Different iPSC-derived neural stem cells shows various spectrums of spontaneous differentiation during long term cultivation

Adelya Albertovna Galiakberova; Olga Igorevna Brovkina; Nikolay Vitalyevich Kondratyev; Alexander Sergeevich Artyuhov; Ekaterina Dmitrievna Momotyuk; Olga Nikolaevna Kulmukhametova; Alexey Aleksandrovich Lagunin; Boris Vladimirovich Shilov; Anton Dmitrievich Zadorozhny; Igor Sergeevitch Zakharov; Larisa Sergeevna Okorokova; Vera Evgenievna Golimbet; Erdem Bairovich Dashinimaev

doi:10.3389/fnmol.2023.1037902

Different iPSC-derived neural stem cells shows various spectrums of spontaneous differentiation during long term cultivation

Front Mol Neurosci. 2023 May 2:16:1037902. doi: 10.3389/fnmol.2023.1037902. eCollection 2023.

Authors

Adelya Albertovna Galiakberova^{1

2}, Olga Igorevna Brovkina³, Nikolay Vitalyevich Kondratyev⁴, Alexander Sergeevich Artyuhov¹, Ekaterina Dmitrievna Momotyuk¹, Olga Nikolaevna Kulmukhametova⁵, Alexey Aleksandrovich Lagunin^{5

6}, Boris Vladimirovich Shilov⁵, Anton Dmitrievich Zadorozhny⁵, Igor Sergeevitch Zakharov⁷, Larisa Sergeevna Okorokova⁸, Vera Evgenievna Golimbet⁴, Erdem Bairovich Dashinimaev^{1

7

9}

Affiliations

¹ Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Pirogov Russian National Research Medical University, Moscow, Russia.
² Faculty of Biology, Lomonosov Moscow State University, Moscow, Russia.
³ Federal Research and Clinical Center, Federal Medical-Biological Agency of Russia, Moscow, Russia.
⁴ Mental Health Research Center, Moscow, Russia.
⁵ Pirogov Russian National Research Medical University, Moscow, Russia.
⁶ Department of Bioinformatics, Institute of Biomedical Chemistry, Moscow, Russia.
⁷ Department of Bioinformatics, Koltzov Institute of Developmental Biology, Russian Academy of Sciences, Moscow, Russia.
⁸ AcademGene LLC, Novosibirsk, Russia.
⁹ Moscow Institute of Physics and Technology (State University), Dolgoprudny, Russia.

Abstract

Introduction: Culturing of human neural stem cells (NSCs) derived from induced pluripotent stem cells (iPSC) is a promising area of research, as these cells have the potential to treat a wide range of neurological, neurodegenerative and psychiatric diseases. However, the development of optimal protocols for the production and long-term culturing of NSCs remains a challenge. One of the most important aspects of this problem is to determine the stability of NSCs during long-term in vitro passaging. To address this problem, our study was aimed at investigating the spontaneous differentiation profile in different iPSC-derived human NSCs cultures during long-term cultivation using.

Methods: Four different IPSC lines were used to generate NSC and spontaneously differentiated neural cultures using DUAL SMAD inhibition. These cells were analyzed at different passages using immunocytochemistry, qPCR, bulk transcriptomes and scRNA-seq.

Results: We found that various NSC lines generate significantly different spectrums of differentiated neural cells, which can also change significantly during long-term cultivation in vitro.

Discussion: Our results indicate that both internal (genetic and epigenetic) and external (conditions and duration of cultivation) factors influence the stability of NSCs. These results have important implications for the development of optimal NSCs culturing protocols and highlight the need to further investigate the factors influencing the stability of these cells in vitro.

Keywords: differentiation; iPSC; neural culture; neural stem cells; sc-RNA-seq.