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Glycomics of mammalian aging

Boek - Dissertatie

N-glycosylation is the mirror of the status of the cell. N-glycosylation changes in relation to cellular activation, embryonic development, organogenesis and differentiation have been thoroughly studied. But little is known about the involvement of N-glycosylation in the aging process. Therefore, we analyzed how the N-glycosylation status of serum proteins is altered during aging in humans. We questioned whether N-glycosylation of serum proteins could be a suitable aging biomarker because no effective and easy to measure parameter for physiological age has yet been found (see I.3).After studying age-related N-glycan alterations in humans, we investigated whether this phenomenon also occurs in other species, such as mice. If alterations in N-glycosylation in mice can also be used as an aging biomarker, different anti-aging interventions can be monitored in mice by determining the N-glycosylation status. An advantage of using mice is the ability to control their housing environment. To determine whether age-related glycosylation alteration is inherent to the organism, we can study whether mice isolated from environmental influences (i.e. in specific pathogen free conditions; SPF) also exhibit the same N-glycosylation changes. Another advantage is that wile the influence of genetic background cannot be controlled in humans, genetically identical inbred mouse strains are available. Therefore, we analyzed the age-related N-glycosylation alterations in genetically identical mice, namely C57BL/6, housed in SPF conditions. Moreover, to further confirm the age-related nature of the N-glycosylation changes, the N-glycosylation status was analyzed in short-lived and long-lived mice (see I.1.6.2).Next, we explored the mechanism behind these N-glycosylation alterations. Are N-glycosylation changes the cause of aging (since N-glycosylation regulates important pathways in the organism) or are they the consequence of aging? We hypothesized that structural changes in N-glycosylation could be caused by alterations in the biosynthesis pathway, degradation in the blood, changes in clearance capacity, or a combination of all.Age-related alteration in gene expression might affect the synthesis of N-glycans in the ER and Golgi. Some glycosyltransferases and glycosidases might be up or down regulated, which could lead to incomplete or aberrant N-glycan structures. If the structure of a certain glycan changes with age, it could be easy to predict which gene is responsible for the fluctuation because biosynthesis of glycans occurs through the consecutive actions of well defined glycosyltransferases. Therefore, gene expression levels and protein levels of different glyco-genes were measured. If a certain glyco-gene is suspected of being involved, a transgenic mouse overexpressing that glyco-gene can be made and studied. Also the regulation of interesting glyco-genes can be analyzed by promoter studies. We also investigated if there is a link between these glyco-genes and essential “aging” molecules, e.g. GH and IGF-I (see I.1.5.3).Another mechanism that could explain the observed N-glycosylation differences is degradation. While circulating in the blood, the N-glycan of an N-glycoprotein can be attacked by glycosidases. These enzymes are normally present in the lysosomes but some leakage could occur. To tackle this possibility, we measured the amounts and activities of glycosidases in serum.A third mechanism that was investigated is clearance. Proteins in the blood are cleared mostly by the kidneys and the liver. Proteins smaller than 45 kDa are cleared by the kidneys but most serum N-glycoproteins are larger. These glycoproteins can be cleared from the circulation by lectins that recognize a certain glycan structure (see I.2.3.3). We explored the possibility that clearance capacity is decreased during the aging process. The gene expression level of different clearance receptors was measured and an in vivo clearance tests was performed.Finally, we aimed to investigate possible longevity of candidate long-living mice. Studies performed in lower organisms have shown that overexpressing chaperones extends lifespan (see I.1.5.4). We analyzed the N-glycan profile of chaperone-overexpressing mice to see if we could find any retardation of the age-related N-glycosylation changes that could predict the extension of life in these mice. We also followed the lifespan of these mice and investigated gene expression levels and protein activity of candidate molecules.
Aantal pagina's: 1
Jaar van publicatie:2009