|Art der Förderung:||Standard Projekt|
|Institution:||Philipps-Universität Marburg, Abteilung für Neurologie|
|Projektleiter:||Prof. Dr. Richard Dodel|
|Laufzeit:||01. November 2000 - 31. Oktober 2002|
Beim Fortschreiten der Alzheimer-Erkrankung wird der Aktivierung des Immunsystems eine bedeutende Rolle zugewiesen. Das Immunsystem aktiviert und steuert Entzündungs-reaktionen im Körper. Die wichtigsten Bestandteile der Entzündungsreaktion im Blut wie im Gehirn stellen die Interleukine, eine Gruppe von Immunfaktoren, dar. Im Gehirn von Alzheimer-Patienten werden verschiedene Interleukine in den Amyloid-Ablagerungen nachgewiesen.
Es bestehen Hinweise, dass erhöhte Mengen von Interleukinen im Gehirn das Absterben von Nervenzellen verstärken. In ersten Untersuchungen konnte Dr. Richard Dodel von der Philipps-Universität in Marburg nachweisen, dass eine genetische Veränderung des Interleukin-1a-Gens das Risiko für die Entstehung der Alzheimer-Krankheit erhöht.
In seinem Forschungsprojekt wird Dr. Dodel den Zusammenhang zwischen der Mutation des Interleukin-1a-Gens und der Entstehung von Alzheimer weiter untersuchen. Zusätzlich soll der Einfluss der Genveränderung auf den Gehalt von Interleukinen im Blut und in der Gehirnflüssigkeit von Alzheimer-Patienten erforscht werden. Die Ergebnisse werden zur Entwicklung von neuen diagnostischen Methoden und zur Erarbeitung neuartiger therapeutischer Konzepte beitragen, mit denen die Entzündungsreaktion im Gehirn von Alzheimer-Patienten vermindert werden kann.
During the first year of grant support we have started with all proposed projects. The results are listed in conjunction with their proposed aims as outlined in the summary section of our grant application. In addition, we have started a new project with the title „Naturally occurring antibodies against Abeta and their role in the pathogenesis of Alzheimer's disease (AD)"
1. The first aim would be to test whether these preliminary results could be verified in a larger cohort (>700 individuals) of AD patients and controls. The data suggest that this polymorphism, whose odds ratio to develop AD increases with the gene dose (i.e. having one or two alleles), may be as important as apolipoprotein E. Apolipoprotein E is the major riskfactor known to date for AD.
We have finished the genotyping of IL-1A (-889) in a large cohort of AD patients (n=259) and controls (192) from three different centres. The number of patients was reduced according to power calculations. We found an increased risk for AD for heterozygous carriers and more pronounced for individuals homozygous for IL-1A(889) allele. No evidence was established for an interaction between the IL-1A and the apoE 4 polymorphisms (carriers and homozygotes), age, or gender with regard to conferred risk. The data strongly support an association between the IL-1A (-889) allele 2, especially in homozygotes, and later-onset AD.
In an additional sample, which consists of autopsy-proven familial AD patients, we investigated whether the polymorphism IL-1A (-889) has an increased risk for AD. We determined the prevalence of this polymorphism in 319 individuals with late onset autopsy- confirmed familial AD or non-demented individuals in these families from the Indiana Alzheimer Disease Center. Our results showed that l4% of 100 neuropathologically confirmed late onset AD patients were homozygous for allele 2 of IL-1A compared to 5.9% of 219 non-demented individuals with a family history of AD. Our data support an association between allele2 of IL-1a, especially homozygous individuals, and AD risk in familial AD. The results were published as an article and abstract (Du et al., 2001; Du et al., 2002). An additional paper is submitted.
We investigated whether the increased risk of the polymorphism IL-1A(-889) to develop AD is also associated with an increased risk for other neurodegenerative diseases, including Parkinson's disease (PD). The observed risk to develop AD seems not to be true in patients with PD. In our study, no significant difference was found in the risk for carriers of the polymorphism, as compared with controls. In addition, we did not see an age-dependent difference, as was postulated for a polymorphism in the IL-1ß gene in PD. The results from this study demonstrate that the IL-1A (-889) polymorphism is not a risk factor for patients with PD and that different pathogenetic mechanisms may be involved in the neurodegenerative process associated with PD and AD. The results were published as an article (Dodel et al., 2001)
2. Mutations or polymorphisms in the regulatory region of a gene are known to change the expression levels of that gene. Therefore, we hypothesize that the mutation in the regulatory region of IL-1α may also lead to a change in the concentration of IL-1 or other interleukins, who are related to IL-1. To characterize the effect of having different alleles (wildtyp, heterozygous, homozygous) we would investigate the concentration of IL-1α the cerebrospinal fluid (CSF) and the blood from individuals who have none (wildtyp), one (heterozygous), or two (heterozygous) alleles. In addition, we would investigate the concentration of other interleukins, like IL-β, IL-6, IL-1 Receptor, and IL-10. As neuronal or glial cells are not available for further characterization of the interleukin response to pathogens, we will use isolated mononuclear cells from the blood to further characterize the influence of being a carrier of these polymorphisms. Cells will be activated by lipopolysaccharides (which are parts of the bacterial membrane, which induce an intense immune reaction) and the release of the forementioned interleukins will be investigated. We hypothesize that the release of IL-1α and probably other interleukins are different depending of the allele carrier status of the donor.
Using ELISA technique we have not found a significant difference in IL-1 levels in patients with AD who are carrier of the IL-IA (-889) compared to AD patients, who lack this polymorphism and normal controls. In collaboration with the Dept. of Neurology Indiana University Medical School (Alzheimer Research Center of Excellence) we included 38 patients with AD carrying the polymorphism and 50 AD patients lacking the polymorphisms and 60 normal controls. No significant difference was found in these three samples. From the 38 AD patients and IL-IA (-889) carriers we purified PBMC and challenged them with different pro-inflammatory proteins (LPS, IL-6, etc.) and measured IL-1 in the medium. These experiments are currently ongoing. No results are yet available.
3. In the last year several mutations in different genes of the interleukin family have been linked to an increased or decreased risk to develop AD (e.g. IL-6). Therefore, we would investigate in the large cohort available to us (including more than 700 patients and controls), whether these results can be reproduced in our study. In addition, we would investigate several other polymorphisms in genes of the interleukin family for their risk to develop AD.
The immunoregulatory IL-10 downregulates synthesis of pro-inflammatory cytokines such as IL-1. Recent evidence suggests that some polymorphisms in the IL-10 promoter may have functional effects on IL-1 secretion. 406 AD patients and 251 unrelated non-demented control were investigated for the presence of three polymorphisms on IL-10 promoter (-1087A/G, 824Cn, -597C/A). No significant differences in the allelic distribution of the analyzed IL-10 polymorphisms have been found between AD and controls. We conclude that polymorphisms in the IL-10 promoter do not increase the risk to develop AD. Preliminary results are available.
4. Finally, we want to investigate how and by which mechanisms these polymorphisms influence the gene expression. The specific regions of the gene of different carriers will be cloned into different cells and the expression levels will be measured. The results from these experiments should give a more clear-cut evidence of the functional effects of these polymorphisms.
In this project we investigated the effect on expression of IL-1α in transfected cells which carry different IL-l polymorphisms, including IL-1α (-889), IL-1α (+4845). Cells were transiently transfected using cationic liposomes using a 1426 bp fragment covering the IL-1α 5'flanking sequences. Luciferase reporter plasmid pGl2-Basic (Promega) was used in a reporter gene assay to examine IL-1promoter activity. The experiments are currently on-going. No results have been obtained, yet.
Dodel, R., Lohmuller, F., Du, Y., Eastwood, B., Gocke, P., Oertel, W. & Gasser, T. (2001).
A polymorphism in the intronic region of the IL-1alpha gene and the risk for Parkinson's disease. Neurology, 56:982-983.
Du, Y., Dodel, R., Hampel, H., Buerger, K., Lin, S., Eastwood, B., Bales, K., Gao, F., Moeller, H.-J., Oertel, W., Farlow, M. and Paul, S. (2001). Reduced levels of amyloid beta-peptide antibody in Alzheimer disease. Neurology, 57:801-805.
Du, Y., Dodel, R. C., Eastwood, B. J., Bales, K. R., Gao, F., Lohmuller, F., Muller, U., Kurz, A., Zimmer, R., Evans, R. M., Hake, A., Gasser, T., Oertel, W. H., Griffin, W. S. T., Paul, S. M. and Farlow, M. R. (2000). Association of an interleukin 1alpha polymorphism with Alzheimer's disease. Neurology, 55:480-484.