Investigation of myelin membrane adhesion and compaction in the central nervous system

Abstract

Myelin is a multi-layered membrane which enwraps the axons in peripheral (PNS) and central nervous system (CNS). The formation and assembly of this structure is a multi-step process that is regulated by a variety of extracellular factors. In the CNS, myelin is produced by oligodendrocytes. During development, the progenitors of these cells differentiate into mature oligodendrocytes that start to enwrap axons by myelin membrane sheaths after receiving the appropriate signal(s) from the microenvironment. However, the responsible signals to initiate this process are unknown. Here, we showed that oligodendrocytes release small microvesicles, exosomes, into the extracellular space that prevent the terminal differentiation of oligodendrocytes and subsequently myelin formation. These inhibitory effects were revealed to be mediated by activity of the RhoA-ROCK signalling cascade. Importantly, the exosome release by oligodendrocytes was significantly reduced when cells were incubated with the conditioned medium from neurons. Our results suggest that exosomes produced by oligodendrocytes maintain the cells in pre-myelinating stage, whereas in the presence of neuronal signals, exosomes secretion by oligodendrocytes is reduced and the autoinhibitory signals are relieved. Thus neurons may regulate the formation and release of oligodendroglial-derived exosomes in order to coordinate myelin membrane biogenesis and assembly. In the second part of the thesis, the question of how myelin compaction is mediated was addressed. Whereas MBP is known to organize the interaction between myelin membranes from cytoplasmic side, the molecular mechanisms underlying the interaction between the outer leaflets still remain unclear. In general, the interaction between two opposite membranes requires the expression of adhesion molecules and the removal of repulsive components. Therefore, we investigated the role of proteolipid protein (PLP), as a putative adhesive molecule, and the glycocalyx, as a repulsive structure, during myelin compaction in the CNS. We analyzed the adhesion of purified myelin particles with the primary oligodendrocytes in order to mimic the interaction between myelin layers. Using this system we showed that PLP increases the adhesiveness of myelin membrane. We also found that PLP enhances physical stability of myelin using single particle force spectrometry. In addition, we observed a significant reduction in the glycocalyx during oligodendrocyte maturation which correlated with an increase in their surface affinity towards myelin particles. Further analysis indicated that the negative charge of sugar moieties, mainly sialic acid, is responsible for the reduction in myelin adhesiveness. Therefore, we propose that the adhesive properties of PLP along with the reduction of the glycocalyx, orchestrate myelin membrane adhesion and compaction in the CNS.