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Alginate-based 3D culture of human induced pluripotent stem cells and their differentiation to functional dopaminergic neurons

ABSTRACT: Alginate-based 3D culture of human induced pluripotent stem cells and their differentiation to functional dopaminergic neurons

PROVIDER: PRJNA740008 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR14880023_1.fastq.gz	Fastqsanger.gz
	SRR14880023_2.fastq.gz	Fastqsanger.gz
	SRR14880024_1.fastq.gz	Fastqsanger.gz
	SRR14880024_2.fastq.gz	Fastqsanger.gz
	SRR14880025_1.fastq.gz	Fastqsanger.gz

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Project description:Human induced pluripotent stem cells (hiPSCs) represent an unlimited cell source for the generation of in vitro models of patient-specific dopaminergic (DA) neurons, posing a possibility to overcome the restricted accessibility to disease-affected tissue for mechanistic studies on Parkinson’s disease (PD). However, the complexity of the human brain is not fully recapitulated by existing monolayer culture methods. Consequently, neurons differentiated in a three dimensional (3D) in vitro culture system might better mimic the in vivo cellular environment for basic mechanistic studies and represent better predictors of drug responses in vivo. Thus, in this work we established a new in vitro cell culture model based on the microencapsulation of hiPSCs in small alginate/fibronectin beads and their differentiation to DA neurons. Optimisation of hydrogel matrix concentrations and composition resulted in high viability of embedded iPSCs. Neural differentiation capacity and DA neuronal yield, analyzed by qRT-PCR, immunofluorescence, and western blot analyses, were increased in the 3D neuronal cultures compared to neurons generated using the conventional 2D culture system. Additionally, electrophysiological parameters and metabolic switching profile supported the increased functionality and an anticipated metabolic resetting of neurons grown in alginate scaffolds with respect to the 2D neurons. We also report long-term maintenance of neuronal cultures and mature functional properties. Furthermore, our findings indicate that our 3D model system can recapitulate mitochondrial superoxide production as an important mitochondrial phenotype observed in PD patients’-derived neurons and that this phenotype might be detectable earlier during neuronal differentiation. Taken together, these results indicate that our alginate-based 3D culture system offers an advantageous strategy for the reliable and rapid derivation of mature and functional DA neurons from iPSCs.

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Alginate-based 3D culture of human induced pluripotent stem cells and their differentiation to functional dopaminergic neurons

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