| Abstract No.: | B-B2030 |
| Country: | Canada |
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| Title: | PLATELET DERIVED GROWTH FACTOR (PDGF) INDUCED ACTIVATION OF ERK1/2 IN OLIGODENDROCYTE PROGENITORS IS TIME AND CONCENTRATION DEPENDENT. |
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| Authors/Affiliations: | 1 Zhicheng Zhou; 3 Joumana Mustapha*; 2 Emma Frost;
1 Dept. Pathology and Manitoba Institute of Child Health, University of Manitoba, Winnipeg, MB, Canada; 2 Dept.s of Pathology; Human Anatomy and Cell Science; Biochemistry and Medical Genetics, Winnipeg, MB, Canada; and 3 MICH, University of Manitoba, Winnipeg, MB, Canada
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| Content: | OBJECTIVE: To clarify the role of PDGF induced intracellular signaling in the regulation of oligodendrocyte progenitor (OP) migration. INTRODUCTION: Oligodendrocytes are the myelinating cells of the CNS. In the developing brain, they originate as progenitors in the germinal matrix (GM). From there they migrate extensively to populate white matter tracts1. OP migration is induced in response to numerous local cues including soluble signaling proteins derived from neurons. One such factor, Platelet-derived growth factor (PDGF), is essential for the development of myelin2. OP response to PDGF differs according to the signaling pathway activated by receptor phosphorylation, i.e. migration is regulated via activation of the extracellular regulated kinase (ERK) signaling pathway, whereas proliferation is regulated via the PI3K signaling pathway3. We have recently shown that PDGF (10ng/ml) induced migration requires transient (<30 mins) exposure, and that this migration is sustained for up to 72 hours. This study furthers that finding by showing that low concentrations of PDGF activate ERK without inducing OP migration. MATERIALS AND METHODS: OPs were isolated from neonatal rat pups (P0-P1) as described previously4. ERK activation: PDGF induced ERK activation in OP was assessed by Western Blot analysis. Briefly, OPs were exposed to PDGF at 0.01-10ng/ml for periods from 5 mins to 2 hours. Cell lysates were electrophoresed on a 12% polyacrylamide gel. After electrotransfer to PVDH membrane, blots were blocked in 5% skim milk before being exposed to antibodies against phosphorylated ERK1/2, total ERK1/2 and GAPDH, a standard housekeeping protein. Bands were visualized using an HRP conjugated secondary antibody enhanced with ECL reagent and exposed to X-ray film Migration: The agarose drop assay was used to analyze the migration of OP after exposure to PDGF, as described previously5. RESULTS: All concentrations of PDGF at all durations of exposure activated ERK1/2 in OP. Whereas PDGF induced OP migration required either transient exposure (>10 minutes) to high concentrations (10ng/ml), or sustained exposure (<2 hours) to low concentrations (<1 ng/ml). CONCLUSION: We show OP migration is only activated at 1ng/ml PDGF (a physiologically relevant concentration), with continuous exposure. Whereas ERK phosphorylation occurs in response to all PDGF concentrations and durations this is supported by previous studies that show that duration of receptor occupancy determines the downstream effects of ligands. We hypothesize that OP migration requires a threshold level of ERK activation that is not achieved by exposing OPC to low concentrations of PDGF for short periods of time. In addition, we hypothesize that ERK activation alone is not sufficient to drive OP migration. This work was funded by the Manitoba Children’s Hospital Foundation.
(1) Levison, S. W.et al. Development (1993) 119, 611
(2) Fruttiger, M.et al. Development (1999) 126, 457
(3) Ebner, S.et al. J Neurosci Res (2000) 62, 336
(4) Armstrong, R. C. Methods (1998) 16, 282
(5) Frost, E. E.et al. In Extracellular Matrix Protocols (2000) 139, 265
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