Efficient Generation of Trunk Neural Crest and Sympathetic Neurons from Human Pluripotent Stem Cells Via a Neuromesodermal Axial Progenitor Intermediate

Frith, Thomas J.R.; Tsakiridis, Anestis

doi:10.1002/cpsc.81

Cited by 35 publications

(37 citation statements)

References 18 publications

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“…In another key study, Muotri and colleagues reported a protocol where cortical organoids were induced with electrophysiologically active neurons with glutamatergic and GABAergic signaling ( Trujillo et al, 2018 ). We designed our protocol based on these existing reports and made use of selected hormones, such as human insulin, transferrin, and progesterone, which have been found to be protective or support the survival and differentiation of dopaminergic neurons and neural crest lineage cells ( Ciarlo et al, 2017 ; Frith and Tsakiridis, 2019 ). Of note, our findings suggest that the use of an orbit shaker might have contributed to quick induction of hBSOs, through the achievement of an ideal concentration gradient of growth factors.…”

Section: Discussionmentioning

confidence: 99%

Brainstem Organoids From Human Pluripotent Stem Cells

et al. 2020

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The brainstem is a posterior region of the brain, composed of three parts, midbrain, pons, and medulla oblongata. It is critical in controlling heartbeat, blood pressure, and respiration, all of which are life-sustaining functions, and therefore, damages to or disorders of the brainstem can be lethal. Brain organoids derived from human pluripotent stem cells (hPSCs) 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 are limited in the extent hPSCs recapitulate human brain development and hence are not able to fully elucidate the diseases affecting various components of the brain such as brainstem. Here, we developed a method to generate human brainstem organoids (hBSOs), containing midbrain/hindbrain progenitors, noradrenergic and cholinergic neurons, dopaminergic neurons, and neural crest lineage cells. Single-cell RNA sequence (scRNA-seq) analysis, together with evidence from proteomics and electrophysiology, revealed that the cellular population in these organoids was similar to that of the human brainstem, which raises the possibility of making use of hBSOs in investigating central nervous system disorders affecting brainstem and in efficient drug screenings.

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Section: Discussionmentioning

confidence: 99%

Brainstem Organoids From Human Pluripotent Stem Cells

et al. 2020

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“…In another key study, Muotri and colleagues reported a protocol where cortical organoids were induced with electrophysiologically-active neurons with glutamatergic and GABAergic signaling (Trujillo et al, 2018). We designed our protocol based on these existing reports, and made use of selected hormones, such as human insulin, transferrin, and progesterone, that have been found to support the survival and differentiation of dopaminergic neurons and neural crest lineage cells (Ciarlo et al, 2017;Frith and Tsakiridis, 2019). Of note, our findings suggest that the use of an orbit shaker might have contributed to quick induction of hBSOs, through the achievement of an ideal concentration gradient of growth factors.…”

Section: Discussionmentioning

confidence: 99%

Brainstem organoids from human pluripotent stem cells contain neural crest population

Eura

Matsui

Luginbühl

et al. 2019

Preprint

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The 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. IntroductionThe brainstem is a posterior region of the brain between the deep structures of the cerebral hemispheres. It connects the cerebrum with the spinal cord and is divided into three parts: midbrain, pons and medulla oblongata. They contain multiple nuclei and small fiber tracts widely projecting to the cerebrum cortex, basal ganglia and other parts of the cerebrum. Brainstem functions such as alertness, heartbeat, blood pressure and respiration are considered to be more vital for life than that of the cortex. Therefore, damages to or disorders of brainstem such as infarction, hemorrhage, tumors, or any neurodegenerative diseases may lead to death.Of those, Parkinson's disease (PD) is the most well-known degenerative disease, showing progressive voluntary movement impairments, including rigidity, akinesia, tremor, and postural instability. These symptoms are caused by the loss of dopaminergic neurons at midbrain substantia nigra. In addition, non-motor symptoms, such as autonomic dysfunction, sleep disorder, and depression in PD patients, are thought to be derived from impairments of the serotonergic or noradrenergic system in the brainstem. However, the mechanisms driving these symptoms have yet to be determined. Hence, we need the development of models that can recapitulate the human midbrain and surrounding brainstem to elucidate the process of neural degeneration in this area.Recent progress on protocols for inducing organs in-a-dish (organoids) provides potentials for the modeling of various diseases (Clevers, 2016). Organoids mimic the structure of organs Eura, Matsui, Luginbühl et al.

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“…Differentiation of NCC was carried out according to previously described procedures with minor modifications [27]. hPSCs were plated at 4 × 10 4 cells/cm 2 on plates coated with vitronectin in Essential 6 medium supplemented with CHIR99021 (1.4 μM), FGF2 (100 ng/ml), BMP4 (20 ng/ml), and SB431542 (10 μM, Peprotech).…”

Section: Differentiationmentioning

confidence: 99%

The Rho-associated kinase inhibitor fasudil can replace Y-27632 for use in human pluripotent stem cell research

Lee

Choi

et al. 2020

PLoS ONE

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Poor survival of human pluripotent stem cells (hPSCs) following freezing, thawing, or passaging hinders the maintenance and differentiation of stem cells. Rho-associated kinases (ROCKs) play a crucial role in hPSC survival. To date, a typical ROCK inhibitor, Y-27632, has been the primary agent used in hPSC research. Here, we report that another ROCK inhibitor, fasudil, can be used as an alternative and is cheaper than Y-27632. It increased hPSC growth following thawing and passaging, like Y-27632, and did not affect pluripotency, differentiation ability, and chromosome integrity. Furthermore, fasudil promoted retinal pigment epithelium (RPE) differentiation and the survival of neural crest cells (NCCs) during differentiation. It was also useful for single-cell passaging of hPSCs and during aggregation. These findings suggest that fasudil can replace Y-27632 for use in stem research.

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Efficient Generation of Trunk Neural Crest and Sympathetic Neurons from Human Pluripotent Stem Cells Via a Neuromesodermal Axial Progenitor Intermediate

Cited by 35 publications

References 18 publications

Brainstem Organoids From Human Pluripotent Stem Cells

Brainstem Organoids From Human Pluripotent Stem Cells

Brainstem organoids from human pluripotent stem cells contain neural crest population

The Rho-associated kinase inhibitor fasudil can replace Y-27632 for use in human pluripotent stem cell research

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