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Alexandra M. Bandeian1, Maria Chiara G. Monaco2, and Eugene O. Major 2
1University of Michigan, Ann Arbor, MI; 2Laboratory of Molecular Medicine and Neuroscience, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892, USA
Differentiation of human neural progenitors into neuronal and glial cell types offers a model to study and compare molecular regulation of neural cell lineage development. In vitro expansion of neural progenitors from fetal central nervous system (CNS) tissue has been well characterized. Despite the identification and isolation of glial progenitors from adult human sub-cortical white matter and development of various culture conditions to direct differentiation of fetal neural progenitors into myelin producing oligodendrocytes, acquiring sufficient human oligodendrocytes for in vitro experimentation remains difficult. The need remains for a culture system to produce cells of the oligodendrocyte lineage suitable for in vitro experimentation.
Culture of primary human oligodendrocytes could, for example, be a useful model to study the pathogenesis of neurotropic infectious agents like the human polyomavirus, JCV. The JC virus (JCV) is a human polyomavirus that infects oligodendrocytes, which are myelin-producing cells in the central nervous system, resulting in the fatal demyelinating disease progressive multifocal leukoencephalopathy (PML) (Ferenczy, 2012). In vivo, oligodendrocytes are the most susceptible cells to infection while neural progenitors have very low levels of susceptibility. Because oligodendrocytes stem from progenitor cells, cellular events take place during differentiation, either on the cell surface or within the cell, that allow for higher rates of infection by JCV. There is no working animal model for studying JCV, so our lab has used human fetal brain cells for experimentation.
Neural progenitor cells were cultured in DMEM-F12 serum-free media supplemented with oligodendrocyte specific growth factors (Monaco, 2012). Using these conditions, the majority of the cells in culture maintain a morphology characterized by few processes and express markers of pre-oligodendrocyte cells, such as A2B5 and O-4. When we remove the four growth factors (GF) (bFGF, PDGF-AA, Shh, NT-3), the cells start to acquire more processes and express markers specific of oligodendrocyte differentiation, such as GalC and myelin basic protein (MBP). Phenotypic characterization was performed using video microscopy and immunofluorescence staining. Observing oligodendrocyte differentiation by tracking phenotypical changes of the cells can be very useful in finding the mode of infection and replication of the virus, which can ultimately help future JCV research.
Last Modified December 14, 2012