Co-culture of Human Stem Cell Derived Neurons and Oligodendrocyte Progenitor Cells

Dooves, Stephanie; Nadadhur, Aishwarya G.; Gasparotto, Lisa; Heine, Vivi M.

doi:10.21769/bioprotoc.3350

Cited by 6 publications

(7 citation statements)

References 17 publications

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“…Cortical neuron differentiation was performed as previously described(22) using dual SMAD inhibition and consequent maturation of cells into both glutamatergic and GABAergic neurons. For myelinating cultures, human iPSCs were differentiated into glial progenitor cells as previously described(23). See supplementary method section for more details on culture protocols.…”

Section: Methodsmentioning

confidence: 99%

“…Therefore, we decided to analyze oligodendrocyte maturation and myelination on human neuron-oligodendrocyte cocultures. We altered a previously published protocol for neuron-OPC culture (23) to generate a myelinating co-culture. Indeed, after 40 days of co-culture we observed a robust generation of mature oligodendrocytes (defined as MBP + cells) and myelination, shown by overlap between MBP and NF200 staining (Fig.…”

Section: H Neurons Show Robust Myelination In Vitromentioning

confidence: 99%

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Cortical interneuron development is affected in leukodystrophy 4H

Dooves

Kok

Holmes

et al. 2022

Preprint

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4H leukodystrophy is a rare genetic disorder classically characterized by hypomyelination, hypodontia and hypogonadotropic hypogonadism. With the discovery that 4H is caused by mutations that affect RNA polymerase III, mainly involved in the transcription of small non-coding RNAs, also patients with atypical presentations with mainly a neuronal phenotype were identified. Pathomechanisms of 4H brain abnormalities are still unknown and research is hampered by a lack of preclinical models. We aimed to identify cells and pathways that are affected by 4H mutations using induced pluripotent stem cell models. RNA sequencing analysis on induced pluripotent stem cell-derived cerebellar cells revealed several differentially expressed genes between 4H patients and control samples, including reduced ARX expression. As ARX is involved in early brain and interneuron development, we studied and confirmed interneuron changes in primary tissue of 4H patients. Subsequently, we studied interneuron changes in more depth and analyzed induced pluripotent stem cell-derived cortical neuron cultures for changes in neuronal morphology, synaptic balance, network activity and myelination. We showed a decreased percentage of GABAergic synapses in 4H, which correlated to increased neuronal network activity. Treatment of cultures with GABA antagonists led to a significant increase in neuronal network activity in control cells but not in 4H cells, also pointing to lack of inhibitory activity in 4H. Myelination and oligodendrocyte maturation in cultures with 4H neurons was normal, and treatment with sonic hedgehog agonist SAG did not improve 4H related neuronal phenotypes. qPCR analysis revealed increased expression of parvalbumin interneuron marker ERBB4, suggesting that the development rather than generation of interneurons may be affected in 4H. Together, these results indicate that interneurons are involved, possibly parvalbumin interneurons, in disease mechanisms of 4H leukodystrophy.

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Section: Methodsmentioning

confidence: 99%

Section: H Neurons Show Robust Myelination In Vitromentioning

confidence: 99%

Cortical interneuron development is affected in leukodystrophy 4H

Dooves

Kok

Holmes

et al. 2022

Preprint

Self Cite

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“… 68 , 69 In line with this, co‐cultures of human iPSC‐derived neurons and oligodendrocytes, which allow the study of neuron–glia cross‐talk and myelination processes, led to the observation of neuronal alterations related to tuberous sclerosis complex that were not apparent in neuronal monocultures. 70 These issues show that iPSC‐derived neurons, while invaluable for studying both coding and noncoding gene variants in the context of a human genetic background, should be optimized for the questions being asked and interpreted with care.…”

Section: Functional Consequences Of a Noncoding Aud ...mentioning

confidence: 99%

5. Collaborative Study on the Genetics of Alcoholism: Functional genomics

Gameiro‐Ros

Popova

Prytkova

et al. 2023

Genes Brain and Behavior

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Alcohol Use Disorder is a complex genetic disorder, involving genetic, neural, and environmental factors, and their interactions. The Collaborative Study on the Genetics of Alcoholism (COGA) has been investigating these factors and identified putative alcohol use disorder risk genes through genome‐wide association studies. In this review, we describe advances made by COGA in elucidating the functional changes induced by alcohol use disorder risk genes using multimodal approaches with human cell lines and brain tissue. These studies involve investigating gene regulation in lymphoblastoid cells from COGA participants and in post‐mortem brain tissues. High throughput reporter assays are being used to identify single nucleotide polymorphisms in which alternate alleles differ in driving gene expression. Specific single nucleotide polymorphisms (both coding or noncoding) have been modeled using induced pluripotent stem cells derived from COGA participants to evaluate the effects of genetic variants on transcriptomics, neuronal excitability, synaptic physiology, and the response to ethanol in human neurons from individuals with and without alcohol use disorder. We provide a perspective on future studies, such as using polygenic risk scores and populations of induced pluripotent stem cell‐derived neurons to identify signaling pathways related with responses to alcohol. Starting with genes or loci associated with alcohol use disorder, COGA has demonstrated that integration of multimodal data within COGA participants and functional studies can reveal mechanisms linking genomic variants with alcohol use disorder, and potential targets for future treatments.

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“…This strategy was successfully applied to differentiate PSCs into a variety of neuronal subtypes relevant to neurodegenerative and neuropsychiatric diseases [ 46 , 47 ], such as hippocampus CA3 pyramidal neurons, which exhibit the electrophysiological properties of mossy fibers of the dendate gyrus [ 48 ]; hypothalamic-like neurons capable of secreting orexigenic and anorexigenic neuropeptides and responding appropriately to the metabolic hormones ghrelin and leptin [ 49 ]; GABAergic interneurons of the cortex and the basal ganglia [ 50 , 51 ]; serotoninergic neurons of the raphe nuclei [ 52 , 53 ]; dopaminergic neurons of the substantia nigra [ 44 , 54 ]; cortical pyramidal neurons [ 55 , 56 , 57 ]; and spinal motoneurons secreting acetylcholine [ 58 , 59 ]. Glial cells can also be derived from PSCs, including astrocytes [ 60 , 61 , 62 , 63 , 64 ], oligodendrocytes [ 65 , 66 , 67 , 68 ] and microglia [ 69 , 70 , 71 , 72 ], allowing the co-culture and recapitulation of cell-autonomous and non-cell-autonomous mechanisms leading to disease progression to be conducted [ 73 , 74 , 75 , 76 , 77 , 78 , 79 , 80 , 81 , 82 ]. Next to these “growth-factor”-mediated protocols, several groups have described methods of direct conversion of PSC into sub-types of neurons using forced expression of transcription factors.…”

Section: Integration Of Psc-derived Models In the Process Of Drug Discoverymentioning

confidence: 99%

Contribution of Human Pluripotent Stem Cell-Based Models to Drug Discovery for Neurological Disorders

Benchoua

Lasbareilles

Tournois

2021

Cells

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One of the major obstacles to the identification of therapeutic interventions for central nervous system disorders has been the difficulty in studying the step-by-step progression of diseases in neuronal networks that are amenable to drug screening. Recent advances in the field of human pluripotent stem cell (PSC) biology offers the capability to create patient-specific human neurons with defined clinical profiles using reprogramming technology, which provides unprecedented opportunities for both the investigation of pathogenic mechanisms of brain disorders and the discovery of novel therapeutic strategies via drug screening. Many examples not only of the creation of human pluripotent stem cells as models of monogenic neurological disorders, but also of more challenging cases of complex multifactorial disorders now exist. Here, we review the state-of-the art brain cell types obtainable from PSCs and amenable to compound-screening formats. We then provide examples illustrating how these models contribute to the definition of new molecular or functional targets for drug discovery and to the design of novel pharmacological approaches for rare genetic disorders, as well as frequent neurodegenerative diseases and psychiatric disorders.

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Co-culture of Human Stem Cell Derived Neurons and Oligodendrocyte Progenitor Cells

Cited by 6 publications

References 17 publications

Cortical interneuron development is affected in leukodystrophy 4H

Cortical interneuron development is affected in leukodystrophy 4H

5. Collaborative Study on the Genetics of Alcoholism: Functional genomics

Contribution of Human Pluripotent Stem Cell-Based Models to Drug Discovery for Neurological Disorders

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