Date of Degree

2010

Document Type

PhD diss.

Degree Name

PhD (Doctor of Philosophy)

Department

Immunology

First Advisor

Brian K. Martin

Abstract

Demyelination in the CNS is known to involve several immune effector mechanisms, including complement proteins. For this dissertation project the central hypothesis that C3 and downstream effector complement proteins exacerbate demyelination through activation of glial cells was tested. To investigate the role of C3 and downstream complement proteins in demyelination and remyelination pathology in vivo we utilized the cuprizone model. We used C3 knockout mice (C3-/-), which are lacking the central C3 protein and subsequently all downstream complement effector proteins, and transgenic mice expressing C3a or C5a under the control of the glial GFAP promoter. Interestingly, we found no changes in demyelination or remyelination pathology between C3-/- and control mice. However, C3a and C5a transgenic mice had exacerbated demyelination and slightly delayed remyelination in the corpus callosum compared to WT mice. Transgenic mice had increased cellularity in the corpus callosum due to increased activation and/or migration of microglia. There was also evidence of T cells in the corpus callosum during demyelination in C5a transgenic mice, suggesting C5a may modulate BBB permeability. During early remyelination oligodendrocytes migrated to the corpus callosum in higher numbers in C3a and C5a transgenic mice, thus enabling these mice to remyelinate as effectively as WT mice by the end of the ten week study.

To determine the effects of anaphylatoxins on individual glial subsets, we created murine recombinant C3a and C5a proteins. We found that the MAPK pathway proteins JNK1 and ERK1/2 were activated in glia upon stimulation with recombinant anaphylatoxin proteins. When microglia and mixed glial cultures were stimulated with C3a and/or C5a, we observed an increase in the production of proinflammatory cytokines and chemokines. In contrast, anaphylatoxin-treated primary astrocytes had suppressed cytokine and chemokine production compared to untreated astrocytes. In vitro, primary microglia and astrocytes did not significantly migrate in response to stimulation with C3a or C5a proteins, suggesting migration may not be a primary anaphylatoxin-mediated function in the CNS. Overall, our findings show that anaphylatoxin production in the brain plays a negative proinflammatory role during demyelination and that anaphylatoxin proteins can activate individual subsets of glia, initiating the production of inflammatory mediators.

Pages

xi, 142

Bibliography

128-142

Copyright

Copyright 2010 Sarah A. Ingersoll