriptomic, structural, and functional properties with minimal off-target effects 49 from gene editing. In vivo, hypoimmunogenic cardiac organoids restored contractile function in 50 infarcted rat hearts and demonstrated superior graft retention and immune evasion in humanized 51 mice compared to wild-type counterparts. These findings establish the therapeutic potential of 52 hypoimmunogenic hPSC-CMs in the cardiac organoid platform, laying the foundation for off-the shelf cardiac cell therapies to treat cardiovascular disease, the leading cause of death worldwide.
2025-06-08 | GSE299172 | GEO
Project description:Human pluripotent stem cell (hPSC)-derived cardiac therapies hold great promise for heart 41 regeneration but face major translational barriers due to allogeneic immune rejection. Here, we 42 engineered hypoimmunogenic hPSCs using a two-step CRISPR-Cas9 strategy: (1) B2M 43 knockout, eliminating HLA class I surface expression, and (2) knock-in of HLA-E or HLA-G trimer 44 constructs in the AAVS1 safe harbor locus to confer robust immune evasion. Hypoimmunogenic 45 hPSCs maintained pluripotency, efficiently differentiated into cardiac cell types that resisted both 46 T and NK cell-mediated cytotoxicity in vitro, and self-assembled into engineered cardiac 47 organoids. Comprehensive analyses of the hypoimmunogenic cells and organoids revealed 48 preservation of transcriptomic, structural, and functional properties with minimal off-target effects 49 from gene editing. In vivo, hypoimmunogenic cardiac organoids restored contractile function in 50 infarcted rat hearts and demonstrated superior graft retention and immune evasion in humanized 51 mice compared to wild-type counterparts. These findings establish the therapeutic potential of 52 hypoimmunogenic hPSC-CMs in the cardiac organoid platform, laying the foundation for off-the shelf cardiac cell therapies to treat cardiovascular disease, the leading cause of death worldwide.
2025-06-08 | GSE299171 | GEO
Project description:Single cell sequencing of hypoimmunogenic neonatal islet beta cells
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Project description:Morphogen gradients critically regulate the coordinated emergence of endodermal and mesodermal lineages during liver bud formation, yet current human hepatic organoid models inadequately replicate these spatial cues. Here we demonstrate that precise modulation of Activin A concentrations within a dynamic 3D vertical-wheel bioreactor system enables the co-emergence and spatial organization of hepatic epithelium and septum transversum mesenchyme-like populations from human pluripotent stem cells. High Activin levels induce uniform definitive endoderm and enhanced hepatic maturation, whereas low levels promote epithelial tube–like structures encased by mesenchymal cells, recapitulating early liver diverticulum architecture. This gradient-based approach autonomously generates parenchymal and non-parenchymal hepatic lineages, providing a scalable platform to model human liver organogenesis and engineer ventral foregut–derived tissues.
2025-11-30 | GSE311252 | GEO
Project description:Hypoimmunogenic human pluripotent stem cells (hPSCs) are expected to serve as an unlimited cell source for generating universally compatible "off-the-shelf" cell grafts. However, whether the engineered hypoimmunogenic hPSCs still preserve their advantages of unlimited self-renewal and multilineage differentiation to yield functional tissue cells remains unclear. Here, we systematically studied the self-renewal and differentiation potency of three types of hypoimmunogenic hPSCs, established through the biallelic lesion of B2M gene to remove all surface expression of classical and nonclassical HLA class I molecules (B2Mnull), biallelic homologous recombination of nonclassical HLA-G1 to the B2M loci to knockout B2M while expressing membrane-bound β2m-HLA-G1 fusion proteins (B2MmHLAG), and ectopic expression of soluble and secreted β2m-HLA-G5 fusion proteins in B2MmHLAG hPSCs (B2Mm/sHLAG) in the most widely used WA09 human embryonic stem cells. Our results showed that hypoimmunogenic hPSCs with variable expression patterns of HLA molecules and immune compromising spectrums retained their normal self-renewal capacity and three-germ-layer differentiation potency. More importantly, as exemplified by neurons, cardiomyocytes and hepatocytes, hypoimmunogenic hPSC-derived tissue cells were fully functional as of their morphology, electrophysiological properties, macromolecule transportation and metabolic regulation. Our findings thus indicate that engineered hypoimmunogenic hPSCs hold great promise of serving as an unlimited universal cell source for cell therapeutics.
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Project description:This two-paper Series focuses on recent advances and applications of regenerative medicine that could benefit paediatric patients. Innovations in genomic, stem-cell, and tissue-based technologies have created progress in disease modelling and new therapies for congenital and incurable paediatric diseases. Prenatal approaches present unique opportunities associated with substantial biotechnical, medical, and ethical obstacles. Maternal plasma fetal DNA analysis is increasingly adopted as a noninvasive prenatal screening or diagnostic test for chromosomal and monogenic disorders. The molecular basis for cell-free DNA detection stimulated the development of circulating tumour DNA testing for adult cancers. In-utero stem-cell, gene, gene-modified cell (and to a lesser extent, tissue-based) therapies have shown early clinical promise in a wide range of paediatric disorders. Fetal cells for postnatal treatment and artificial placenta for ex-utero fetal therapies are new frontiers in this exciting field.
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Project description:Mesenchymal stem/stromal cells (MSC) are currently the best candidate therapeutic cells for regenerative medicine related to osteoarticular, muscular, vascular and inflammatory diseases, although these cells remain heterogeneous and necessitate a better biological characterization. We and others recently described that MSC originate from two types of perivascular cells, namely pericytes and adventitial cells and contain the in situ counterpart of MSC in developing and adult human organs, which can be prospectively purified using well defined cell surface markers. Pericytes encircle endothelial cells of capillaries and microvessels and express the adhesion molecule CD146 and the PDGFRβ, but lack endothelial and haematopoietic markers such as CD34, CD31, vWF (von Willebrand factor), the ligand for Ulex europaeus 1 (UEA1) and CD45 respectively. The proteoglycan NG2 is a pericyte marker exclusively associated with the arterial system. Besides its expression in smooth muscle cells, smooth muscle actin (αSMA) is also detected in subsets of pericytes. Adventitial cells surround the largest vessels and, opposite to pericytes, are not closely associated to endothelial cells. Adventitial cells express CD34 and lack αSMA and all endothelial and haematopoietic cell markers, as for pericytes. Altogether, pericytes and adventitial perivascular cells express in situ and in culture markers of MSC and display capacities to differentiate towards osteogenic, adipogenic and chondrogenic cell lineages. Importantly, adventitial cells can differentiate into pericyte-like cells under inductive conditions in vitro. Altogether, using purified perivascular cells instead of MSC may bring higher benefits to regenerative medicine, including the possibility, for the first time, to use these cells uncultured.
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Project description:The pancreas became one of the first objects of regenerative medicine, since other possibilities of dealing with the pancreatic endocrine insufficiency were clearly exhausted. The number of people living with diabetes mellitus is currently approaching half a billion, hence the crucial relevance of new methods to stimulate regeneration of the insulin-secreting β-cells of the islets of Langerhans. Natural restrictions on the islet regeneration are very tight; nevertheless, the islets are capable of physiological regeneration via β-cell self-replication, direct differentiation of multipotent progenitor cells and spontaneous α- to β- or δ- to β-cell conversion (trans-differentiation). The existing preclinical models of β-cell dysfunction or ablation (induced surgically, chemically or genetically) have significantly expanded our understanding of reparative regeneration of the islets and possible ways of its stimulation. The ultimate goal, sufficient level of functional activity of β-cells or their substitutes can be achieved by two prospective broad strategies: β-cell replacement and β-cell regeneration. The "regeneration" strategy aims to maintain a preserved population of β-cells through in situ exposure to biologically active substances that improve β-cell survival, replication and insulin secretion, or to evoke the intrinsic adaptive mechanisms triggering the spontaneous non-β- to β-cell conversion. The "replacement" strategy implies transplantation of β-cells (as non-disintegrated pancreatic material or isolated donor islets) or β-like cells obtained ex vivo from progenitors or mature somatic cells (for example, hepatocytes or α-cells) under the action of small-molecule inducers or by genetic modification. We believe that the huge volume of experimental and clinical studies will finally allow a safe and effective solution to a seemingly simple goal-restoration of the functionally active β-cells, the innermost hope of millions of people globally.
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