Neural Stem Cells: Source, Culture, Identification, Differentiation

Sunday, September 17, 2023 Leave a comment

Neural stem cells (NSCs) are primitive cells with self-renewal and multi-differentiation potential that are preserved during the development of the nervous system. With the deepening of the understanding of neural stem cells, the prospect, and value of their clinical application have gained the attention of researchers, and they are one of the hotspots of life science research nowadays.

Bio-Sample Sources

It has been demonstrated that NSCs can be isolated from various parts of the embryonic nervous system as well as from adult mammals.

NSCs isolated from human embryos and the adult CNS share the same proliferation kinetics, differentiate into the three major nervous system cell types, and express the same family of regulatory genes. However, NSCs from different individual sources have their own unique gene expression and differentiation potential.
In the study of mammalian embryonic NSCs, stem cells have been isolated from the cerebral cortex, hippocampus, striatum, olfactory bulb, ventricles along the ventricles including lateral ventricles, the third and fourth ventricles, the mesencephalon, midbrain, cerebellum, spinal cord, and the retina and spinal cord.

Culture Systems

Over the past several decades, several culture systems have been developed that attempt to recapitulate the distinct in vivo developmental stages of the nervous system, enabling the isolation and expansion of different NSC populations at different stages of development.

Serum-free culture system

Purified embryonic stem cells are cultured into neural progenitor cells, which then form cell spheres. Theoretically, this system is suitable for studying the mechanisms of early neurogenesis. Stem cells grow in serum-free culture in a suspended spherical shape without wall attachment or protrusion, and then differentiate into mature neuronal cells by wall attachment after changing the culture medium.

Markers for Identification

For in vitro in situ characterization of NSCs, the most widely used methods are intermediate neurofilament protein (Nestin) and Musashi, an RNA-binding protein, as well as Vimentin, glial cell marker (GFAP), CD133, and other methods.

Markers	Description
Nestin	Nestin belongs to class IV intermediate filament proteins and is expressed only in the neuroepithelium of the early embryo and ceases to be expressed after birth. Nestin ceases to be expressed when neural precursor cells differentiate in a terminal direction into neurons and glial cells. Therefore, Nestin is widely used for the identification of NSCs.
Musashi	Musashi is an RNA-binding protein that is selectively expressed in NSCs/progenitor cells of various mammals. And it plays an important role in maintaining stem cell status and differentiation.
Vimentin	Vimentin is a waveform protein that belongs to class III intermediate filament proteins. The expression of vimentin is relatively early, starting after neural migration. It is also used as a marker for neural progenitor cells because its expression declines after the completion of differentiation.
CD133	CD133 is a cell surface antigen. Sorted CD133-positive embryonic brain cells can proliferate to form cell spheres, whereas CD133-negative cells cannot form cell spheres.

Differentiation

Currently, most researchers agree that there are two mechanisms for the differentiation of NSCs, cellular gene regulation, and exogenous signaling.

Cellular gene regulation
Cellular gene regulation refers to the regulation of the development of NSCs by their transcription factors and other functional proteins. Many transcription factors are involved in the proliferation and differentiation of NSCs. They are activated by a certain pathway at a specific time, which causes or shuts down the expression of downstream genes and ultimately determines the fate of NSCs.
Exogenous signaling
Exogenous signaling refers to the regulation of NSC development by the microenvironment, including cytokines and microenvironment. Currently, it is widely believed that mitogenic signals such as epidermal growth factor (EGF) and basic fibroblast growth factor (FGF2) play an important role in the proliferation and differentiation of NSCs, and can maintain the self-renewal ability of NSCs.

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