Development of O4+/O1- immunopanned pro-oligodendroglia in vitro.
In this study, O4+/O1- pro-oligodendroglia isolated by immunopanning from cerebral hemispheres of P3-P5 rats were evaluated during their maturation in culture. Immunopanning yielded 3-4 x 10(5) cells/cerebrum, with 98% O4+ and 6% O1+. There was heterogeneity in the morphologies of immunopanned cells ranging from simple bipolar cells to more complex multipolar cells. As a first step in determining potential differentiative responses of mature oligodendroglia, we examined glial fibrillary acidic protein (GFAP) expression in response to fetal bovine serum (FBS) by cultures established from O4+/O1- immunopanned cells grown for 1, 14, or 21 days, exposed to 20% FBS for 6-7 days and fixed and immunostained on days 7, 21 or 28 in culture (DIC). When immunopanned cells were exposed to FBS following 1 day in serum-free medium, 88% expressed GFAP and when immunopanned cells were cultured for 14 days prior to FBS exposure, 78% expressed GFAP. By contrast, when cells were cultured for 21 days prior to FBS exposure (when a majority of the cells expressed O1 and myelin basic protein (MBP)), only 19% of the cells expressed GFAP (p < 0.001). Cells that were O4+/GFAP- even in the presence of FBS often exhibited a mature oligodendroglial morphology. Among immunopanned cells that responded to FBS by expression of GFAP, both process-bearing (similar to type 2 astroglia) and flattened, polygonal (similar to type 1 astroglia) GFAP+ cells were observed. These results confirm the utility of immunopanning for the isolation of pro-oligodendroglia and demonstrate that oligodendroglia that develop in vitro from O4+/O1- immunopanned cells become resistant to GFAP induction by FBS.
Ingraham
C A
CA
Department of Psychiatry, Albany Medical College, NY 12208, USA. christine_ingraham@ccgateway.amc.edu
Rising
L J
LJ
Morihisa
J M
JM
D000818:Animals; D000831:Animals, Newborn; D000911:Antibodies, Monoclonal; D002417:Cattle / Q000097:blood; D002478:Cells, Cultured; D016922:Cellular Senescence / Q000502:physiology; D005312:Fetal Blood / Q000502:physiology; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D007158:Immunologic Techniques; D008858:Microscopy, Phase-Contrast; D004676:Myelin Basic Protein / Q000378:metabolism; D009836:Oligodendroglia / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013234:Stem Cells / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D013997:Time Factors