Effects of transgenic overexpression of polysialyltransferase (PST) on myelination and impacts of increased sulfatide accumulation in arylsulfatase A deficient mice and its relevance to human metachromatic leukodystrophy

Abstract

The process of oligodendrocyte differentiation and myelination involves downregulation, biosynthesis and degradation of various lipids, proteins and carbohydrates. Alterations in these metabolic pathways can lead to neuropathological symptoms and demyelinating diseases like Metachromatic leukodystrophy (MLD). This study aims at addressing the roles of two molecules namely (a) polysialic acid (PSA), an oligosaccharide and (b) sulfatide, a glycosphingolipid, in the process of oligodendrocyte differentiation, myelination and in demyelinating disorder, MLD.<br /> Oligodendrocyte precursor cells express neural cell adhesion molecule (NCAM) which is post-translationally modified by addition of polysialic acid (PSA). As these cells undergo differentiation, they downregulate the synthesis of PSA. To understand whether downregulation of PSA is an essential criteria for oligodendrocyte differentiation and myelination, transgenic mice overexpressing polysialyltransferase, PST, under the control of proteolipid protein (PLP) promoter was generated, so that, these mice do not downregulate polysialylation and show an increased level of PSA on NCAM-120 in mature oligodendrocytes. The transgenic mice sustained PSA modification of NCAM-120 in all developmental stages. These mice displayed partial co-localization of PSA and myelin basic protein (MBP) with localization of PSA-NCAM in myelin. Sustained PSA expression resulted in a decrease in myelin content and MBP protein. Structural analysis of these mice showed normal myelination in the central nervous system (CNS) and peripheral nervous system (PNS), but adult mice displayed structural abnormalities like axonal degeneration and redundant myelin. In situ hybridization indicated a reduction in the number of differentiated oligodendrocytes in the forebrain with no reduction in the number of oligodendrocyte precursor cells. Analysis of primary oligodendrocyte cultures displayed a less differentiated morphology with fewer processes and membranous extensions without any changes in the localization of NCAM and myelin associated glycoprotein (MAG) and with no significant difference in the percentage of MBP positive cells. Altogether these observations indicate that downregulation of PSA is essential for myelination but is not the only prerequisite for efficient myelin formation and that PSA is also involved in myelin maintenance and stability.<br /> Lipids play an important role in maintaining membrane stoichiometry as well as in the process of myelination. De-regulation in lipid degradation could result in disorders like MLD. MLD is a lysosomal sulfatide storage disorder characterized by accumulation of sulfatide in the nervous system and other organs. The disease is caused by deficiency in the sulfatide degrading enzyme arylsulfatase A (ASA). MLD patients suffer from neurological symptoms and progressive demyelination. ASA(-/-) mice, the only animal model of MLD, in part displayed some, but less severe symptoms and no demyelination like MLD patients, thus resembling the earlier stages of human MLD. Since sulfatide storage is critical for MLD disease manifestations, it was hypothesized that increased sulfatide storage in ASA(-/-) mice might display the neuropathological symptoms of the disease. Hence, in order to increase sulfatide storage in ASA(-/-) mice, transgenic/ASA(-/-) [tg/ASA(-/-)] mice overexpressing galactosylceramide sulfotransferase (CST) under the control of PLP promoter was generated and analyzed. These tg/ASA(-/-) mice displayed a significant increase in sulfatide, in both, central and peripheral nervous system. Animals older than one year displayed severe neurological defects like gait disturbances, hind limb paralysis and decreased nerve conduction velocity. Demyelination and hypertrophic neuropathy of peripheral nerves was prevalent in these mice, along with demyelination of the central nervous system. Biochemical analysis showed a decrease in MBP. Altogether these observations indicate that, increasing sulfatide storage resulted in neuropathological symptoms and demyelination akin to the advanced stages of MLD.