Der bidirektionale Zusammenhang zwischen Sulfatiden und den molekularen Mechanismen der Alzheimer-Erkrankung

Abstract

Die Alzheimer-Erkrankung ist eine neurodegenerative Erkrankung die durch Ablagerung von extrazellulären senilen Plaques gekennzeichnet ist. Der Hauptbestandteil dieser Plaques wird durch das Peptid Amyloid-β (Aβ) gebildet, welches durch die sequentielle Proteolyse des Amyloiden Vorläufer Proteins (APP) entsteht. Sulfatide gehören zur Gruppe der Sphingolipide und sind ein wichtiger Membranbestandteil insbesondere der Lipid Rafts des Zentralennervensystems. In einigen Studien konnte gezeigt werden, dass der Sulfatid-Gehalt im Gehirn von Alzheimer-Patienten bereits in frühen Stadien der Erkrankung reduziert ist. Die molekularen Mechanismen der wechselseitigen Beeinflussung von Sulfatiden und der Alzheimer-Erkrankung näher zu untersuchen war Ziel dieser Arbeit. Es konnte sowohl im Zellkulturmodell als auch im Mausmodell gezeigt werden, dass durch die intrazelluläre Domäne von APP (AICD) die Sulfatid-Synthese in Folge einer reduzierten Genexpression des Enzyms Cerebrosid-Sulfotransferase gehemmt wird. Umgekehrt führten Sulfatide im Zellkulturmodell zu einer reduzierten Bildung von Aβ in Folge einer gesteigerten Degradation der β-Sekretase und einer Inhibition der g-Sekretase insbesondere innerhalb der Lipid-Rafts. Zuletzt zeigte sich auch eine Reduktion der Aβ-Aggregation unter Zugabe von Sulfatiden. Es ließ sich somit ein bidirektionaler Zusammenhang zwischen Sulfatiden und der APP Prozessierung darstellen, welcher im Falle der Alzheimer Erkrankung zu einem Teufelskreis führen könnte.

Bidirectional link between sulfatides and the molecular mechanisms of Alzheimer’s disease Alzheimer's disease is a neurodegenerative disease characterized by the deposition of extracellular senile plaques. The main component of these plaques is amyloid-β (Aβ) peptide, which is formed by the se-quential proteolysis of the amyloid precursor protein (APP) (MASTERS et al., 1985). Two different processing pathways exist. In the amyloidogenic processing β-secretase BACE1 starts the cleavage, whereas the non-amyloidogenic pathway is initiated by the α-secretase (ZHANG et al., 2012). The mul-tienzyme complex of the γ-secretase then catalyzes the subsequent proteolysis of the remaining mem-brane fragments. During the amyloidogenic processing Aβ-peptides of different lengths are formed by the γ-secretase, while the non-amyloidogenic processing prevents the formation of Aβ (DULIN et al., 2008). In both cases, the APP-intracellular domain (AICD) is released to the cytosol. APP and its pro-cessing products, particularly AICD, are believed to have important physiological functions beside their relevance in Alzheimer's disease (LI et al., 1997; ROBINSON et al., 2014). In this context AICD is discussed to influence transcriptional gene regulation (LAUER et al., 2020). Both, APP and the former mentioned secretases, are transmembrane proteins, suggesting that APP processing is influenced by lipid composition of the cellular membranes. This has already been proofed for some lipids (GRIMM et al., 2011b; PUGLIELLI et al., 2003). Conversely, the processing products of APP also affect the metabo-lism of various lipid classes like cholesterol or sphingolipids (GRIMM et al., 2005; GRIMM et al., 2011a). Sulfatides belong to the group of sphingolipids and are important membrane components, especially in the lipid rafts of the central nervous system (BLOMQVIST et al., 2002). Sulfatide synthesis is mediated by the enzymes ceramide galactosyltransferase (CGT) and cerebroside sulfotransferase (CST), the latter catalyzing the rate-limiting step of the sulfatide synthesis (HIRAHARA et al., 2000). Several studies reported that sulfatide content in the brain of Alzheimer's patients is already reduced in the early stages of the disease (CHENG et al., 2013; HAN et al., 2002; GÓNZALEZ DE SAN ROMÁN et al., 2017). Nevertheless, it remains unclear whether this is cause or consequence of Alzheimer's disease. Investi-gating this controversy was one of the aims of this project. This work first investigated various cell culture models with deficiency for the APP protein family or presenilin. An increase in sulfatide synthesis, sulfatide content and gene expression of the involved en-zymes CGT and CST was detected. Using various methods, these changes could be attributed to the effect of AICD. Increased AICD level, through the addition of synthetic AICD as well as overexpression of the APP fragment, led to a reduced CST gene expression. Moreover, promoter assays revealed a decrease of CST promoter activity in the presence of AICD. Similar findings were also obtained when analyzing murine brain tissue, pointing at the great the in vivo relevance and thus indicating that sulfatide synthesis is directly influenced by APP-processing. On the other hand, we observed that, through an inhibition of the amyloidogenic processing, sulfatides reduce the Aβ level in cell culture experiments, whereas the non-amyloidogenic pathway and Aβ deg-radation remained unaffected. Mechanistically, sulfatides decreased BACE1 protein levels and β-secre-tase activity by accelerating enzyme degradation and reducing endosomal BACE1. Furthermore, sul-fatides inhibited γ-secretase activity directly as well as indirectly by shifting the secretase from raft regions to non-raft regions. The effect of sulfatides on β- and γ-secretase was also confirmed in vivo in murine brain tissue. In addition, elevated sulfatide level caused a delay in Aβ42 aggregation and reduc-tion of toxic oligomers in vitro. In conclusion, the loss of cerebral suldatides in Alzheimer's disease could result in enhanced amyloido-genic processing and Aβ42 aggregation and consequently accelerate the disease pathophysiological pro-cesses. The increased formation of AICD could further promote the loss of sulfatides, resulting in a vicious circle. Raising cerebral sulfatide concentration in Alzheimer's patients therefore represents a promising therapeutic option.