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"MECHANISM OF INSULIN-LIKE GROWTH FACTOR -1 MEDIATED PI3K/AKT/mTOR ACTIVATION IN OLIGODENDROCYTE PROGENITORS AND THE ROLE OF mTOR IN REMYELINATION"

by
Kedar Mahajan
MD/PhD Program
B.S. 2005, Rowan Univeristy, Glassboro, NJ



Thesis Advisor: Terri Wood, Ph.D.
Professor
Department of Neurology and Neurosciences
University Hospital Cancer Center

Wednesday, July 21, 2010
Cancer Center, G1196, 9:30 A.M.


Abstract

Insulin-like growth factor-1 (IGF-1) regulates developmental myelination by promoting oligodendroglial proliferation, survival, and myelin production both in vivo and in vitro. Our previous studies demonstrated that IGF-1 sustains PI3K/Akt phosphorylation in oligodendrocyte progenitor cells (OPCs) to promote proliferation and survival. Here, we investigate whether IGF-1 mediated phosphorylation of PI3K/Akt requires the presence of the IGF type 1 receptor (IGF-1R) in cholesterol enriched microdomains (CEMs; “lipid rafts” or “caveolae”) believed to cluster signaling components. In the absence of stimulation, the IGF-1R is detected in raft fractions and found directly to associate with caveolin-1, an integral structural component of caveolae. Upon acute IGF-1 stimulation, the IGF-1R is lost from raft fractions and its association with caveolin-1 decreases. These data suggest the IGF-1R is localized to caveolae and “primed” to bind ligand. We also report that acute stimulation of OPCs with physiological levels of IGF-1 (20 ng/mL; 2.6 nM) results in phosphorylation of the mammalian target of rapamycin (mTOR) and its downstream signaling targets S6 ribosomal protein (S6), elongation factor 4E binding protein 1 (4E-BP1), and Ser473-Akt. IGF-1 activation of the mTOR pathway in OPCs is sensitive to inhibition by the PI3K inhibitor (LY294002) but not the MEK1/2 inhibitor (U0126). Moreover, inhibition of the IGF-1R by a selective neutralizing antibody (IMC-A12) also attenuates activation of the mTOR pathway in these cells. While mTOR has been recently shown to promote the onset of OPC differentiation and myelination, its role during remyelination has yet to be established. To begin to characterize this role, focal demyelinated lesions were introduced with lysolecithin in the corpus callosum of young adult (8-12 weeks old) C57BL/6J mice. We report demyelination (MBP-/NF-H+ axons) and increased astrogliosis (GFAP+) from 2 to 10 weeks following lysolecithin in the demyelinated lesions as well as increased mTOR and S6 phosphorylation during the remyelination process at 2 weeks following lysolecithin. Accordingly, we conclude that IGF-1/IGF-1R signaling via CEMs in primary rat OPCs mediates PI3K/Akt activation, a key pathway implicated in proliferation and survival, and downstream PI3K/Akt/mTOR activation, implicated in OPC differentiation. We also consider the consequences of mTOR signaling during remyelination.


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