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Fig. 8. Effects of PI 3-kinase inhibition by LY294002 on OBSC neuronal and astrocyte differentiation. OBSCs derived from E14.5 embryos were cultured for 48 hours in differentiation conditions in the presence or absence of 100 ng/ml IGF-I, and they were treated with 30 µM LY294002 for the last 24 hours. (A) P-Akt^Ser473 phosphorylation was strongly inhibited by LY294002, whereas the reduction of P-Akt^Thr308 was moderate. LY294002 produced a small effect on the levels of P-Erk1 (but not on P-Erk2) only in conditions of differentiation. One representative immunoblot out of three experiments is shown. (B) The addition of LY294002 significantly decreased the percentages of TuJ-1-positive cells in the presence of IGF-I and it significantly decreased the percentages of GFAP-positive cells in the presence of BSA and IGF-I (C). (D) Exposure to LY294002 produced a slight but significant increase in the number of TUNEL-positive cells, only in the presence of IGF-I. The results are the average ± s.e.m. from 4-6 cultures. Representative fields of neuronal (TuJ-1; E,F) and astrocyte (GFAP; G,H) morphologies are shown. The number of TUNEL-positive cells was only slightly higher in cultures treated with LY294002 (I,J). Statistical analysis was performed using Student's t test. (*P<0.05, **P<0.01.) Bar, 25 µm.

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