Concurrent isolation and characterization of oligodendrocytes, microglia and astrocytes from adult human spinal cord

Abstract

A cellular preparation of highly enriched oligodendrocytes was obtained from adult human spinal cord by Percoll gradient centrifugation followed by either differential adhesion or fluorescence-activated cell sorting after immunostaining with an antibody against galactocerebroside (Ol). The adherent and O1-negative cell fractions were ⪢96% microglia. The non-adherent and O1-positive fractions were ⪢96% positive for the oligodendrocyte markers O4 and O1, 0–2% positive for glial fibrillary acidic protein, and were devoid of neuronal or microglial markers. If the oligodendrocyte fraction was co-cultured with purified dissociated rat dorsal root ganglion neurons, the oligodendrocytes adhered to the axons and their numbers increased over a 4 week period. However, myelin sheaths were not produced around axons in these cultures. In contrast, if the oligodendrocyte cell fraction was grown alone in culture for ⪢3 weeks, the number of oligodendrocytes decreased and a layer of astrocytes developed underneath the oligodendrocytes. The oligodendrocytes could be eliminated from these cultures by subsequent passaging, thus producing cultures of pure astrocytes. The astrocytes accumulated both K+ and glutamate with kinetic properties similar to those reported for rodent astrocytes. We suggest that these astrocytes arose in part from an O4/O1-positive precursor which did not initially express glial fibrillary acidic protein. These results define a relatively simple method by which highly enriched populations of oligodendrocytes, astrocytes and microglia can be obtained from adult human spinal cord.