Evidence for Heterogeneity of Astrocyte De-Differentiation in vitro: Astrocytes Transform into Intermediate Precursor Cells Following Induction of ACM from Scratch-Insulted Astrocytes

Abstract

Our previous study definitely demonstrated that the mature astrocytes could undergo a de-differentiation process and further transform into pluripotential neural stem cells (NSCs), which might well arise from the effect of diffusible factors released from scratch-insulted astrocytes. However, these neurospheres passaged from one neurosphere-derived from de-differentiated astrocytes possessed a completely distinct characteristic in the differentiation behavior, namely heterogeneity of differentiation. The heterogeneity in cell differentiation has become a crucial but elusive issue. In this study, we show that purified astrocytes could de-differentiate into intermediate precursor cells (IPCs) with addition of scratch-insulted astrocyte-conditioned medium (ACM) to the culture, which can express NG2 and A2B5, the IPCs markers. Apart from the number of NG2(+) and A2B5(+) cells, the percentage of proliferative cells as labeled with BrdU progressively increased with prolonged culture period ranging from 1 to 10 days. Meanwhile, the protein level of A2B5 in cells also increased significantly. These results revealed that not all astrocytes could de-differentiate fully into NSCs directly when induced by ACM, rather they generated intermediate or more restricted precursor cells that might undergo progressive de-differentiation to generate NSCs.

Cell transcriptional state alters genomic patterns of DNA double-strand break repair in human astrocytes

“…Previous investigations have demonstrated that activated neonatal mouse and rat astrocytes can be used to generate neurospheres 22,23 and are sensitized to neoplastic transformation after exposure to IR 41,42 . Furthermore, epidemiologic studies have established links between a variety of human malignancies and childhood exposure to ultraviolet and IR 43–45 .…”

“…Phenotypically, this state of activation is characterized by elevated expression of intermediate filaments 12 , increased secretion of cytokines and extracellular matrix components 13 , cell hypertrophy 14 , proliferation 15 and migration 16 . Activated astrocytes also express elevated levels of the progenitor marker nestin 17–19 , adopt a bipolar morphology 20,21 and can be used to generate neurospheres that are differentiable into neurons and oligodendrocytes 22–24 . Together, these features are suggestive of a more primitive state of differentiation and recapitulate some of the phenotypic attributes of glioma cells 11,25 .…”

Neural Stem Cell Properties of an Astrocyte Subpopulation Sorted by Sedimentation Field-Flow Fractionation

“…30,31 The emergence of neural stem-like cells in the cortex was associated with the presence of GFAP and vimentin expressing cells, two intermediate filaments found in radial glial-like cells and in adult neural stem cells. [32][33][34] The existence of an astrocyte subpopulation that can acquire stem cell characteristics after injury has been described after an invasive CNS lesion, such as stab wounding or cerebral ischemia, but not after diffuse noninvasive injury.…”

Real-Time PCR and Immunocytochemical Study of Chondroitin Sulfate Proteoglycans after Scratch Wounding in Cultured Astrocytes / PCR I IMUNOCITOHEMIJSKA STUDIJA EKSPRESIJE HONDROITIN-SULFATNIH PROTEOGLIKANA NAKON POVREDE ASTROCITA U KULTURI

“…One of such is a scratchwound assay (SW), first established as a simple, reproducible assay for the analysis of cell migration in vitro (13). The SW model was soon adopted by many to investigate different aspects of astroglial response to mechanical injury (14)(15)(16)(17). As a response to SW injury, the following characteristics of reactive astrocytes have been observed: hyperplasia, enhanced expression of extracellular matrix molecules and elongation of hypertrophic processes (18,19).…”