Search Results (210)

Late-onset sporadic Alzheimer’s disease (LOAD) is the most common form of dementia. The disease is characterized by progressive loss of memory and behavioral changes followed by neurodegeneration of all cortical areas. While the contribution of genetic and environmental factors is important, advanced aging remains the most important disease risk factor. Because LOAD does not naturally occur in most animal species, except humans, studies have traditionally relied on the use of transgenic mouse models recapitulating early-onset familial Alzheimer’s disease (EOAD). Hence, the development of more representative LOAD models through reprograming of patient-derived cells into neuronal, glial, and immune cells became a necessity to better understand the disease’s origin and pathophysiology. Herein, and focusing on neurons, we review current work in the field and compare results obtained with two different reprograming methods to generate LOAD patient’s neuronal cells: the induced pluripotent stem cell and induced neuron technologies. We also evaluate if these models can faithfully mimic cellular and molecular pathologies observed in LOAD patients’ brains. Full article

Xeroderma Pigmentosum group C (XP-C) is an autosomal recessive disorder caused by mutations in the XPC gene, leading to defective nucleotide excision repair. This defect leads to genomic instability and a profound cancer predisposition. To model this disease, we generated induced pluripotent stem cells (iPSCs) from an XP-C patient carrying a novel homozygous nonsense mutation in the XPC gene (c.1830C>A). The resulting iPSCs demonstrated typical pluripotent characteristics, including expression of key markers and trilineage differentiation capability. However, genomic assessment revealed progressive karyotypic instability during extended culture. While initial whole-genome sequencing detected no major chromosomal abnormalities, subsequent G-banding analysis identified acquired trisomy 12 in two lines (CL12 and CL27) and a derivative X chromosome in a third line (CL30). These abnormalities were absent in early-passage analyses, indicating that they were acquired and selected for during extended culture. The acquisition of a derivative X chromosome in CL30, alongside recurrent trisomy 12, represents a unique cytogenetic signature likely attributable to the underlying XPC defect. We hypothesize that the loss of GG-NER creates a permissive genomic environment, accelerating the accumulation of DNA damage and chromosomal missegregation under replicative stress. This temporal divergence in genomic integrity highlights how culture pressures drive chromosomal evolution in XP-C iPSCs independently of initial reprogramming. Our findings emphasize that XP-C iPSCs require continuous genomic surveillance and provide a model for investigating how DNA repair deficiencies interact with in vitro culture stress. Full article

Background: Modeling precancerous stages holds the promise to understand early transformation events, thereby offering the potential for personalized, targeted treatment. Because cancer hijacks developmental pathways, precancerous stages could potentially be modeled by reprogramming cancer cells to an induced pluripotent stem cell state and subsequently differentiating them to the target organs using organoid models. Methods: We attempted reprogramming of patient-derived clear cell renal cell carcinoma (ccRCC) cell lines and adjacent normal renal epithelial cell lines using lentivirus or episomal reprogramming vectors. Results: The cancer cells failed to reprogram while the adjacent normal cells reprogrammed with high efficiency. The von Hippel–Lindau factor (VHL) gene was re-expressed in ccRCC cells in an attempt to restore the wild-type phenotype and restore reprogramming. The VHL gene is the major tumor suppressor in ccRCC pathogenesis and a conductor of oxidative-glycolytic glucose metabolism. While its re-expression did restore the epithelial phenotype and oxidative regulation of ccRCC cells, they still failed to stably reprogram. With an optimized reprogramming workflow, VHL-corrected ccRCC cells generate NANOG+ cells; however, they remained dependent on the ectopic expression of the reprogramming factors. Conclusions: We concluded that while VHL expression is necessary for cellular reprogramming of ccRCC cells, other genetic lesions in the ccRCC cells could be preventing the stabilization of the pluripotent state. Full article

The high prevalence and diversity of liver diseases present a significant problem for modern healthcare. Despite FDA approval of gene therapy drugs to treat hemophilia A and B, available treatment methods for other hereditary liver diseases are mainly limited to the frequently ineffective traditional therapies and surgical intervention. In recent years, significant progress has been made in the treatment of hepatitis C, but hepatitis B is still considered an incurable disease. In this regard, the treatment of hereditary and viral liver diseases using gene or cell therapy remains relevant. This review is focused on the current state of the induced pluripotent stem cells (iPSCs) field in the context of modeling and treatment of hereditary, viral, and some other liver diseases, both ex vivo and in vivo. Here we present a detailed discussion of the possible ways of modeling liver diseases ex vivo using iPSCs (reprogramming of patient somatic cells and genetic engineering (GE) of healthy iPSCs), summarize gene editing (GE) and non-GE approaches for the treatment of liver diseases, and demonstrate that iPSCs and their derivatives are widely used to treat liver diseases in vivo. Taken together, we are presenting a comprehensive analysis of 2D and 3D iPSC-based products in the context of liver diseases, discussing the advantages and disadvantages of this platform, including the comparison with other types of stem cells and animal models. This analysis may help understand not only the potential but also the limitations associated with the use of iPSCs in the context of various types of liver diseases. Full article

X chromosome inactivation (XCI) is a fundamental epigenetic process that balances X-linked gene expression between females and males by silencing one X chromosome in female cells. Variability or skewing of XCI can influence the clinical presentation of X-linked disorders. Bain type X-linked intellectual disability syndrome (MRXSB), caused by mutations in the X-linked HNRNPH2 gene, is characterized by intellectual disability, developmental delay, and neurological abnormalities. In female patients, XCI heterogeneity complicates disease modeling and therapeutic development. Induced pluripotent stem cells (iPSCs) offer a unique platform to study patient-specific disease mechanisms, but the dynamics of XCI during iPSC reprogramming, maintenance, and differentiation are not fully understood. In this study, we generated 12 iPSC clones from fibroblasts of a female MRXSB patient heterozygous for the HNRNPH2 c.340C > T mutation. Four clones expressed the mutant HNRNPH2 allele and eight expressed the wild-type allele, indicating X chromosome reactivation (XCR) followed by random XCI during reprogramming. Importantly, these XCI patterns remained stable during long-term iPSC propagation and subsequent differentiation into the three germ layers and neural stem cells. Our findings provide new insights into XCI and XCR dynamics in the context of X-linked neurodevelopmental disorders and emphasize the importance of careful clone selection for accurate disease modeling using iPSC-based approaches. Full article

iPSCs and their derivatives are used to investigate the molecular genetic mechanisms of human diseases, to identify therapeutic targets, and to screen for small molecules. Combining technologies for generating patient-specific iPSC lines and genome editing allows us to create cell models with unique characteristics. We obtained and characterized three iPSC lines by reprogramming peripheral blood mononuclear cells of a patient with Huntington’s disease (HD) using episomal vectors encoding Yamanaka factors. iPSC lines expressed pluripotency marker genes, had normal karyotypes and were capable of differentiating into all three germ layers. The obtained iPSC lines are useful for modeling disease progression in vitro and studying pathological mechanisms of HD, such as ER stress. A transgene of genetically encoded biosensor XBP1-TagRFP was introduced into the iPSCs to visualize ER stress state of cells. The study demonstrated that iPSC-derived medium spiny neurons develop ER stress, though the IRE1-mediated pathway does not seem to be involved in the process. Full article

The capability to generate induced pluripotent stem cells (iPSCs) from adult somatic cells, enabling them to differentiate into any cell type, has been demonstrated in several studies. In humans and mice, iPSCs have been shown to differentiate into primordial germ cells (PGCs), spermatozoa, and oocytes. However, research on iPSCs in deer is novel. Despite the necessity for establishing germplasm banks from endangered cervid species, the collection and cryopreservation of gametes and embryos have proven complex for this group. Therefore, the focus of this study was to establish protocols for deriving stable iPSC lines from Blastocerus dichotomus (Marsh deer) using primary cells derived from antler, adipose tissue, or skin, with the ultimate goal of producing viable gametes in the future. To achieve this, two main reprogramming approaches were tested: (1) transfection using PiggyBac transposons (plasmid PB-TET-MKOS) delivered via electroporation and (2) lentiviral transduction using the STEMCCA system with either human (hOSKM) or murine (mOSKM) reprogramming factors. Both systems utilized murine embryonic fibroblasts (MEFs) as feeder cells. The PiggyBac system was further supplemented with a culture medium containing small molecules to aid reprogramming, including a GSK inhibitor, MEK inhibitor, ALK/TGF inhibitor, and thiazovivin. Initial colony formation was observed; however, these colonies failed to expand post-selection. Despite these challenges, important insights were gained that will inform and guide future studies toward the successful generation of iPSCs in deer. Full article

As an interspecies hybrid inheriting genetic material from horse and donkey lineages, mules provide a unique model for studying allele-specific regulatory dynamics. Here, we isolated adult fibroblasts (AFs) and placental fibroblasts (PFs) from mule tissues and reprogrammed them into induced pluripotent stem cells (iPSCs). Intriguingly, placental fibroblast-derived iPSCs (mpiPSCs) exhibited reduced reprogramming efficiency compared to adult fibroblast-derived iPSCs (maiPSCs). Through allele-specific expression (ASE) analysis, we systematically dissected transcriptional biases in parental cell types and their reprogrammed counterparts, revealing conserved preferential expression of asinine alleles in core pluripotency regulators (e.g., POU5F1/OCT4, SOX2, NANOG) across both cell lineages. Strikingly, mpiPSCs displayed stronger asinine allele dominance than maiPSCs, suggesting tissue-specific parental genomic imprinting. Mechanistic exploration implicated PI3K-AKT signaling as a potential pathway mediating the reprogramming inefficiency in placental fibroblasts. By integrating transcriptomic profiling with ASE technology, this study uncovers allele selection hierarchies during somatic cell reprogramming in hybrids and establishes a framework for understanding how parental genomic conflicts shape pluripotency establishment. These findings advance interspecies iPSC research by delineating allele-specific regulatory networks and providing insights into the molecular constraints of hybrid cellular reprogramming. Full article

Aging, a state of progressive decline in physiological function, is an important risk factor for chronic diseases, ranging from cancer and musculoskeletal frailty to cardiovascular and neurodegenerative diseases. Understanding its cellular basis is critical for developing interventions to extend human health span. This review highlights the crucial role of in vitro models, discussing foundational discoveries like the Hayflick limit and the senescence-associated secretory phenotype (SASP), the utility of immortalized cell lines, and transformative human induced pluripotent stem cells (iPSCs) for aging and disease modeling and rejuvenation studies. We also examine methods to induce senescence and discuss the distinction between chronological time and biological clock, with examples of applying cells from progeroid syndromes and mitochondrial diseases to recapitulate some signaling mechanisms in aging. Although no in vitro model can perfectly recapitulate organismal aging, well-chosen models are invaluable for addressing specific mechanistic questions. We focus on experimental strategies to manipulate cellular aging: from “steering” cells toward resilience to “reversing” age-related phenotypes via senolytics, partial epigenetic reprogramming, and targeted modulation of proteostasis and mitochondrial health. This review ultimately underscores the value of in vitro systems for discovery and therapeutic testing while acknowledging the challenge of translating insights from cell studies into effective, organism-wide strategies to promote healthy aging. Full article

Genome instability in induced pluripotent stem cells (IPSC) poses a significant challenge for their use in research and medicine. Cataloging and precisely describing all the identified aberrations that arise during cell reprogramming, expansion, and differentiation is essential for improving approaches to instability prevention and ensuring genetic quality control. We report the karyotypic analysis of 65 cell lines derived from skin fibroblasts, urinal sediment, and peripheral blood mononuclear cells of 33 individuals, 82% of whom suffer from monogenic genetic disorders not associated with genetic instability. Trisomy of chromosomes 20 and 8 was revealed recurrently, while the 1q arm was the most frequently affected region involved in interstitial duplications and unbalanced translocations with chromosomes 15 and 18. The localization of rearrangement breakpoints identified by SNP arrays within the large DCC gene and histone gene clusters links genetic instability in IPSCs to replication-stress-induced chromosome breakage at common and early replicating fragile sites. Full article

Arteriovenous malformations (AVMs) are congenital vascular anomalies defined by abnormal direct connections between arteries and veins due to their complex structure or endovascular approaches. Pharmacological strategies targeting the underlying molecular mechanisms are thus gaining increasing attention in an effort to determine the mechanism involved in AVM regulation. In this study, we examined 30 human tissue samples, comprising 10 vascular samples, 10 human fibroblasts derived from AVM tissue, and 10 vascular samples derived from healthy individuals. The pharmacological agents thalidomide, U0126, and rapamycin were applied to the isolated endothelial cells (ECs). The pharmacological treatments reduced the proliferation of AVM ECs and downregulated miR-135b-5p, a biomarker associated with AVMs. The expression levels of angiogenesis-related genes, including VEGF, ANG2, FSTL1, and MARCKS, decreased; in comparison, CSPG4, a gene related to capillary networks, was upregulated. Following analysis of these findings, skin samples from 10 AVM patients were reprogrammed into induced pluripotent stem cells (iPSCs) to generate AVM blood vessel organoids. Treatment of these AVM blood vessel organoids with thalidomide, U0126, and rapamycin resulted in a reduction in the expression of the EC markers CD31 and α-SMA. The establishment of AVM blood vessel organoids offers a physiologically relevant in vitro model for disease characterization and drug screening. The authors of future studies should aim to refine this model using advanced techniques, such as microfluidic systems, to more efficiently replicate AVMs’ pathology and support the development of personalized therapies. Full article

Cell-based immunotherapy is a promising treatment strategy for cancer. Particularly in the case of solid tumors, however, this strategy only benefits a minority of patients. A critical limitation to immunotherapy is T cell exhaustion, a terminal differentiation state characterized by loss of self-renewal and cytotoxic capacity. For over a decade, regenerative immunology approaches to overcome exhaustion and restore stem-like features of T cells have been pursued. The reprogramming of tumor-specific T cells back to a less-differentiated, stem-like state using induced pluripotent stem cell (iPSC) technology has been viewed as a powerful and highly appealing strategy to overcome the limitations imposed by exhaustion. However, clinical translation of these approaches has been stymied by the requirement for subsequent iPSC-to-T cell re-maturation strategies, vanishingly low efficiencies, and resource-intensive cell culture protocols. In this review, we discuss the emergence of transcription factor reprogramming to iPSCs, contemporary techniques for T cell reprogramming, as well as techniques for re-differentiation into mature T cells. We discuss the potential clinical utility of T cell reprogramming and re-maturation strategies alongside progress and major roadblocks toward clinical translation. If these challenges can be addressed, transcription factor reprogramming of T cells into iPSCs and subsequent re-maturation into tumor-specific stem-like T cells may represent an incredibly efficacious approach to cancer immunotherapy. Full article

Background: Pulmonary fibrosis is a major disease that leads to the progressive loss of lung function. The disease manifests early, resulting in type 2 respiratory failure. This is likely due to the bronchocentric fibrosis around the major airways, which causes airflow limitation. It affects approximately three million patients worldwide and has a poor prognosis. Skin fibroblasts isolated from patients offer valuable insights into understanding the disease mechanisms, identifying the genetic causes, and developing personalized therapies. However, the use of skin fibroblasts to study a disease that exclusively impacts the lungs is often questioned, particularly since lung fibrosis primarily affects the alveolar epithelium. Method: We report the reprogramming of skin fibroblasts from patients with an atypical early-onset form of lung fibrosis into induced pluripotent stem cells (iPSCs) and subsequently into alveolar epithelial cells. This was achieved using a Sendai virus approach. Results: We show that the reprogrammed cells carry mutations in the calcium-binding protein genes S100A3 and S100A13, leading to diminished protein expression, thus mimicking the patients’ cells. Additionally, we demonstrate that the generated patient iPSCs exhibit aberrant calcium and mitochondrial functions. Conclusions: Due to the lack of a suitable animal model that accurately resembles the human disease, generating patient lung cells from these iPSCs can provide a valuable “disease in a dish” model for studying the atypical form of inherited lung fibrosis. This condition is associated with mutations in the calcium-binding protein genes S100A3 (NM_002960) and S100A13 (NM_001024210), aiding in the understanding of its pathogenesis. Full article

Cardiomyopathies are a heterogeneous group of heart muscle diseases that can lead to heart failure, arrhythmias, and sudden cardiac death. Traditional animal models and in vitro systems have limitations in replicating the complex pathology of human cardiomyopathies. Induced pluripotent stem cells (iPSCs) offer a transformative platform by enabling the generation of patient-specific cardiomyocytes, thus opening new avenues for disease modeling, drug discovery, and regenerative therapy. This process involves reprogramming somatic cells into iPSCs and subsequently differentiating them into functional cardiomyocytes, which can be characterized using techniques such as electrophysiology, contractility assays, and gene expression profiling. iPSC-derived cardiomyocyte (iPSC-CM) platforms are also being explored for drug screening and personalized medicine, including high-throughput testing for cardiotoxicity and the identification of patient-tailored therapies. While iPSC-CMs already serve as valuable models for understanding disease mechanisms and screening drugs, ongoing advances in maturation and bioengineering are bringing iPSC-based therapies closer to clinical application. Furthermore, the integration of multi-omics approaches and artificial intelligence (AI) is enhancing the predictive power of iPSC models. iPSC-based technologies are paving the way for a new era of personalized cardiology, with the potential to revolutionize the management of cardiomyopathies through patient-specific insights and regenerative strategies. Full article

The European wisent (Bison bonasus), an iconic yet genetically vulnerable species, faces ongoing conservation challenges due to a restricted gene pool. Advances in induced pluripotent stem cell (iPSC) technology offer promising prospects for preserving and restoring genetic diversity in endangered species. In this study, we sought to reprogram wisent somatic cells into iPSCs using the PiggyBac transposon system, a non-viral method known for being successfully applied in bovine species. We applied a six-factor reprogramming cocktail (OCT4, SOX2, KLF4, LIN28, c-MYC, NANOG) alongside small-molecule enhancers to fibroblasts isolated from adult wisent tissue. While initial colony formation was observed, the reprogrammed cells exhibited limited proliferation and failed to maintain stable pluripotency, suggesting intrinsic barriers to complete reprogramming. Despite optimizing culture conditions, including hypoxia and extracellular matrix modifications, the reprogramming efficiency remained low. Our findings indicate that wisent somatic cells may require alternative reprogramming strategies, such as new-generation delivery systems and epigenetic modulators, to achieve stable iPSC lines. This study underscores the need for species-specific optimization of reprogramming protocols and highlights the potential of emerging cellular technologies for conservation efforts. Future research integrating advanced reprogramming tools may pave the way for genetic rescue strategies in wisent and other endangered species. Full article