A single‐cell analysis reveals multiple roles of oligodendroglial lineage cells during post‐ischemic regeneration

Abstract

NG2 cells represent precursors of oligodendrocytes under physiological conditions; however, following cerebral ischemia they play an important role in glial scar formation. Here, we compared the expression profiles of oligodendroglial lineage cells, after focal cerebral ischemia (FCI) and in Alzheimer's-like pathology using transgenic mice, which enables genetic fate-mapping of Cspg4-positive NG2 cells and their progeny, based on the expression of red fluorescent protein tdTomato. tdTomato-positive cells possessed the expression profile of NG2 cells and oligodendrocytes; however, based on the expression of cell type-specific genes, we were able to distinguish between them. To shed light on the changes in the expression patterns caused by FCI, we employed self-organizing Kohonen maps, enabling the division of NG2 cells and oligodendrocytes into subpopulations based on similarities in the expression profiles of individual cells. We identified three subpopulations of NG2 cells emerging after FCI: proliferative; astrocyte-like and oligodendrocyte-like NG2 cells; such phenotypes were further confirmed by immunohistochemistry. Oligodendrocytes themselves formed four subpopulations, which reflected the process of oligodendrocytes maturation. Finally, we used 5-ethynyl-2' deoxyuridine (EdU) labeling to reveal that NG2 cells can differentiate directly into reactive astrocytes without preceding proliferation. In contrast, in Alzheimer's-like pathology we failed to identify these subpopulations. Collectively, here we identified several yet unknown differences between the expression profiles of NG2 cells and oligodendrocytes, and characterized specific genes contributing to oligodendrocyte maturation and phenotypical changes of NG2 cells after FCI. Moreover, our results suggest that, unlike in Alzheimer's-like pathology, NG2 cells acquire a multipotent phenotype following FCI.

NG2 Glia: Novel Roles beyond Re-/Myelination

“…To further increase the complexity of this scenario, consistent with previous findings [207,208], in vivo two-photon imaging analyses revealed an assortment of distinct NG2 glia reactions to injury, with some cells starting migration toward the lesion site, others entering cell cycle, and others displaying only hypertrophy and morphological changes [31,86]. Single-cell messenger-RNA (mRNA) profiling of NG2 glia after focal cerebral ischemia also identified at least three subpopulations of cells that could be distinguished according to changes in their expression patterns [19]. Such behavioral and molecular heterogeneity suggests the existence of diverse NG2 glia subsets which respond differently to CNS injury.…”

“…In line with this view, a subpopulation of NG2 glia has been observed to serve as multipotent progenitors, producing protoplasmic astrocytes as well OLs in the embryonic ventral forebrain or following some kinds of CNS injury [19,[24][25][26][27]. It has also been proposed that adult NG2 glia may activate spontaneous neurogenic events within restricted brain areas [28].…”

“…These observations suggest that in basal conditions such an abundant pool of resident NG2 glia may exert additional roles beyond their functions in oligodendrogenesis and myelin production. Although the molecular aspects of NG2 glia response to injury are far from being completely understood, data accumulated so far indicate that, phenotypically, NG2 glia provide a stereotypic reaction (i.e., increased NG2 expression, retraction of cell processes, cell body swelling, cell proliferation and migration toward the lesion site) to almost all kind of injury, independently of the extent of myelin loss [14,[16][17][18][19]. Further, upon experimental demyelination in the subcortical white matter, transient populations of NG2 glia coming from the subventricular zone (SVZ) of the lateral ventricles immediately amplify and invade the lesion site, but ultimately seem not to contribute to myelin repair, and are lost [20][21][22].…”

Oligomeric amyloid beta prevents myelination in a clusterin-dependent manner

“…While we describe the transcriptional profile of OPCs during homeostasis, it is important to note that understanding the role of OPCs in the healthy brain will provide a necessary foundation for examining any protective or detrimental novel functions in desease pathology. Previous work has demonstrated that OPCs exhibit significant transcriptional heterogeneity and disease-associated signatures in models of multiple sclerosis and cerebral ischemia, and showed significant transcriptional changes in human Alzheimer's disease patients 18,59,76 .…”

A Clonal NG2-Glia Cell Response in a Mouse Model of Multiple Sclerosis

“…This morphology is characterized by thickened and highly ramified processes, as opposed to the relatively thin and unbranched prolongations of non-reactive NG2-glia [17]. Furthermore, NG2-glia display both molecular and behavioral heterogeneity following brain injury [17,21,22], producing both beneficial and deleterious effects [8,21,[23][24][25][26][27]. However, little is known about the existence of certain subsets of NG2-glia that produce a specific response to brain damage and whether their fate may already be determined early in embryonic development.…”