Abstract
SummaryPostnatal brain neural stem and progenitor cells (NSPCs) cluster in anatomically inaccessible stem cell niches, such as the subependymal zone (SEZ). Here, we describe a method for the isolation of NSPCs from live animals, which we term “milking.” The intracerebroventricular injection of a release cocktail, containing neuraminidase, integrin-β1-blocking antibody, and fibroblast growth factor 2, induces the controlled flow of NSPCs in the cerebrospinal fluid, where they are collected via liquid biopsies. Isolated cells retain key in vivo self-renewal properties and their cell-type profile reflects the cell composition of their source area, while the function of the niche is sustained even 8 months post-milking. By changing the target area more caudally, we also isolate oligodendrocyte progenitor cells (OPCs) from the corpus callosum. This novel approach for sampling NSPCs and OPCs paves the way for performing longitudinal studies in experimental animals, for more in vivo relevant cell culture assays, and for future clinical neuro-regenerative applications.
Highlights
Tissue-specific stem cells (t-SCs) reside in mature tissues in order to maintain homeostasis and to contribute to regeneration, functions they can perform because they retain the cardinal properties of stem cells: self-renewal and the potential to generate multiple, tissue-specific, cell types
Release and collection of neural stem and progenitor cells (NSPCs) NSPCs of the subependymal zone (SEZ) are positioned adjacent to ependymal cells, with Neural stem cells (NSCs) remaining in direct contact with the cerebrospinal fluid (CSF) via intercalating mono-ciliated processes (Doetsch et al, 1999, 2002)
We designed a strategy to compromise the integrity of the lateral ventricle wall, thereby allowing NSPCs to enter the CSF and to be collected via liquid biopsies
Summary
Tissue-specific stem cells (t-SCs) reside in mature tissues in order to maintain homeostasis and to contribute to regeneration, functions they can perform because they retain the cardinal properties of stem cells: self-renewal and the potential to generate multiple, tissue-specific, cell types. Postnatal brain neural stem and progenitor cells (NSPCs) reside in anatomically restricted niches of the postnatal brain, such as the subependymal zone [(SEZ) known as the subventricular zone] of the lateral walls of the lateral ventricles and the subgranular zone of the dentate gyrus (Obernier and Alvarez-Buylla, 2019). Neural stem cells (NSCs) clustering in the SEZ generate transitamplifying progenitors that subsequently give rise to progeny of neuronal or oligodendroglial commitment (neuroblasts and oligodendroblasts, respectively) (Kazanis et al, 2017). Due to the scarcity of these cells, especially in the adult human brain (Coletti et al, 2018; Sanai et al, 2011), and their location in inaccessible areas, in vitro experimental work aiming at deciphering the properties of endogenous NSC populations has relied on the postmortem isolation of animal NSPCs (Pastrana et al, 2009) and their clinical use remains challenging, albeit of great importance
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