Efficient Derivation of Excitatory and Inhibitory Neurons from Human Pluripotent Stem Cells Stably Expressing Direct Reprogramming Factors

Song, Saera; Ashok, Archana; Williams, Damian J.; Kaufman, Maria; Duff, Karen; Sproul, Andrew A.

doi:10.1002/cpz1.141

Cited by 9 publications

(10 citation statements)

References 49 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…All human cell line work was performed on de-identified cell lines and approved by the Columbia University Institutional Review Board (IRB). IMR90 cl.4 hiPSCs (WiCell) (Yu et al, 2007; Yu et al, 2009; Chen et al, 2011; Hu et al, 2011) were grown in StemFlex media (Thermo Scientific) on Cultrex (Biotechne) and passaged with ReLeSR as described previously (Song et al, 2021). Human neural progenitor cells (NPCs) were generated using dual SMAD inhibition as non-adherent embryoid bodies, followed by plating on polyornithine (10 ug/mL, Sigma P4957)/laminin (10 ug/mL, R&D Systems 3400-010-02) and subsequent manual rosette selection and expansion, as described previously (Topol et al, 2015; Sun et al, 2019).…”

Section: Methodsmentioning

confidence: 99%

ZCCHC17 modulates neuronal RNA splicing and supports cognitive resilience in Alzheimer’s disease

Bartosch

Youth

Hansen

et al. 2023

Preprint

Self Cite

View full text Add to dashboard Cite

ZCCHC17 is a putative master regulator of synaptic gene dysfunction in Alzheimer's Disease (AD), and ZCCHC17 protein declines early in AD brain tissue, before significant gliosis or neuronal loss. Here, we investigate the function of ZCCHC17 and its role in AD pathogenesis. Co-immunoprecipitation of ZCCHC17 followed by mass spectrometry analysis in human iPSC-derived neurons reveals that ZCCHC17's binding partners are enriched for RNA splicing proteins. ZCCHC17 knockdown results in widespread RNA splicing changes that significantly overlap with splicing changes found in AD brain tissue, with synaptic genes commonly affected. ZCCHC17 expression correlates with cognitive resilience in AD patients, and we uncover an APOE4 dependent negative correlation of ZCCHC17 expression with tangle burden. Furthermore, a majority of ZCCHC17 interactors also co-IP with known tau interactors, and we find significant overlap between alternatively spliced genes in ZCCHC17 knockdown and tau overexpression neurons. These results demonstrate ZCCHC17's role in neuronal RNA processing and its interaction with pathology and cognitive resilience in AD, and suggest that maintenance of ZCCHC17 function may be a therapeutic strategy for preserving cognitive function in the setting of AD pathology.

show abstract

Section: Methodsmentioning

confidence: 99%

ZCCHC17 modulates neuronal RNA splicing and supports cognitive resilience in Alzheimer’s disease

Bartosch

Youth

Hansen

et al. 2023

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…We recommend adhering to the user's own preferences for methods involving RNA [quantitative reverse transcription PCR (RT-qPCR); single nucleus/single cell RNA sequencing (sn/scRNA-seq)] and protein (immunocytochemistry; western blot) analyses. Suggestions from existing Current Protocols articles include immunofluorescence staining on dissociated neurons (Song et al, 2021) or whole organoids (Lewis-Israeli et al, 2022), and electrophysiological characterization using patch clamp (Song et al, 2021). Additional functional characterization can be achieved using calcium imaging techniques (Galiakberova et al, 2022;Jiang et al, 2018;Sharma, Saha, Joseph, Krishnan, & Raghu, 2020;Vőfély et al, 2018) or multi-electrode array (MEA) platforms from companies such as Axion Biosystems (www.axionbiosystems.com) or Maxwell Biosystems (www.mxwbio.com).…”

Section: Strategic Planningmentioning

confidence: 99%

A Robust Pipeline for the Multi‐Stage Accelerated Differentiation of Functional 3D Cortical Organoids from Human Pluripotent Stem Cells

et al. 2023

View full text Add to dashboard Cite

Disordered cellular development, abnormal neuroanatomical formations, and dysfunction of neuronal circuitry are among the pathological manifestations of cortical regions in the brain that are often implicated in complex neurodevelopmental disorders. With the advancement of stem cell methodologies such as cerebral organoid generation, it is possible to study these processes in vitro using 3D cellular platforms that mirror key developmental stages occurring throughout embryonic neurogenesis. Patterning‐based stem cell models of directed neuronal development offer one approach to accomplish this, but these protocols often require protracted periods of cell culture to generate diverse cell types and current methods are plagued by a lack of specificity, reproducibility, and temporal control over cell derivation. Although ectopic expression of transcription factors offers another avenue to rapidly generate neurons, this process of direct lineage conversion bypasses critical junctures of neurodevelopment during which disease‐relevant manifestations may occur. Here, we present a directed differentiation approach for generating human pluripotent stem cell (hPSC)‐derived cortical organoids with accelerated lineage specification to generate functionally mature cortical neurons in a shorter timeline than previously established protocols. This novel protocol provides precise guidance for the specification of neuronal cell type identity as well as temporal control over the pace at which cortical lineage trajectories are established. Furthermore, we present assays that can be used as tools to interrogate stage‐specific developmental signaling mechanisms. By recapitulating major components of embryonic neurogenesis, this protocol allows for improved in vitro modeling of cortical development while providing a platform that can be utilized to uncover disease‐specific mechanisms of disordered development at various stages across the differentiation timeline. © 2023 Wiley Periodicals LLC. Basic Protocol 1: 3D hPSC neural induction Support Protocol 1: Neural rosette formation assay Support Protocol 2: Neurosphere generation Support Protocol 3: Enzymatic dissociation, NSC expansion, and cryopreservation Basic Protocol 2: 3D neural progenitor expansion Basic Protocol 3: 3D accelerated cortical lineage patterning and terminal differentiation

show abstract

“…42 Indeed, the targeted generation of a homogenous cellular state from a pluripotent stem cell is now frequently enforced using gene overexpression. 43 In 2006, the laboratory of Takahashi and Yamanaka performed a limited genetic screen in mouse actively selecting for combinations of genes able to dedifferentiate an adult fibroblast to a pluripotent state equivalent to the inner cell mass of the blastocyst. 8 Subsequently demonstrated using human somatic cells, 44 this astounding observation has revolutionized the stem cell field, providing an opportunity to recreate pluripotent stem cells from any individual.…”

Section: Adult Kidney Stem Cellsmentioning

confidence: 99%

“…This concept of dedifferentiation or transdifferentiation via the expression of critical gene networks is now readily being applied with global transcriptional profiling and sophisticated bioinformatic algorithms predicting how to change one cellular state into another 42 . Indeed, the targeted generation of a homogenous cellular state from a pluripotent stem cell is now frequently enforced using gene overexpression 43 …”

Section: Proposed Renal Therapeutic Options In 2006mentioning

confidence: 99%

Regrow or Repair: An Update on Potential Regenerative Therapies for the Kidney

Little

Humphreys

2022

JASN

View full text Add to dashboard Cite

Fifteen years ago, this journal published a review outlining future options for regenerating the kidney. At that time, stem cell populations were being identified in multiple tissues, the concept of stem cell recruitment to a site of injury was of great interest, and the possibility of postnatal renal stem cells was growing in momentum. Since that time, we have seen the advent of human induced pluripotent stem cells, substantial advances in our capacity to both sequence and edit the genome, global and spatial transcriptional analysis down to the single-cell level, and a pandemic that has challenged our delivery of health care to all. This article will look back over this period of time to see how our view of kidney development, disease, repair, and regeneration has changed and envision a future for kidney regeneration and repair over the next 15 years.

show abstract

Efficient Derivation of Excitatory and Inhibitory Neurons from Human Pluripotent Stem Cells Stably Expressing Direct Reprogramming Factors

Abstract: A. (2021). Efficient derivation of excitatory and inhibitory neurons from human pluripotent stem cells stably expressing direct reprogramming factors. Current Protocols, 1, e141.

Cited by 9 publications

References 49 publications

ZCCHC17 modulates neuronal RNA splicing and supports cognitive resilience in Alzheimer’s disease

ZCCHC17 modulates neuronal RNA splicing and supports cognitive resilience in Alzheimer’s disease

A Robust Pipeline for the Multi‐Stage Accelerated Differentiation of Functional 3D Cortical Organoids from Human Pluripotent Stem Cells

Regrow or Repair: An Update on Potential Regenerative Therapies for the Kidney

Contact Info

Product

Resources

About