Cell-based screen for agents useful for reducing neuronal demyelination or promoting neuronal remyelination
This invention is in the field of neurology. Specifically, the invention relates to the discovery and characterization of molecular components that play a role in neuronal demyelination or remyelination. In addition, the invention relates to the generation of an animal model that exhibits hypomyelination. The compositions and methods embodied in the present invention are particularly useful for drug screening and/or treatment of demyelination disorders.
1. An in vitro method of developing a biologically active agent that reduces neuronal demyelination comprising:
(a) contacting a candidate biologically active agent with an oligodendrocyte;
(b) detecting an altered expression of a GADD34 gene or GADD34 gene product of said oligodendrocyte relative to a control cell, wherein said GADD34 gene or GADD34 gene product mediates its biological effect through the pancreatic endoplasmic reticulum (ER) kinase gene (PERK) pathway; and
(c) selecting said candidate biologically active agent as a candidate that reduces neuronal demyelination, if the level of expression of said GADD34 gene or GADD34 gene product of said oligodendrocyte is decreased relative to said control cell,
wherein said level of said GADD34 gene or GADD34 gene product of said oligodendrocyte is measured by a level of eIF-2α phosphorylation of said oligodendrocyte,
wherein an increased level of eIF-2α phosphorylation of said oligodendrocyte is indicative of said candidate biologically active agent reducing neuronal demyelination.
2. The method of claim 1 , wherein said detecting step involves an immunoassay.
3. The method of claim 1 , wherein said detecting step involves a hybridization assay.
4. The method of claim 1 , wherein said biologically active agent is selected from the group consisting of an antisense oligonucleotide, peptide, an antibody, a liposome, a small interfering RNA, a small organic and an inorganic compound.
5. The method of claim 1 , wherein said oligodendrocyte is from a demyelinated lesion in a subject's central nervous system.
6. The method of claim 5 , wherein said subject is a transgenic animal.
7. The method of claim 6 , wherein said transgenic animal having (a) stably integrated into the genome of said animal a transgenic nucleotide sequence encoding interferon-gamma (INF-γ).
8. The method of claim 7 , wherein said transgenic animal having (a) stably integrated into the genome of said animal a transgenic nucleotide sequence encoding interferon-gamma (INF-γ), and (b) an altered expression of at least one other gene; wherein upon expression of said INF-γ, said animal exhibits a greater degree of demyelination relative to a transgenic animal having a stably integrated transgenic nucleotide sequence encoding interferon-gamma (INF-γ) as in (a), but lacking said altered expression of said at least one other gene.
9. The method of claim 8 , wherein said at least one other gene is correlated with endoplasmic reticulum stress.
10. The method of claim 5 , wherein said subject comprises a heterozygous knock-out of pancreatic ER kinase gene (PERK), and stably integrated into the genome of said animal a transgenic nucleotide sequence comprising interferon-gamma (INF-γ).