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Neuroinflammatory astrocytes generated from cord blood-derived human induced pluripotent stem cells.

ABSTRACT: BACKGROUND:Astrocytes respond to central nervous system (CNS) injury and disease by transforming to a reactive astrogliosis cell state that can contribute to either CNS dysfunction or repair. Neuroinflammation is a powerful driver of a harmful A1 astrogliosis phenotype associated with in vitro neurotoxicity and histopathology in human neurodegenerative diseases. Here we report a protocol for the rapid development of a human cell culture model of neuroinflammatory astrogliosis using induced pluripotent stem cells (iPSCs). METHODS:Using RNA sequencing and in vitro cell assays, we measured transcriptional and cellular effects of chronic exposure of human iPSC-derived astrocytes to the cytokines TNF? (tumor necrosis factor alpha) or IL-1? (interleukin-1 beta). RESULTS:We show TNF? and IL-1? induce pro-inflammatory gene signatures but by widely different magnitudes. TNF? treatment results in 606 differential expressed genes, the suppression of glutamate-uptake, and increased phagocytic activity in astrocyte cultures. In contrast, IL-1? effects are attenuated to 33 differential expressed genes and no significant effects on glutamate-uptake or increased phagocytic activity. CONCLUSION:Our approach demonstrates a rapid tool for modeling neuroinflammatory human astrocytic responses in nervous system trauma and disease. In particular, we reveal a model for robust TNF?-induced human astrogliosis suitable for the study of neurotoxic A1 astrocytes.

SUBMITTER: Zhou Q

PROVIDER: S-EPMC6688278 | biostudies-literature | 2019 Aug

REPOSITORIES: biostudies-literature

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Neuroinflammatory astrocytes generated from cord blood-derived human induced pluripotent stem cells.

Zhou Qiong Q Viollet Coralie C Efthymiou Anastasia A Khayrullina Guzal G Moritz Kasey E KE Wilkerson Matthew D MD Sukumar Gauthaman G Dalgard Clifton L CL Doughty Martin L ML

Journal of neuroinflammation 20190809 1

<h4>Background</h4>Astrocytes respond to central nervous system (CNS) injury and disease by transforming to a reactive astrogliosis cell state that can contribute to either CNS dysfunction or repair. Neuroinflammation is a powerful driver of a harmful A1 astrogliosis phenotype associated with in vitro neurotoxicity and histopathology in human neurodegenerative diseases. Here we report a protocol for the rapid development of a human cell culture model of neuroinflammatory astrogliosis using induc ...[more]

PMID: 31395092

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Project description:IntroductionHuman induced pluripotent stem cells (hiPSCs) offer great promise for regenerative therapies or in vitro modelling of neurodegenerative disorders like Parkinson's disease. Currently, widely used cell sources for the generation of hiPSCs are somatic cells obtained from aged individuals. However, a critical issue concerning the potential clinical use of these iPSCs is mutations that accumulate over lifetime and are transferred onto iPSCs during reprogramming which may influence the functionality of cells differentiated from them. The aim of our study was to establish a differentiation strategy to efficiently generate neurons including dopaminergic cells from human cord blood-derived iPSCs (hCBiPSCs) as a juvenescent cell source and prove their functional maturation in vitro.MethodsThe differentiation of hCBiPSCs was initiated by inhibition of transforming growth factor-β and bone morphogenetic protein signaling using the small molecules dorsomorphin and SB 431542 before final maturation was carried out. hCBiPSCs and differentiated neurons were characterized by immunocytochemistry and quantitative real time-polymerase chain reaction. Since functional investigations of hCBiPSC-derived neurons are indispensable prior to clinical applications, we performed detailed analysis of essential ion channel properties using whole-cell patch-clamp recordings and calcium imaging.ResultsA Sox1 and Pax6 positive neuronal progenitor cell population was efficiently induced from hCBiPSCs using a newly established differentiation protocol. Neuronal progenitor cells could be further maturated into dopaminergic neurons expressing tyrosine hydroxylase, the dopamine transporter and engrailed 1. Differentiated hCBiPSCs exhibited voltage-gated ion currents, were able to fire action potentials and displayed synaptic activity indicating synapse formation. Application of the neurotransmitters GABA, glutamate and acetylcholine induced depolarizing calcium signal changes in neuronal cells providing evidence for the excitatory effects of these ligand-gated ion channels during maturation in vitro.ConclusionsThis study demonstrates for the first time that hCBiPSCs can be used as a juvenescent cell source to generate a large number of functional neurons including dopaminergic cells which may serve for the development of novel regenerative treatment strategies.

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Neuroinflammatory astrocytes generated from cord blood-derived human induced pluripotent stem cells.

Publications

Neuroinflammatory astrocytes generated from cord blood-derived human induced pluripotent stem cells.

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