Mixed primary culture and clonal analysis provide evidence that NG2 proteoglycan-expressing cells after spinal cord injury are glial progenitors.
NG2(+) cells in the adult rat spinal cord proliferate after spinal cord injury (SCI) and are postulated to differentiate into mature glia to replace some of those lost to injury. To further study these putative endogenous precursors, tissue at 3 days after SCI or from uninjured adults was dissociated, myelin partially removed and replicate cultures grown in serum-containing or serum-free medium with or without growth factors for up to 7 days in vitro (DIV). Cell yield after SCI was 5-6 times higher than from the normal adult. Most cells were OX42(+) microglia/macrophages but there were also more than twice the normal number of NG2(+) cells. Most of these coexpressed A2B5 or nestin, as would be expected for glial progenitors. Few cells initially expressed mature astrocyte (GFAP) or oligodendrocyte (CC1) markers, but more did at 7 DIV, suggesting differentiation of glial precursors in vitro. To test the hypothesis that NG2(+) cells after SCI express progenitor-like properties, we prepared free-floating sphere and single cell cultures from purified suspension of NG2(+) cells from injured spinal cord. We found that sphere cultures could be passaged in free-floating subcultures, and upon attachment the spheres clonally derived from an acutely purified single cell differentiated into oligodendrocytes and rarely astrocytes. Taken together, these data support the hypothesis that SCI stimulates proliferation of NG2(+) cells that are glial progenitor cells. Better understanding the intrinsic properties of the NG2(+) cells stimulated by SCI may permit future therapeutic manipulations to improve recovery after SCI.
Yoo
Soonmoon
S
Department of Neuroscience, Georgetown University Medical Center, Washington, DC 20057, USA.
Wrathall
Jean R
JR
D000818:Animals; D000941:Antigens / Q000378:metabolism*; D001253:Astrocytes / Q000166:cytology / Q000378:metabolism; D015415:Biomarkers / Q000032:analysis / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D019070:Cell Lineage / Q000502:physiology; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D002999:Clone Cells / Q000166:cytology / Q000378:metabolism; D018920:Coculture Techniques; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D005260:Female; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology; D007381:Intermediate Filament Proteins / Q000378:metabolism; D017628:Microglia / Q000166:cytology / Q000378:metabolism; D009416:Nerve Regeneration / Q000502:physiology*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D064231:Nestin; D009457:Neuroglia / Q000166:cytology / Q000378:metabolism*; D009836:Oligodendroglia / Q000166:cytology / Q000378:metabolism; D011509:Proteoglycans / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D020127:Recovery of Function / Q000502:physiology; D013119:Spinal Cord Injuries / Q000378:metabolism / Q000503:physiopathology*; D013234:Stem Cells / Q000166:cytology / Q000378:metabolism*; D000099216:Chondroitin Sulfate Proteoglycan 4
D000941:Antigens; D015415:Biomarkers; D016895:Culture Media, Serum-Free; D036341:Intercellular Signaling Peptides and Proteins; D007381:Intermediate Filament Proteins; D009419:Nerve Tissue Proteins; D064231:Nestin; D011509:Proteoglycans; D000099216:Chondroitin Sulfate Proteoglycan 4; C577486:Nes protein, rat