GM-CSF acts synergistically with other proinflammatory cytokines to induce tissue damage in an in vitro model of the CNS
Diana Arseni1, Colin P. Glover2, Julia Edgar1, Chris Linington1,
1University of Glasgow; 2Medimmune Ltd., Granta Park, Cambridge. CB21 6GH
Granulocyte-macrophage colony stimulating factor (GM-CSF) plays multiple roles in the development, maintenance and regulation of the immune system (1). In experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (2), the absence of GM-CSF expression by CNS antigen-specific Th1/17 T cells abolishes their encephalitogenic potential suggesting that GM-CSF plays a non-redundant role in disease development, an effect attributed to the inability of GM-CSF-/- T cells to activate myeloid cells within the CNS (3). To address the mechanisms involved, we investigate the effects of GM-CSF in myelinated cultures derived from embryonic mouse spinal cord (4,5,6). Our data demonstrate GM-CSF is able to “activate” microglia in this culture system inducing marked changes in their morphology and enhancing their phagocytic potential. However, in isolation, this response is unable to damage myelin or axons. RNAseq analysis revealed these effects were associated with upregulation of a small set of immune related genes including Mgl2, Clec4n and a variety of chemokine and chemokine receptors (eg. Ccr2, Ccr1, Ccl12, Ccl6).
These data suggest GM-CSF synergises with other effector mechanisms to mediate tissue damage in EAE. We therefore investigated the ability of GM-CSF to potentiate the cytotoxic activity of IFN-γ and TNF-α in myelinated cultures. These experiments revealed a combinatory effect of all three cytokines was required to induce rapid and extensive loss of myelin and axons. This effect was associated with rapid induction of iNOS in microglia, accompanied by elevated NO levels, which was only observed when all three cytokines were present. Our data demonstrate GM-CSF acts synergistically to lower the threshold for IFN-γ and TNF-α to induce tissue damage.
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