Facioscapulohumeral muscular dystrophy (FSHD) represents a major unmet clinical need arising from the progressive weakness and atrophy of skeletal muscles. The dearth of adequate experimental models has severely hampered our understanding of the disease. To date, no treatment is available for FSHD. Human embryonic stem cells (hESCs) potentially represent a renewable source of skeletal muscle cells (SkMCs) and provide an alternative to invasive patient biopsies. We developed a scalable monolayer system to differentiate hESCs into mature SkMCs within 26 days, without cell sorting or genetic manipulation. Here we show that SkMCs derived from FSHD1-affected hESC lines exclusively express the FSHD pathogenic marker double homeobox 4 and exhibit some of the defects reported in FSHD. FSHD1 myotubes are thinner when compared with unaffected and Becker muscular dystrophy myotubes, and differentially regulate genes involved in cell cycle control, oxidative stress response, and cell adhesion. This cellular model will be a powerful tool for studying FSHD and will ultimately assist in the development of effective treatments for muscular dystrophies. STEM CELLS TRANSLATIONAL MEDICINE 2016;5:1145-1161 SIGNIFICANCEThis work describes an efficient and highly scalable monolayer system to differentiate human pluripotent stem cells (hPSCs) into skeletal muscle cells (SkMCs) and demonstrates disease-specific phenotypes in SkMCs derived from both embryonic and induced hPSCs affected with facioscapulohumeral muscular dystrophy. This study represents the first human stem cell-based cellular model for a muscular dystrophy that is suitable for high-throughput screening and drug development.
Notch signaling is a key regulator of cell-fate decisions and is essential for proper neuroectodermal development. There, it favors the formation of ectoderm, promotes maintenance of neural stem cells, inhibits differentiation into neurons, and commits neural progenitors to a glial fate. In this report, we explore downstream effects of Notch important for astroglial differentiation. Transient activation of Notch1 during early stages of neuroectodermal differentiation of embryonic stem cells resulted in an increase of neural stem cells, a reduction in neurons, an induction of astroglial cell differentiation, and an induction of neural crest (NC) development. Transient or continuous activation of Notch1 during neuroectodermal differentiation led to upregulation of Sox9 expression. Knockdown of the Notch1-induced Sox9 expression reversed Notch1-induced astroglial cell differentiation, increase in neural stem cells, and the decrease in neurons, whereas the Notch1 effects on NC development were hardly affected by knockdown of Sox9 expression. These findings reveal a critical role for Notch-mediated upregulation of Sox9 in a select set of neural lineage determination steps controlled by Notch.
We are moving into a new era of stem cell research where many possibilities for treatment of degenerative, chronic and/or fatal diseases and injuries are becoming primed for clinical trial. These reports have led millions of people worldwide to hope that regenerative medicine is about to revolutionise biomedicine: either through transplantation of cells grown in the laboratory, or by finding ways to stimulate a patient's intrinsic stem cells to repair diseased and damaged organs. While major contributions of stem cells to drug discovery, safety and efficacy testing, as well as modelling ‘diseases in a dish’ are also expected, it is the in vivo use of stem cells that has captured the general public's attention. However, public misconceptions of stem cell potential and applications can leave patients vulnerable to the influences of profit driven entities selling unproven treatments without solid scientific basis or appropriate clinical testing or follow up. This review provides a brief history of stem cell clinical translation together with an overview of the properties, potential, and current clinical application of various stem cell types. In doing so it presents a clearer picture of the inherent risks and opportunities associated with stem cell research translation, and thus offers a framework to help realise invested expectations more quickly, safely and effectively.
The Notch signaling pathway controls cell fate choices at multiple steps during cell lineage progression. To produce the cell fate choice appropriate for a particular stage in the cell lineage, Notch signaling needs to interpret the cell context information for each stage and convert it into the appropriate cell fate instruction. The molecular basis for this temporal context-dependent Notch signaling output is poorly understood, and to study this, we have engineered a mouse embryonic stem (ES) cell line, in which short pulses of activated Notch can be produced at different stages of in vitro neural differentiation. Activation of Notch signaling for 6h specifically at day 3 during neural induction in the ES cells led to significantly enhanced cell proliferation, accompanied by Notch-mediated activation of cyclin D1 expression. A reduction of cyclin-D1-expressing cells in the developing CNS of Notch signaling-deficient mouse embryos was also observed. Expression of a dominant negative form of cyclin D1 in the ES cells abrogated the Notch-induced proliferative response, and, conversely, a constitutively active form of cyclin D1 mimicked the effect of Notch on cell proliferation. In conclusion, the data define a novel temporal context-dependent function of Notch and a critical role for cyclin D1 in the Notch-induced proliferation in ES cells.
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