scholarly journals Stepwise, non-adherent differentiation of human pluripotent stem cells to generate basal forebrain cholinergic neurons via hedgehog signaling

2013 ◽  
Vol 11 (3) ◽  
pp. 1206-1221 ◽  
Author(s):  
Lucy A. Crompton ◽  
Meg L. Byrne ◽  
Hannah Taylor ◽  
Talitha L. Kerrigan ◽  
Gilles Bru-Mercier ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Basal Forebrain ◽  
Hedgehog Signaling ◽  
Cholinergic Neurons ◽  
Human Pluripotent Stem Cells ◽  
Basal Forebrain Cholinergic Neurons

scholarly journals A Simple Differentiation Protocol for Generation of Induced Pluripotent Stem Cell-derived Basal Forebrain Cholinergic Neurons for Alzheimer’s Disease and Frontotemporal Dementia Disease Modeling

2020 ◽  
Author(s):  
Sonia Sanz Muñoz ◽  
Martin Engel ◽  
Rachelle Balez ◽  
Dzung Do-Ha ◽  
Mauricio Castro Cabral-Da-Silva ◽  
...  
Keyword(s):  
Stem Cells ◽  
Growth Factors ◽  
Frontotemporal Dementia ◽  
Small Molecule ◽  
Pluripotent Stem Cells ◽  
Basal Forebrain ◽  
Cholinergic Neurons ◽  
Basal Forebrain Cholinergic Neurons ◽  
Differentiation Protocol ◽  
Induced Pluripotent

The study of neurodegenerative diseases using pluripotent stem cells requires new methods to assess neurodevelopment and neurodegeneration of specific neuronal subtypes. The cholinergic system, characterized by its use of the neurotransmitter acetylcholine, is one of the first to degenerate in Alzheimer’s disease and is also affected in frontotemporal dementia. We developed a differentiation protocol to generate basal forebrain cholinergic neurons (BFCNs) from induced pluripotent stem cells (iPSCs) aided by the use of small molecule inhibitors and growth factors. Ten iPSC lines were successfully differentiated into BFCNs using this protocol. The neuronal cultures were characterised through RNA and protein expression, and functional analysis of neurons was confirmed by whole-cell patch clamp. We have developed a reliable protocol using only small molecule inhibitors and growth factors, while avoiding transfection or cell sorting methods, to achieve a BFCN culture that expresses the characteristic markers of cholinergic neurons.

Gli3-mediated hedgehog inhibition in human pluripotent stem cells initiates and augments developmental programming of adult hematopoiesis

Blood ◽  
2013 ◽  
Vol 121 (9) ◽  
pp. 1543-1552 ◽  
Author(s):  
Brendan A. S. McIntyre ◽  
Veronica Ramos-Mejia ◽  
Shravanti Rampalli ◽  
Rami Mechael ◽  
Jong-Hee Lee ◽  
...  
Keyword(s):  
Stem Cells ◽  
Pluripotent Stem Cells ◽  
Hedgehog Signaling ◽  
Hematopoietic Cells ◽  
Developmental State ◽  
Developmental Programming ◽  
Human Pluripotent Stem Cells ◽  
Key Points

Key Points Transient inhibition of hedgehog signaling augments hematopoiesis in hPSC-derived EBs. Hedgehog inhibition initiates an advancement in the developmental state of hematopoietic cells derived from hPSCs.

scholarly journals Brainstem organoids from human pluripotent stem cells contain neural crest population

2019 ◽  
Author(s):  
Nobuyuki Eura ◽  
Takeshi K. Matsui ◽  
Joachim Luginbühl ◽  
Masaya Matsubayashi ◽  
Hitoki Nanaura ◽  
...  
Keyword(s):  
Stem Cells ◽  
Neural Crest ◽  
Pluripotent Stem Cells ◽  
Dopaminergic Neurons ◽  
Cholinergic Neurons ◽  
Human Pluripotent Stem Cells ◽  
Neural Crest Stem Cells ◽  
Human Brain Development ◽  
Cellular Population ◽  
The Brain

SummaryThe brainstem controls heartbeat, blood pressure and respiration, which are life-sustaining functions, therefore, disorders of the brainstem can be lethal. Brain organoids derived from human pluripotent stem cells recapitulate the course of human brain development and are expected to be useful for medical research on central nervous system disorders. However, existing organoid models have limitations, hampering the elucidation of diseases affecting specific components of the brain. Here, we developed a method to generate human brainstem organoids (hBSOs), containing neural crest stem cells as well as midbrain/hindbrain progenitors, noradrenergic and cholinergic neurons, and dopaminergic neurons, demonstrated by specific electrophysiological signatures. Single-cell RNA sequence analysis, together with proteomics and electrophysiology, revealed that the cellular population in these organoids was similar to that of the human brainstem and neural crest, which raises the possibility of making use of hBSOs in grafting for transplantation, efficient drug screenings and modeling the neural crest diseases.

scholarly journals A Simple Differentiation Protocol for Generation of Induced Pluripotent Stem Cell-Derived Basal Forebrain-Like Cholinergic Neurons for Alzheimer’s Disease and Frontotemporal Dementia Disease Modeling

Cells ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 2018 ◽  
Author(s):  
Sonia Sanz Muñoz ◽  
Martin Engel ◽  
Rachelle Balez ◽  
Dzung Do-Ha ◽  
Mauricio Castro Cabral-da-Silva ◽  
...  
Keyword(s):  
Stem Cells ◽  
Growth Factors ◽  
Frontotemporal Dementia ◽  
Small Molecule ◽  
Pluripotent Stem Cells ◽  
Basal Forebrain ◽  
Small Molecule Inhibitors ◽  
Cholinergic Neurons ◽  
Differentiation Protocol ◽  
Induced Pluripotent

The study of neurodegenerative diseases using pluripotent stem cells requires new methods to assess neurodevelopment and neurodegeneration of specific neuronal subtypes. The cholinergic system, characterized by its use of the neurotransmitter acetylcholine, is one of the first to degenerate in Alzheimer’s disease and is also affected in frontotemporal dementia. We developed a differentiation protocol to generate basal forebrain-like cholinergic neurons (BFCNs) from induced pluripotent stem cells (iPSCs) aided by the use of small molecule inhibitors and growth factors. Ten iPSC lines were successfully differentiated into BFCNs using this protocol. The neuronal cultures were characterised through RNA and protein expression, and functional analysis of neurons was confirmed by whole-cell patch clamp. We have developed a reliable protocol using only small molecule inhibitors and growth factors, while avoiding transfection or cell sorting methods, to achieve a BFCN culture that expresses the characteristic markers of cholinergic neurons.

scholarly journals The Controlled Generation of Functional Basal Forebrain Cholinergic Neurons from Human Embryonic Stem Cells

Stem Cells ◽  
2011 ◽  
Vol 29 (5) ◽  
pp. 802-811 ◽  
Author(s):  
Christopher J. Bissonnette ◽  
Ljuba Lyass ◽  
Bula J. Bhattacharyya ◽  
Abdelhak Belmadani ◽  
Richard J. Miller ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Basal Forebrain ◽  
Embryonic Stem ◽  
Cholinergic Neurons ◽  
Basal Forebrain Cholinergic Neurons

scholarly journals Ventral Telencephalic Patterning Protocols for Induced Pluripotent Stem Cells

2021 ◽  
Vol 9 ◽  
Author(s):  
Victor Krajka ◽  
Maximilian Naujock ◽  
Martje G. Pauly ◽  
Felix Stengel ◽  
Britta Meier ◽  
...  
Keyword(s):  
Stem Cells ◽  
Induced Pluripotent Stem Cells ◽  
Pluripotent Stem Cells ◽  
Basal Forebrain ◽  
Cholinergic Neurons ◽  
Neural Induction ◽  
Patient Specific ◽  
Electrophysiological Properties ◽  
Wide Range ◽  
Induced Pluripotent

The differentiation of human induced pluripotent stem cells (hiPSCs) into specific cell types for disease modeling and restorative therapies is a key research agenda and offers the possibility to obtain patient-specific cells of interest for a wide range of diseases. Basal forebrain cholinergic neurons (BFCNs) play a particular role in the pathophysiology of Alzheimer’s dementia and isolated dystonias. In this work, various directed differentiation protocols based on monolayer neural induction were tested for their effectiveness in promoting a ventral telencephalic phenotype and generating BFCN. Ventralizing factors [i.e., purmorphamine and Sonic hedgehog (SHH)] were applied at different time points, time intervals, and concentrations. In addition, caudal identity was prevented by the use of a small molecule XAV-939 that inhibits the Wnt-pathway. After patterning, gene expression profiles were analyzed by quantitative PCR (qPCR). Rostro-ventral patterning is most effective when initiated simultaneously with neural induction. The most promising combination of patterning factors was 0.5 μM of purmorphamine and 1 μM of XAV-939, which induces the highest expression of transcription factors specific for the medial ganglionic eminence, the source of GABAergic inter- and cholinergic neurons in the telencephalon. Upon maturation of cells, the immune phenotype, as well as electrophysiological properties were investigated showing the presence of marker proteins specific for BFCN (choline acetyltransferase, ISL1, p75, and NKX2.1) and GABAergic neurons. Moreover, a considerable fraction of measured cells displayed mature electrophysiological properties. Synaptic boutons containing the vesicular acetylcholine transporter (VACHT) could be observed in the vicinity of the cells. This work will help to generate basal forebrain interneurons from hiPSCs, providing a promising platform for modeling neurological diseases, such as Alzheimer’s disease or Dystonia.

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