Allogeneic natural killer (NK) cell therapy has been effective in treating cancer. Many studies have tested NK cell therapy using human pluripotent stem cells (hPSCs). However, the impacts of the origin of PSC-NK cells on competence are unclear. In this study, several types of hPSCs, including human-induced PSCs (hiPSCs) generated from CD34+, CD3-CD56+, and CD56- cells in umbilical cord blood (UCB), three lines of human embryonic stem cells (hESCs, ES-1. ES-2 and ES-3) and MHC I knockout (B2M-KO)-ESCs were used to differentiate into NK cells and their capacities were analyzed. All PSC types could differentiate into NK cells. Among the iPSC-derived NK cells (iPSC-NKs) and ESC-derived NK cells (ES-NKs), 34+ iPSCs and ES-3 had a higher growth rate and cytotoxicity, respectively, ES-3 also showed better efficacy than 34+ iPSCs. B2M-KO was similar to the wild type. These results suggest that the screening for differentiation of PSCs into NK cells prior to selecting the PSC lines for the development of NK cell immunotherapy is an essential process for universal allotransplantation, including the chimeric antigen receptor (CAR).

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Human retinal organoids have become indispensable tools for retinal disease modeling and drug screening. Despite its versatile applications, the long timeframe for their differentiation and maturation limits the throughput of such research. Here, we successfully shortened this timeframe by accelerating human retinal organoid development using unique pharmacological approaches. Our method comprised three key steps: 1) a modified self-formed ectodermal autonomous multizone (SEAM) method, including dual SMAD inhibition and bone morphogenetic protein 4 treatment, for initial neural retinal induction; 2) the concurrent use of a Sonic hedgehog agonist SAG, activin A, and all-trans retinoic acid for rapid retinal cell specification; and 3) switching to SAG treatment alone for robust retinal maturation and lamination. The generated retinal organoids preserved typical morphological features of mature retinal organoids, including hair-like surface structures and well-organized outer layers. These features were substantiated by the spatial immunostaining patterns of several retinal cell markers, including rhodopsin and L/M opsin expression in the outermost layer, which was accompanied by reduced ectopic cone photoreceptor generation. Importantly, our method required only 90 days for retinal organoid maturation, which is approximately two-thirds the time necessary for other conventional methods. These results indicate that thoroughly optimized pharmacological interventions play a pivotal role in rapid and precise photoreceptor development during human retinal organoid differentiation and maturation. Thus, our present method may expedite human retinal organoid research, eventually contributing to the development of better treatment options for various degenerative retinal diseases.

The presence of residual undifferentiated pluripotent stem cells (PSCs) in PSC-derived cell therapy products (CTPs) is a major safety issue for their clinical application, due to the potential risk of PSC-derived tumor formation. An international multidisciplinary multisite study to evaluate a droplet digital PCR (ddPCR) approach to detect residual undifferentiated PSCs in PSC-derived CTPs was conducted as part of the Health and Environmental Sciences Institute Cell Therapy-TRAcking, Circulation & Safety Technical Committee. To evaluate the use of ddPCR in quantifying residual iPSCs in a cell sample, different quantities of induced pluripotent stem cells (iPSCs) were spiked into a background of iPSC-derived cardiomyocytes (CMs) to mimic different concentrations of residual iPSCs. A one step reverse transcription ddPCR (RT-ddPCR) was performed to measure mRNA levels of several iPSC-specific markers and to evaluate the assay performance (precision, sensitivity, and specificity) between and within laboratories. The RT-ddPCR assay variability was initially assessed by measuring the same RNA samples across all participating facilities. Subsequently, each facility independently conducted the entire process, incorporating the spiking step, to discern the parameters influencing potential variability. Our results show that a RT-ddPCR assay targeting ESRG, LINC00678, and LIN28A genes offers a highly sensitive and robust detection of impurities of iPSC-derived CMs and that the main contribution to variability between laboratories is the iPSC-spiking procedure, and not the RT-ddPCR. The RT-ddPCR assay would be generally applicable for tumorigenicity evaluation of PSC-derived CTPs with appropriate marker genes suitable for each CTP.

Despite improvements in patient outcomes, pediatric cancer remains a leading cause of non-accidental death in children. Recent genetic analysis of patients with pediatric cancers indicates an important role for both germline genetic predisposition and cancer-specific somatic driver mutations. Increasingly, evidence demonstrates that the developmental timepoint at which the cancer cell-of-origin transforms is critical to tumor identity and therapeutic response. Therefore, future therapeutic development would be bolstered by the use of disease models that faithfully recapitulate the genetic context, cell-of-origin, and developmental window of vulnerability in pediatric cancers. Human stem cells have the potential to incorporate all of these characteristics into a pediatric cancer model, while serving as a platform for rapid genetic and pharmacological testing. In this review, we describe how human stem cells have been used to model pediatric cancers and how these models compare to other pediatric cancer model modalities.
Today, pediatric cancer is a leading cause of non-accidental death in children. In order to further improve outcomes, it is important for researchers and clinicians alike to recognize how pediatric cancers are distinct from adult cancers. Inherited risk of cancer may play a greater role in pediatric cancer risk, and subsequent tumor-specific acquired driver mutations initiate tumor formation. However, there is substantial interaction between inherited and acquired mutations, which supports consideration of both simultaneously. Recent advancements in biotechnology, have improved matching between early cells of development and pediatric cancer cells, although cell-of-origin for certain pediatric central nervous system tumors remain elusive. Increasingly, evidence, particularly in pediatric medulloblastoma, demonstrates that the developmental timepoint at which the cancer cell-of-origin transforms is critical to tumor identity and therapeutic response. Therefore, future therapeutic development would be bolstered by the use of disease models that faithfully recapitulate the genetic context, cell-of-origin, and developmental window of pediatric cancers. Human stem cells have the potential to incorporate all of these characteristics into a pediatric cancer model, while serving as a platform for rapid genetic and pharmacological testing. In this review, we describe how human stem cells have been used to model pediatric cancers, how human these models compare to other pediatric cancer model modalities, and how these models can be improved in the future.

Mechanisms controlling trophoblast proliferation and differentiation during embryo implantation are poorly understood. Human trophoblast stem cells (TSC) and BMP4/A83-01/PD173074-treated pluripotent stem cell-derived trophoblast cells (BAP) are two widely employed, contemporary models to study trophoblast development and function, but how faithfully they mimic early trophoblast cells has not been fully examined. We evaluated the transcriptomes of trophoblast cells from BAP and TSC and directly compared them with those from peri-implantation human embryos during extended embryo culture (EEC) between embryonic day 8 to 12. The BAP and TSC grouped closely with trophoblast cells from EEC within each trophoblast sublineage following dimensional analysis and unsupervised hierarchical clustering. However, subtle differences in transcriptional programs existed within each trophoblast sublineage. We also validated the presence of six genes in peri-implantation human embryos by immunolocalization. Our analysis reveals that both BAP and TSC models have features of peri-implantation trophoblasts, while maintaining minor transcriptomic differences, and thus serve as valuable tools for studying implantation in lieu of human embryos.

Human embryonic stem cells and human induced pluripotent stem cells may be used to create 3D tissues called brain organoids. They duplicate the physiological and pathological characteristics of human brain tissue more faithfully in terms of both structure and function, and they more precisely resemble the morphology and cellular structure of the human embryonic brain. This makes them valuable models for both drug screening and in vitro studies on the development of the human brain and associated disorders. The technical breakthroughs enabled by brain organoids have a significant impact on the research of different brain regions, brain development and sickness, the connections between the brain and other tissues and organs, and brain evolution. This article discusses the development of brain organoids, their use in diabetes research, and their progress.

With the current environmental impact of large-scale animal production and societal concerns about the welfare of farm animals, researchers are questioning whether we can cultivate animal cells for the purpose of food production. This review focuses on a pivotal aspect of the cellular agriculture domain: cells. We summarised information on the various cell types from farm animals currently used for the development of cultured meat, including mesenchymal stromal cells, myoblasts, and pluripotent stem cells. The review delves into the advantages and limitations of each cell type and considers factors like the selection of the appropriate cell source, as well as cell culture conditions that influence cell performance. As current research in cultured meat seeks to create muscle fibers to mimic the texture and nutritional profile of meat, we focused on the myogenic differentiation capacity of the cells. The most commonly used cell type for this purpose are myoblasts or satellite cells, but given their limited proliferation capacity, efforts are underway to formulate myogenic differentiation protocols for mesenchymal stromal cells and pluripotent stem cells. The multipotent character of the latter cell types might enable the creation of other tissues found in meat, such as adipose and connective tissues. This review can help guiding the selection of a cell type or culture conditions in the context of cultured meat development.

Cell size is a crucial physical property that significantly impacts cellular physiology and function. However, the influence of cell size on stem cell specification remains largely unknown. Here, we investigated the dynamic changes in cell size during the differentiation of human pluripotent stem cells into definitive endoderm (DE). Interestingly, cell size exhibited a gradual decrease as DE differentiation progressed with higher stiffness. Furthermore, the application of hypertonic pressure or chemical to accelerate the reduction in cell size significantly and specifically enhanced DE differentiation. By functionally intervening in mechanosensitive elements, we have identified actomyosin activity as a crucial mediator of both DE differentiation and cell size reduction. Mechanistically, the reduction in cell size induces actomyosin-dependent angiomotin (AMOT) nuclear translocation, which suppresses Yes-associated protein (YAP) activity and thus facilitates DE differentiation. Together, our study has established a novel connection between cell size diminution and DE differentiation, which is mediated by AMOT nuclear translocation. Additionally, our findings suggest that the application of osmotic pressure can effectively promote human endodermal lineage differentiation.

Reactive astrocytes are known to exert detrimental effects upon neurons in several neurodegenerative diseases, yet our understanding of how astrocytes promote neurotoxicity remains incomplete, especially in human systems. In this study, we leveraged human pluripotent stem cell (hPSC) models to examine how reactivity alters astrocyte function and mediates neurodegeneration. hPSC-derived astrocytes were induced to a reactive phenotype, at which point they exhibited a hypertrophic profile and increased complement C3 expression. Functionally, reactive astrocytes displayed decreased intracellular calcium, elevated phagocytic capacity, and decreased contribution to the blood-brain barrier. Subsequently, co-culture of reactive astrocytes with a variety of neuronal cell types promoted morphological and functional alterations. Furthermore, when reactivity was induced in astrocytes from patient-specific hPSCs (glaucoma, Alzheimer\'s disease, and amyotrophic lateral sclerosis), the reactive state exacerbated astrocytic disease-associated phenotypes. These results demonstrate how reactive astrocytes modulate neurodegeneration, significantly contributing to our understanding of a role for reactive astrocytes in neurodegenerative diseases.