Kasti, M; (2013) Experimental Neuroinflammation: A Study of Hypoxia and Protein Translation. Doctoral thesis , UCL (University College London).

Abstract

Multiple sclerosis is a chronic inflammatory disease of the CNS associated with widespread primary demyelination and axonal degeneration. The mechanisms underlying the expression of neurological deficits are incompletely understood. Recent histological findings are consistent with a view that active MS lesions may be hypoxic (i.e. suffer a low oxygen concentration). For example, inflammatory ‘Pattern III’ MS lesions express hypoxia inducible factor-1a (HIF-1a), a master regulator of genes whose function is to bias a cell for survival under hypoxic conditions. Hypoxia also results in a number of other consequences designed to limit energy expenditure, including the inhibition of protein translation by the phosphorylation of the eukaryotic initiation factor (eIF-2a), and the formation of stress granules (SGs). Both hypoxia and the inhibition of protein synthesis could cause neurological deficits and thus contribute to the neurological deficits of MS. The aim of the present study is to explore experimental models of MS for evidence of a) hypoxia and b) inhibition of protein translation. The experimental models of inflammatory demyelinating lesions were induced either by the intraspinal injection of lipopolysaccharide (LPS) into the rat dorsal column (LPS-DC, focal lesion), or by immunization of rats or mice to induce experimental autoimmune encephalomyelitis (EAE, disseminated lesions). Animals were examined daily for the expression of any neurological deficit, and tissue hypoxia was detected during life by the systemic administration of pimonidazole, a marker for hypoxia, several hours before termination. Tissue was taken at different stages of lesion development (1-28 days post injection) and examined immunohistochemically for the presence of hypoxia, determined by the expression of binding for pimonidazole, and for the inhibition of protein translation by examining the expression of eIF-2a and SGs. Other markers of disease activity were also examined, including a marker of microglial/macrophage activation (ED-1), HIF-1a, and glucose transporter-1 (GLUT-1). In LPS lesions, labelling for pimonidazole was most intense at the site of injection, 24 hours later. In EAE, labelling for pimonidazole was present as early as 2 days post immunization, but it was expressed more strongly when animals were exhibiting a neurological deficit, subsiding thereafter. In animals injected with LPS, eIF-2a and SGs were expressed most intensely 24 hours post LPS injection, localised to the spinal motor neurons. In EAE, eIF-2a and SGs were expressed in spinal motor neurons, and in cerebellar neurons, at the onset of neurological deficits. These findings reveal for the first time that inflammatory demyelinating lesions are associated with the presence of tissue hypoxia and markers of the inhibition of protein synthesis. It appears that these phenomena may contribute to the expression of neurological deficits, opening new opportunities for therapy.

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