Phase contrast images and immunocytochemical analysis
of differentiating
MLPCs in a 2D environment. (A) Undifferentiated MLPCs (Und) exhibited
fibroblast morphology. Cells after 15 days in differentiation medium
without NE, forskolin (For), or K252a retained their fibroblast morphology.
Process formation was visible in the medium without growth factors
PDGF-AA, EGF, and bFGF (PEF). Refractile cell bodies and an increased
level of process formation were observed in the presence of all factors
indicated above for days 15–20. Cells lost their multipolar
morphology and became bipolar or spindle-shaped after approximately
20–24 days. The scale bars are 100 μm (20× and 40×
magnification). (B) Immunocytochemical analysis of differentiating
MLPCs in a 2D environment. The untreated MLPCs showed negative staining
for A2B5 and faint staining for O4. After differentiating for 15 days,
cells exhibited positive staining for A2B5 and O4, characteristics
of immature oligodendrocyte precursor cells. The scale bars are 100
μm (rows 1 and 2, 20× magnification; row 3, 40× magnification).
(C) MLPCs do not differentiate into committed oligodendrocytes in
a 2D environment. After differentiation for 8 days, 72.4% of cells
were positive for A2B5 and 69.9% for O4, and after 15 days, 70.3%
of MLPCs exhibited positive staining for A2B5 and 69.7% for O4. After
20 days, 35.0% of cells remained A2B5 positive and 49.7% O4 positive.
Expression of O1 galactocerebroside and MBP was absent in both untreated
and differentiating cells. Error bars represent the SD.