Rapid and robust generation of long-term self-renewing human neural stem cells with the ability to generate mature astroglia.

Thomas Palm,Jens Christian Schwamborn,Claudia Träger,Ralf-Leslie Meilenbrock,Johannes Brockhaus,Sarah Nickels,Silvia Bolognin,Markus Missler,Miriam Schreitmüller,Johannes Meiser

doi:10.1038/srep16321

Thomas Palm, Jens Christian Schwamborn + Show 8 more

Open Access

https://doi.org/10.1038/srep16321

Copy DOI

Abstract

Induced pluripotent stem cell bear the potential to differentiate into any desired cell type and hold large promise for disease-in-a-dish cell-modeling approaches. With the latest advances in the field of reprogramming technology, the generation of patient-specific cells has become a standard technology. However, directed and homogenous differentiation of human pluripotent stem cells into desired specific cell types remains an experimental challenge. Here, we report the development of a novel hiPSCs-based protocol enabling the generation of expandable homogenous human neural stem cells (hNSCs) that can be maintained under self-renewing conditions over high passage numbers. Our newly generated hNSCs retained differentiation potential as evidenced by the reliable generation of mature astrocytes that display typical properties as glutamate up-take and expression of aquaporin-4. The hNSC-derived astrocytes showed high activity of pyruvate carboxylase as assessed by stable isotope assisted metabolic profiling. Moreover, using a cell transplantation approach, we showed that grafted hNSCs were not only able to survive but also to differentiate into astroglial in vivo. Engraftments of pluripotent stem cells derived from somatic cells carry an inherent tumor formation potential. Our results demonstrate that hNSCs with self-renewing and differentiation potential may provide a safer alternative strategy, with promising applications especially for neurodegenerative disorders.

Highlights

Time and cost consuming maintenance of pluripotent stem cells as well as potential immune rejection and teratoma formation[6]
In the astrocyte cultures we found that glutamine provided similar amounts of carbons for the synthesis of citrate as glucose (22%) (Fig. 5d)
Based on stable isotope experiments we found higher relative serine biosynthesis rates in multi-lineage differentiation cultures (MLDCs) (24%) as in pure astrocyte cultures (5%) (Fig. 6b,c) where most serine originated from the medium (M0 isotopologue abundance)

Summary

Introduction

Time and cost consuming maintenance of pluripotent stem cells as well as potential immune rejection and teratoma formation[6]. It has been shown that glia cells have a direct, non-cell autonomous effect on motor neuron survival in a mouse model of amyptrophic lateral sclerosis[11] Despite this growing evidence, astrocytes are less investigated compared to neurons. In the field of neurodegenerative diseases, hundreds of compounds have been successfully used in animal models to ameliorate induced-neuropathology or cognitive deficits This has not translated into effective disease-modifying therapies for humans. The use of iPSCs overcomes the legal/ethical concerns as well as the risks of immune rejection of human embryonic stem cells (hESCs). Despite these advantages, iPSCs, as well as hESCs, have the inherent potential for teratoma formation[6]. We put a specific emphasis on the generation of long-term expandable human astrocyte cultures, which can be of use for modeling of disease-specific pathological traits

Methods

Results

Conclusion