No description available.

Oligodendrocytes are cells located in the central nervous system (CNS) that are responsible for the production of the lipid rich membrane, myelin. Myelin and the process of making and wrapping myelin around an axon (also known as myelination) are critical for normal development since they ensure proper signal conduction in the vertebrate CNS. The loss or damage of this myelin, which is typically associated with the demyelinating disease multiple sclerosis (MS), is associated with improper axonal protection along with disrupted nerve signaling which can lead to a variety of different debilitating phenotypic responses. It has been shown that there are MS lesions in which oligodendrocyte progenitors are present. However, while these cells are thought to possess the intrinsic ability to myelinate, they do not efficiently mature and/or repair the myelin sheath within the MS lesion. The reasons for this block in differentiation are currently not fully understood. A critical and thorough understanding of oligodendrocyte ix development provides the foundation needed for future research to potentially provide therapeutic targets for stimulating proper maturation and efficient remyelination from the oligodendrocyte progenitors that are present within the MS brain. In the search for regulators of oligodendrocyte development and potential therapeutic targets, the data generated as part of my thesis provided evidence that CaMKII (more specifically CaMKIIβ) is a regulator of oligodendrocyte myelination and maturation. Using pharmacological inhibitors or siRNA-mediated knockdown of this protein resulted in improper formation of the oligodendrocyte process network. Interestingly, siRNA-mediated knockdown of CaMKIIβ appeared to play no noticeable role in the genetic regulation of specific oligodendrocyte developmental markers. Furthermore, an overall reduction of the thickness of the compact myelin was observed in the ventral spinal cord of CaMKIIβ knockout mice. These findings emphasize the importance of CaMKIIβ in oligodendrocyte myelination and maturation. To further investigate CaMKIIβ’s role in the regulation of CNS myelination, the effect of glutamate signaling on CaMKIIβ and in particular its actin binding site were assessed. These data showed that signaling via glutamate transporters promote an increase of process network in oligodendrocytes. This effect was associated with a transient increase in intracellular calcium concentration and a change in the phosphorylation of at least one serine residue present within CaMKIIβ’s actin binding site. Changes in phosphorylation of CaMKIIβ’s actin binding site suggested that CaMKIIβ detaches from filamentous F-actin and x allows for remodeling of the oligodendrocyte’s actin cytoskeleton. This was demonstrated by overexpressing CaMKIIβ actin binding mutant constructs to alter phosphorylation of serine residues to either always allow actin binding (CaMKIIβallA) or never allow actin binding (CaMKIIβallD). The overexpression of CaMKIIβallD alone demonstrated a decrease in the process network of oligodendrocytes and inhibited the effect of glutamate on the process network. In contrast, the overexpression of CaMKIIβallA and CaMKIIβWT alone showed normal process network formation along with a significant increase in the process network after stimulation of glutamate. The above data strongly suggest that there is a significant relationship between sodium dependent glutamate transporters/CaMKIIβ activation and the oligodendrocyte cytoskeleton in the role of regulation of oligodendrocyte differentiation and CNS myelination. The data presented in this dissertation provides overwhelming evidence that CaMKIIβ plays a significant role in the proper formation of the oligodendrocyte complex process network and myelination. CaMKIIβ’s relationship with glutamate and the actin cytoskeleton could lay the foundation for future research not only for the signaling of oligodendrocyte process formation and remyelination but also for future targets for MS therapies.

Charcot-Marie-Tooth disease (CMT) is a cohort of human hereditary disorders of the peripheral nervous system (PNS) which exhibit symptoms like sensory dysfunction, muscle weakness and gait disturbances. Different mutations are described as causation for this neuropathy, such as a duplication of chromosome 17 comprising the gene for the peripheral myelin protein-22 (PMP22). Based on different animal models former studies identified immune cells, i.e. macrophages and T-lymphocytes, as crucial mediators of pathology in these neuropathies. In this study, PMP22-overexpressing mice (PMP22tg, C61), serving as a model for a specific type of CMT – CMT1A – were crossbred with immune-deficient mutant mice to examine the impact of the immune system on nerve pathology. Crossbreeding of PMP22tg mice with recombination activating gene-1 (RAG-1) deficient mice, lacking mature T- and B-lymphocytes, caused no striking alterations of pathogenesis in peripheral nerves of mutant mice. In contrast, crossbreeding of PMP22tg myelin mutants with mice deficient in the chemokine monocyte chemoattractant protein-1 (MCP-1, CCL2) caused an amelioration of the demyelinating phenotype of peripheral nerves when MCP-1 was either reduced or completely absent. Furthermore, functional investigations, i.e. neurographic recordings and examinations of the grip strength of the extremities, revealed an amelioration in PMP22tg/MCP-1-/- mice in regard to a symptomatic improvement in the compound action muscle potential (CMAP) and stronger grip strength of the hindlimbs. Interestingly, peripheral nerves of PMP22tg mice showed an irregular distribution of potassium channels in presence of MCP-1, whereas the absence of MCP-1 in the myelin mutants rescued the ion channel distribution and resulted in a more wild type-like phenotype. Having shown the impact of MCP-1 as an important mediator of nerve pathology in PMP22/MCP-1 double mutants, the regulation of this chemokine became an important target for potential treatment strategies. We found that the signaling cascade MEK1/2/ERK1/2 was more strongly activated in peripheral nerves of PMP22tg mice compared to nerves of wild type mice. This activation corresponded to an increase in MCP-1 mRNA expression in peripheral nerves at the same age. Furthermore, a MEK1/2-inhibitor was used in vivo to confirm the regulation of MCP-1 by the MEK1/2/ERK1/2 pathway. After a treatment period of three weeks, a clear reduction of ERK1/2-phosphorylation as well as a reduction of MCP-1 mRNA expression was observed, accompanied by a decline in macrophage number in peripheral nerves of PMP22tg mice. These observations suggest that the expression of MCP-1 is crucial for the neuropathological progression in a mouse model for CMT1A. Therefore, this chemokine could provide a basis for a putative treatment strategy of inherited neuropathies. / Die Charcot-Marie-Tooth Erkrankungen (CMT) sind eine Gruppe von humanen, erblichen Erkrankungen des peripheren Nervensystems (PNS), welche Symptome wie sensible Störungen, Muskelschwäche und Gangstörungen verursachen können. Verschiedene Mutationen, z.B. eine Duplikation des Chromosoms 17, welches das Gen für das periphere Myelinprotein-22 (PMP22) enthält, sind als Ursache für diese Neuropathie beschrieben. Anhand verschiedener Tiermodelle wurde in früheren Studien gezeigt, dass Immunzellen, insbesondere Makrophagen und T-Lymphozyten, maßgeblich an der Pathogenese dieser Neuropathien beteiligt sind. In der vorliegenden Studie wurden PMP22-überexprimierende Mäuse (PMP22tg, C61) als Modell einer spezifischen CMT-Form – CMT1A – mit immun-defizienten Mutanten verkreuzt, um die modulierende Rolle des Immunsystems innerhalb der Pathogenese peripherer Nerven untersuchen zu können. Die Verkreuzung von PMP22tg Mäusen mit „recombination activating gene-1“-defizienten Mutanten (RAG-1-/-), die keine reifen T- und B-Lymphozyten besitzen, resultierte in keiner deutlich veränderten Pathologie der peripheren Nerven. Im Gegensatz hierzu führte die Verkreuzung der Myelinmutanten mit Mäusen, defizient für das Chemokin „monocyte chemoattractant protein-1“ (MCP-1), zu einer Abschwächung des demyelinisierenden Phänotyps in peripheren Nerven, wenn MCP-1 reduziert war oder völlig fehlte. Funktionelle Analysen, wie elektrophysiologische Messungen und Untersuchungen der Kraft in den Extremitäten, zeigten zudem in PMP22tg/MCP-1-/- Mäusen eine symptomatische Verbesserung, was sich in einer höheren Amplitude (compound muscle action potential, CMAP) und einer erhöhten Kraft in den Hinterpfoten der Mäuse widerspiegelte. Interessanterweise zeigten periphere Nerven der PMP22tg Mäuse eine abnorme Verteilung von Kalium-Kanälen, wohingegen das Fehlen von MCP-1 in den Myelinmutanten zu einer Verteilung dieser Ionenkanäle führte, die ähnlich zu Wildtyp-Mäusen war. Da MCP-1 in den PMP22/MCP-1 Doppelmutanten einen deutlichen Einfluss auf die Pathogenese aufwies, wurde die Regulation dieses Chemokins im Hinblick auf mögliche Therapie-Ansätze untersucht. Diese Untersuchung zeigte, dass die MEK1/2/ERK1/2-Signalkaskade in peripheren Nerven von PMP22tg Mäusen stärker aktiviert wird als in Nerven von Wildtyp-Tieren. Die Aktivierung dieser Signalkaskade ging dabei mit einer erhöhten MCP-1 mRNA Expression in peripheren Nerven von Tieren des gleichen Alters einher. Ergänzend wurde ein MEK1/2-Inhibitor in vivo verwendet, um die Regulation von MCP-1 durch die MEK1/2/ERK1/2 Kaskade zu bestätigen. Nach einer Behandlungszeit von drei Wochen wurde eine deutliche Reduktion der ERK1/2-Phosphorylierung, sowie eine Reduktion der MCP-1 mRNA Expression und eine geringere Makrophagen-Anzahl in peripheren Nerven von PMP22tg Mäusen detektiert. Diese Untersuchungen zeigen, dass die Expression von MCP-1 entscheidend für den neuropathologischen Verlauf in einem Mausmodell für CMT1A ist. Somit bietet dieses Chemokin eine Basis für die Entwicklung neuer Behandlungsstrategien peripherer Neuropathien.

published_or_final_version / abstract / toc / Biochemistry / Master / Master of Philosophy

No description available.

Correct myelin formation and maintenance is essential for normal functioning and is affected in the demyelinating disease, Multiple Sclerosis (MS). To better understand this disease and identify important targets in promoting remyelination, the study of developmental myelination is important. Galanin, a 29 amino acid neuropeptide has been identified as a potentially important modulator in early myelin development. In our Galanin transgenic mouse model, myelin basic protein (MBP) levels are highly elevated at postnatal day 10 compared to the wild type. A preliminary investigation of Galanin’s behavior at various doses in vitro, yielded results that agreed with Galanin’s effect in vivo. Proteolipid protein (PLP) was highly elevated in the 10nM dose in vitro indicating Galanin exerts its effects in a time and dose dependent manner. Overall, this study identifies Galanin as a potentially important modulator of developmental myelination that may become a therapeutic target in future studies of demyelinating diseases.

Univ., Diss.--Würzburg, 1999.

Thesis (M. Phil.)--University of Hong Kong, 2005. / Title proper from title frame. Also available in printed format.

No description available.

No description available.