|
|
|
|
|
|
|
||||
| Home Help Feedback Subscriptions Archive Search Table of Contents | ||||
|
Fig. 4. LIF promotes, whereas BMP4 inhibits, a GFAP-expressing multipotential stem
cell fate and neuron production. (A) Experimental paradigm for the cytokine
induction, rGFAP promoter and FACS selection, and stem cell analysis of
GFAP+ cells. Neurospheres were expanded, infected with the
rGFAPp-EGFP retrovirus, further expanded, plated for 5 days to differentiate
with or without cytokines and then sorted on the basis of eGFP expression
(green). Sorted cells were plated in a neurosphere-forming assay to assess
self-renewal and subsequent spheres were plated for differentiation to assess
the ability to form neurons (red), astrocytes (purple) and oligodendrocytes
(blue). (B,G) In the neurosphere-forming assay, 2% of control GFAP+
cells self-renew (green indicates cells that are GFP positive). (C-E,G) LIF
and noggin treatments each increase the percentage of GFAP+ cells
that form neurospheres (green). LIF treatment also increases neurosphere size,
which is further increased by additional BMP inhibition. (F,G) BMP4 treatment
prevents the self-renewal of GFAP+ cells. (H) A single GFP-positive
neurosphere plated for differentiation gives rise to oligodendrocytes (O4,
blue), neurons (ß-tubulin, red) and astrocytes (GFAP, pink),
demonstrating multipotentiality. (I-M) GFP-expressing neurosphere populations
were dissociated to assess neuronal progeny. (I,M) GFAP-expressing (GFP
positive, green) cells produce few neurons (red). (J,M) Noggin treatment alone
does not significantly change neuron production. (K,M) LIF treatment
significantly increases the number of neurons generated. (L,M) LIF treatment
with BMP inhibition further increases neuron production. (G)
*P<0.05, **P<0.01; (M)
*P<0.01, **P<0.005 ANOVA. Scale
bars: 100 µm in B-F; 20 µm in H-L.
|
