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Differentiation from human pluripotent stem cells of cortical neurons of the superficial layers amenable to psychiatric disease modeling and high-throughput drug screening.

Boissart C, Poulet A, Georges P, Darville H, Julita E, Delorme R, Bourgeron T, Peschanski M, Benchoua A - Transl Psychiatry (2013)

Bottom Line: Addressing this issue, we have successfully captured a stable population of self-renewing late cortical progenitors (LCPs) that synchronously and massively differentiate into glutamatergic cortical neurons of the upper layers.This has allowed us to analyze the capability of LCPs at differentiating into post mitotic neurons as well as extending and branching neurites in response to a collection of selected bioactive molecules.LCPs and cortical neurons of the upper layers were successfully produced from patient-derived-induced PSC, indicating that this system enables functional studies of individual-specific cortical neurons ex vivo for disease modeling and therapeutic purposes.

View Article: PubMed Central - PubMed

Affiliation: Neuroplasticity and Therapeutics, CECS, I-STEM, AFM, Evry Cedex, France.

ABSTRACT
Cortical neurons of the superficial layers (II-IV) represent a pivotal neuronal population involved in the higher cognitive functions of the human and are particularly affected by psychiatric diseases with developmental manifestations such as schizophrenia and autism. Differentiation protocols of human pluripotent stem cells (PSC) into cortical neurons have been achieved, opening the way to in vitro modeling of neuropsychiatric diseases. However, these protocols commonly result in the asynchronous production of neurons typical for the different layers of the cortex within an extended period of culture, thus precluding the analysis of specific subtypes of neurons in a standardized manner. Addressing this issue, we have successfully captured a stable population of self-renewing late cortical progenitors (LCPs) that synchronously and massively differentiate into glutamatergic cortical neurons of the upper layers. The short time course of differentiation into neurons of these progenitors has made them amenable to high-throughput assays. This has allowed us to analyze the capability of LCPs at differentiating into post mitotic neurons as well as extending and branching neurites in response to a collection of selected bioactive molecules. LCPs and cortical neurons of the upper layers were successfully produced from patient-derived-induced PSC, indicating that this system enables functional studies of individual-specific cortical neurons ex vivo for disease modeling and therapeutic purposes.

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Related in: MedlinePlus

Differentiation of induced pluripotent stem cells (iPSC) into neurons of the superficial layers. (a) Representative immunocytochemistry of late cortical progenitors (LCP) and cortical neurons produced from four different iPSC lines. 4603 and 1869 iPSC lines were derived from control donors and autism spectrum disorder 1 (ASD1) and ASD2 from autistic children with SHANK-3 mutations. Scale bar=50 μm (b) Kinetic of differentiation of iPSC-derived cortical neurons. (c) Percentage of upper layers neurons produced after 17 days of differentiation of iPSC-derived LCP. For (b, c) results are expressed as mean +/− s.d. of three independent experiments.
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fig5: Differentiation of induced pluripotent stem cells (iPSC) into neurons of the superficial layers. (a) Representative immunocytochemistry of late cortical progenitors (LCP) and cortical neurons produced from four different iPSC lines. 4603 and 1869 iPSC lines were derived from control donors and autism spectrum disorder 1 (ASD1) and ASD2 from autistic children with SHANK-3 mutations. Scale bar=50 μm (b) Kinetic of differentiation of iPSC-derived cortical neurons. (c) Percentage of upper layers neurons produced after 17 days of differentiation of iPSC-derived LCP. For (b, c) results are expressed as mean +/− s.d. of three independent experiments.

Mentions: In order to evaluate whether our protocol was applicable in the context of psychiatric diseases, LCP and neurons of the cortical superficial layers were produced from iPSCs, both from controls and from two autistic patients bearing de novo mutations on SHANK-3 (16). All four lines were efficiently converted into early cortical NEPs and stabilized as proliferating LCPs expressing Sox2 and nestin (Figure 5a). LCPs readily differentiated into HuCD/Tuj-1-positive neurons with the same efficiency than hESC (Figures 5a and b). The majority of neurons produced were also similarly positive for superficial layers marker in the four lines.


Differentiation from human pluripotent stem cells of cortical neurons of the superficial layers amenable to psychiatric disease modeling and high-throughput drug screening.

Boissart C, Poulet A, Georges P, Darville H, Julita E, Delorme R, Bourgeron T, Peschanski M, Benchoua A - Transl Psychiatry (2013)

Differentiation of induced pluripotent stem cells (iPSC) into neurons of the superficial layers. (a) Representative immunocytochemistry of late cortical progenitors (LCP) and cortical neurons produced from four different iPSC lines. 4603 and 1869 iPSC lines were derived from control donors and autism spectrum disorder 1 (ASD1) and ASD2 from autistic children with SHANK-3 mutations. Scale bar=50 μm (b) Kinetic of differentiation of iPSC-derived cortical neurons. (c) Percentage of upper layers neurons produced after 17 days of differentiation of iPSC-derived LCP. For (b, c) results are expressed as mean +/− s.d. of three independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3756296&req=5

fig5: Differentiation of induced pluripotent stem cells (iPSC) into neurons of the superficial layers. (a) Representative immunocytochemistry of late cortical progenitors (LCP) and cortical neurons produced from four different iPSC lines. 4603 and 1869 iPSC lines were derived from control donors and autism spectrum disorder 1 (ASD1) and ASD2 from autistic children with SHANK-3 mutations. Scale bar=50 μm (b) Kinetic of differentiation of iPSC-derived cortical neurons. (c) Percentage of upper layers neurons produced after 17 days of differentiation of iPSC-derived LCP. For (b, c) results are expressed as mean +/− s.d. of three independent experiments.
Mentions: In order to evaluate whether our protocol was applicable in the context of psychiatric diseases, LCP and neurons of the cortical superficial layers were produced from iPSCs, both from controls and from two autistic patients bearing de novo mutations on SHANK-3 (16). All four lines were efficiently converted into early cortical NEPs and stabilized as proliferating LCPs expressing Sox2 and nestin (Figure 5a). LCPs readily differentiated into HuCD/Tuj-1-positive neurons with the same efficiency than hESC (Figures 5a and b). The majority of neurons produced were also similarly positive for superficial layers marker in the four lines.

Bottom Line: Addressing this issue, we have successfully captured a stable population of self-renewing late cortical progenitors (LCPs) that synchronously and massively differentiate into glutamatergic cortical neurons of the upper layers.This has allowed us to analyze the capability of LCPs at differentiating into post mitotic neurons as well as extending and branching neurites in response to a collection of selected bioactive molecules.LCPs and cortical neurons of the upper layers were successfully produced from patient-derived-induced PSC, indicating that this system enables functional studies of individual-specific cortical neurons ex vivo for disease modeling and therapeutic purposes.

View Article: PubMed Central - PubMed

Affiliation: Neuroplasticity and Therapeutics, CECS, I-STEM, AFM, Evry Cedex, France.

ABSTRACT
Cortical neurons of the superficial layers (II-IV) represent a pivotal neuronal population involved in the higher cognitive functions of the human and are particularly affected by psychiatric diseases with developmental manifestations such as schizophrenia and autism. Differentiation protocols of human pluripotent stem cells (PSC) into cortical neurons have been achieved, opening the way to in vitro modeling of neuropsychiatric diseases. However, these protocols commonly result in the asynchronous production of neurons typical for the different layers of the cortex within an extended period of culture, thus precluding the analysis of specific subtypes of neurons in a standardized manner. Addressing this issue, we have successfully captured a stable population of self-renewing late cortical progenitors (LCPs) that synchronously and massively differentiate into glutamatergic cortical neurons of the upper layers. The short time course of differentiation into neurons of these progenitors has made them amenable to high-throughput assays. This has allowed us to analyze the capability of LCPs at differentiating into post mitotic neurons as well as extending and branching neurites in response to a collection of selected bioactive molecules. LCPs and cortical neurons of the upper layers were successfully produced from patient-derived-induced PSC, indicating that this system enables functional studies of individual-specific cortical neurons ex vivo for disease modeling and therapeutic purposes.

Show MeSH
Related in: MedlinePlus