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Driving efficient and pure skeletal muscle cell differentiation from pluripotent stem cells (PSCs) has been challenging. Here, we report an optimized protocol that generates skeletal muscle progenitor cells with high efficiency and purity in a short period of time. Human induced PSCs (hiPSCs) and murine embryonic stem cells (mESCs) were specified into the mesodermal myogenic fate using distinct and species-specific protocols. We used a specific maturation medium to promote the terminal differentiation of both human and mouse myoblast populations, and generated myotubes associated with a large pool of cell-cycle arrested PAX7 + cells. We also show that myotube maturation is modulated by dish-coating properties, cell density, and percentage of myogenic progenitor cells. Given the high efficiency in the generation of myogenic progenitors and differentiated myofibers, this protocol provides an attractive strategy for tissue engineering, modeling of muscle dystrophies, and evaluation of new therapeutic approaches in vitro., Competing Interests: Declaration of interests R.B.A., C.L., J.E.d.L., and F.R. declare no competing interests. D.A., J.M., and B.O. work or have worked for Phenocell., (Copyright © 2023. Published by Elsevier Inc.)

Waardenburg syndrome type 1 (WS1), a rare genetic disease characterized by pigmentation defects and mild craniofacial anomalies often associated with congenital deafness is caused by heterozygous mutations in the PAX3 gene (2q36.1). We have generated two induced pluripotent stem cell lines (PCli029-A and PCli031-A) from two patients from the same family both carrying the same heterozygous deletion in PAX3 exon 1 (c.-70_85 + 366del). These cells are pluripotent as they can differentiate into ectoderm, mesoderm and endoderm. They also can activate the early neural crest marker SNAI2. These cells will be useful for studying the human neural crest-derived pigment cells., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Brigitte Ontoniente reports a relationship with Phenocell SAS that includes: board membership and employment., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

In this study, we describe a simple and straightforward assay using induced pluripotent stem cell-derived melanocytes and high-throughput flow cytometry, to identify the effect induced by pigment regulating agents on melanin content. The assay is based on the correlation between forward light-scatter characteristics and melanin content, with pigmented cells displaying high light absorption/low forward light scatter, while the opposite is true for lowly pigmented melanocytes, as a result of genetic background or chemical treatments. Orthogonal validation is then performed by regular melanin quantification. Such approach was validated using a set of 80 small molecules and yielded a confirmed hit. The assay described in this study may prove a useful tool to identify modulators of melanogenesis in human melanocytes., (© 2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Despite advances in our understanding and research of induced pluripotent stem cells (iPSCs), their use in clinical practice is still limited due to lack of preclinical experiments. Neural precursors (NPs) derived from a clone of human iPSCs (IMR90) were used to treat a rat spinal cord lesion 1 week after induction. Functional recovery was evaluated using the BBB, beam walking, rotarod, and plantar tests. Lesion morphology, endogenous axonal sprouting, graft survival, and iPSC-NP differentiation were analyzed immunohistochemically. Quantitative polymerase chain reaction (qPCR) was used to evaluate the effect of transplanted iPSC-NPs on endogenous regenerative processes and also to monitor their behavior after transplantation. Human iPSC-NPs robustly survived in the lesion, migrated, and partially filled the lesion cavity during the entire period of observation. Transplanted animals displayed significant motor improvement already from the second week after the transplantation of iPSC-NPs. qPCR revealed the increased expression of human neurotrophins 8 weeks after transplantation. Simultaneously, the white and gray matter were spared in the host tissue. The grafted cells were immunohistochemically positive for doublecortin, MAP2, βIII-tubulin, GFAP, and CNPase 8 weeks after transplantation. Human iPSC-NPs further matured, and 17 weeks after transplantation differentiated toward interneurons, dopaminergic neurons, serotoninergic neurons, and ChAT-positive motoneurons. Human iPSC-NPs possess neurotrophic properties that are associated with significant early functional improvement and the sparing of spinal cord tissue. Their ability to differentiate into tissue-specific neurons leads to the long-term restoration of the lesioned tissue, making the cells a promising candidate for future cell-based therapy of SCI.

The fundamental inaccessibility of the human neural cell types affected by neurological disorders prevents their isolation for in vitro studies of disease mechanisms or for drug screening efforts. Pluripotent stem cells represent a new interesting way to generate models of human neurological disorders, explore the physiopathological mechanisms and develop new therapeutic strategies. Disease-specific human embryonic stem cells were the first source of material to be used to study certain disease states. The recent demonstration that human somatic cells, such as fibroblasts or blood cells, can be genetically converted to induced pluripotent stem cells (hiPSCs) together with the continuous improvement of methods to differentiate these cells into disease-affected neuronal subtypes opens new perspectives to model and understand a large number of human pathologies. This review focuses on the opportunities concerning the use disease-specific human pluripotent stem cells as well as the different challenges that still need to be overcome. We also discuss the recent improvements in the genetic manipulation of human pluripotent stem cells and the consequences of these on disease modeling and drug screening for neurological diseases.

The use of human pluripotent stem cells (PSCs) for cell therapy faces a number of challenges that are progressively answered by results from clinical trials and experimental research. Among these is the control of differentiation before transplantation and the prediction of cell fate after administration into the human brain, two aspects that condition both the safety and efficacy of the approach. For neurological disorders, this includes two steps: firstly, the identification of the optimal maturation stage for transplantation along the continuum that transforms PSCs into fully differentiated neural cell types, together with the derivation of robust protocols for large-scale production of biological products, and, secondly, the understanding of the effects of environmental cues and their possible interference with transplanted cells commitment. This review will firstly summarize our knowledge on developmental processes that have been applied to achieve robust in vitro differentiation of PSCs into neural progenitors. In a second part, we summarize results from experimental and clinical transplantation studies that help understanding the dialogue that establishes between transplanted cells and their host brain.

Human induced pluripotent stem cells (hiPSCs) are a most appealing source for cell replacement therapy in acute brain lesions. We evaluated the potential of hiPSC therapy in stroke by transplanting hiPSC-derived neural progenitor cells (NPCs) into the postischemic striatum. Grafts received host tyrosine hydroxylase-positive afferents and contained developing interneurons and homotopic GABAergic medium spiny neurons that, with time, sent axons to the host substantia nigra. Grafting reversed stroke-induced somatosensory and motor deficits. Grafting also protected the host substantia nigra from the atrophy that follows disruption of reciprocal striatonigral connections. Graft innervation by tyrosine hydoxylase fibers, substantia nigra protection, and somatosensory functional recovery were early events, temporally dissociated from the slow maturation of GABAergic neurons in the grafts and innervation of substantia nigra. This suggests that grafted hiPSC-NPCs initially exert trophic effects on host brain structures, which precede integration and potential pathway reconstruction. We believe that transplantation of NPCs derived from hiPSCs can provide useful interventions to limit the functional consequences of stroke through both neuroprotective effects and reconstruction of impaired pathways.

We tested the neuronal differentiation of human iPS-cells under in vitro conditions. For this purpose we pre-differentiated human (h) iPS-cells into neural stem cells and co-cultivated them with a cortical primary culture from embryonic rats. After 2 days of co-cultivation a certain number of hiPS-cells exhibited a clear neuronal morphology combined with expression of betaIII-tubulin and doublecortin. In addition, we found hiPS-cells without neuronal differentiation and cells already expressing betaIII-tubulin but not having yet distinctive axonal and dendritic processes. Human neuronal progenitors, starting neuronal differentiation, were contacted by both neuronal processes from rat neurons and oligodendrocytes, indicating a possible instructive influence by the primary culture on human cells. After 7 days of co-cultivation, however, we observed a complete degeneration of human iPS-derived cells and phagocytosis by microglial cells. Immunocytochemical stainings surprisingly revealed that microglial cells of the cortical primary culture express both CD8 and T-cell receptors.

With the development of stem cell (SC) biology, cell-based therapy has become a highly challenging field for experimental and clinical research. Among neurological disorders, stroke has pioneered the clinical application of SC. Safety concerns have prevailed for pilot clinical studies and important preclinical work is ongoing to help SC therapy reach the level of generalization. Stroke is classically divided into an acute, a subacute and a chronic phase. Each phase is defined by a complex array of events with overlapping and distinct kinetics that lead to both rapid tissue degeneration and long-lasting remodeling. Each SC type possesses intrinsic properties - transposed via cell-autonomous and non-cell-autonomous signaling - that would more specifically address some of these events. A better definition of what is expected from SC therapy in stroke might help assign SC sources to the acute or chronic phases and possibly optimize their use in the clinic., (Copyright © 2011 S. Karger AG, Basel.)

The purpose of the present study was to investigate the potential cardioprotective effects of an original approach based on the properties of the X chromosome-linked Inhibitor of Apoptosis (XIAP), the most effective endogenous inhibitor of apoptosis. For this purpose, the C-terminal part of XIAP (BIR3 and RING domains) was fused to the protein transduction domain (PTD) of the HIV1 transactivator of transcription, which confers to fused protein the ability to cross cell membranes. This protein, so-called PTD-BIR3/RING, was administered intravenously in C57BL/6J mice subjected to 30 min coronary artery occlusion and 24 h of reperfusion. Administration of PTD-BIR3/RING at 5 min before and 30 min after the onset of reperfusion reduced infarct size vs control (23+/-2% vs 41+/-4% and 27+/-4% vs 41+/-3%, respectively, p<0.05). Similar reduction in infarct size was observed when PTD-BIR3/RING was administered prior to ischemia (28+/-1% vs 44+/-3%). In addition to inhibition of caspase-3 and -9 activities, PTD-BIR3/RING induced an inhibition of caspase-8 and several other actors of the apoptotic pathways. In conclusion, this study demonstrates that the administration of PTD-BIR3/RING reduces myocardial infarct size even when injected during reperfusion through interruption of caspase activation by pharmacologically mimicking endogenous XIAP.