Functional maturation of human neural stem cells in a 3D bioengineered brain model enriched with fetal brain-derived matrix

Disha Sood,Sabato Santaniello,Karl Deisseroth,Dana M Cairns,Lauren D Black,Jayanth M Dabbi,Charu Ramakrishnan,David L Kaplan

doi:10.1038/s41598-019-54248-1

Disha Sood, Sabato Santaniello + Show 6 more

Open Access

https://doi.org/10.1038/s41598-019-54248-1

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Abstract

Brain extracellular matrix (ECM) is often overlooked in vitro brain tissue models, despite its instructive roles during development. Using developmental stage-sourced brain ECM in reproducible 3D bioengineered culture systems, we demonstrate enhanced functional differentiation of human induced neural stem cells (hiNSCs) into healthy neurons and astrocytes. Particularly, fetal brain tissue-derived ECM supported long-term maintenance of differentiated neurons, demonstrated by morphology, gene expression and secretome profiling. Astrocytes were evident within the second month of differentiation, and reactive astrogliosis was inhibited in brain ECM-enriched cultures when compared to unsupplemented cultures. Functional maturation of the differentiated hiNSCs within fetal ECM-enriched cultures was confirmed by calcium signaling and spectral/cluster analysis. Additionally, the study identified native biochemical cues in decellularized ECM with notable comparisons between fetal and adult brain-derived ECMs. The development of novel brain-specific biomaterials for generating mature in vitro brain models provides an important path forward for interrogation of neuron-glia interactions.

Highlights

A common limitation of current 3D in vitro brain models is that the extracellular matrix (ECM) content is often not considered in detail, even though brain ECM is dynamic during development and plays a crucial role in cell signaling and homeostasis[18]
The impact of some specific brain-ECM constituents such as laminin[25,26] and adult brain-derived ECM on cell differentiation, synapse formation and mechanical properties has been studied in isolation[27,28,29]; the study of composite, scaffold-based 3D in vitro systems to investigate the bioactivity of ECM from different developmental stages over long-term differentiation of human induced neural stem cells into both neurons and astrocytes is lacking
With the likelihood of epigenetic changes to be maintained in directly reprogrammed cells in comparison to induced pluripotent stem cells (iPSCs), as noted in previous studies[40,41], we anticipate that human induced neural stem cells (hiNSCs)-based models will better recapitulate neurodegenerative disease phenotypes. hiNSCs were cultured in bioengineered silk protein scaffold-based 3D tissue constructs infused with collagen type I (CLG1, previously shown to be compatible with brain cells42) hydrogels supplemented with native brain-derived ECM

Summary

Introduction

A common limitation of current 3D in vitro brain models is that the ECM content is often not considered in detail, even though brain ECM is dynamic during development and plays a crucial role in cell signaling and homeostasis[18]. During development, ECM guides the compartmentalization of functional brain microdomains, and contributes to the sophisticated architecture and function of the brain[23] Such native ECM signals are important for differentiation of neural progenitor/stem cells[24]. We investigated the effects of brain-derived ECM from two different developmental stages (fetal versus adult) on the differentiation of previously characterized hiNSCs39 into neurons and healthy astrocytes, when cultured within relevant environmental cues (biochemical factors and 3D topology) (Fig. 1). HiNSCs were cultured in bioengineered silk protein scaffold-based 3D tissue constructs infused with collagen type I (CLG1, previously shown to be compatible with brain cells42) hydrogels supplemented with native brain-derived ECM (fetal or adult). One-way ANOVA with Tukey’s post hoc for multiple comparisons. *p < 0.0431, **p < 0.0071

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