Short title:
Replacement of animal experiments in Parkinson’s research
Background:
Research in Parkinson’s disease is experiencing an upswing at the moment, on one hand due to a lack of curative drugs for the large number of patients, on the other hand due to the recent discovery of the PARK genes in families affected by the genetic form of the disease. Drug testing is nearly exclusively performed in vivo in the so-called MPP, methamphetamine and 6-hydroxydopamine models requiring tens of thousands of animals. These animal experiments impose medium to very severe stress on animals (depending on the protocol used), and employ animal species ranging from mice to rats and primates. Sometimes primary neuronal cultures of rats are used for mechanistic studies, but these are very difficult to handle. Human neurons, which would be most relevant, are not usually available, and neuroblastoma cells are only a very poor substitute.
Methods/approach:
We have developed a human neuronal model cell line (LUHMES) that may be applied for the study of Parkinson’s disease with respect to mechanisms of degeneration and mechanisms and efficacy of drugs. We want to establish these absolutely unique cells as standard model for Parkinson’s disease by validation against the established animal models. Moreover, we want to introduce the known human mutated PARK genes, which can then be tested in combination with the established environmental triggers of Parkinson’s disease (e.g. MPP) and largely substitute transgenic models for this purpose.
Study plan:
In a first stage, the expression of tyrosine hydroxylase in LUHMES cells will be augmented to make these cells as similar as possible to human brain cells in vivo. Secondly, an MPP model of Parkinson’s disease will be established in the cells and compared to the already functioning methamphetamine model. Thirdly, the models will be validated, using the currently known drugs that show efficacy in animal models. In a final step, mutated PARK genes, e.g. DJ-1 will be introduced into the cells and their interaction with MPP and methamphetamine will be tested. Data will continuously be made available to the scientific community to foster the use of the cells.
Relevance to 3R:
Animal models are the main tool for drug development and testing of mutated Parkinson genes. Our calculations show that around 100000 animals (used otherwise in severely stressful experiments) could be saved, even if our model would be used only in a percentage of cases as alternative in vitro method in Parkinson’s research. Research data will be immediately usable by others and create an immediate impact.
Abbreviations:
PD: Parkinson’s disease; PARK gene: gene found to be mutated in a familial form of Parkinson’s disease; MPP: 1-methyl-4-phenylpyridinium iodide.
Outcome:
Schildknecht S,
Poeltl D, Nagel DM, Matt F, Scholz D, Lotharius J, Salvo-Vargas A, Leist M (2009). Requirement of a dopaminergic neuronal
phenotype for toxicity of low concentrations of 1-methyl-4-phenylpyridinium to
human cells.
Toxicol Appl
Pharmacol,
241, 23-35
Stiegler N, Krug A,
Matt F, Leist M (2011) Assessment of
chemical-induced impairment of human neurite outgrowth by multiparametric live
cell imaging in high-density cultures. Toxicol
Sci 121: 73 – 87
Scholz D, Pöltl D,
Genewsky A, Weng M, Waldmann T, Schildknecht S, Leist M (2011) Rapid, complete and large-scale generation of
post-mitotic neurons from the human LUHMES cell line. J Neurochem 119, 957-971
Zimmer B,
Schildknecht S, Kuegler PB, Tanavde V, Kadereit S, Leist M (2011) Sensitivity of dopaminergic neuron differentiation
from stem cells to chronic low-dose methylmercury exposure. Toxicol Sci 121,
357 – 367
Pöltl D, Schildknecht
S, Karreman C, Leist M (2012). Uncoupling
of ATP-depletion and cell death in human dopaminergic neurons. NeuroToxicol 33, 769-779
Schoenenberger F,
Krug A, Leist M, Ferrando-May E,
Merhof D (2012). An advanced image processing approach based on parallel growth
and overlap handling to quantify neurite growth. TICSP (ISSN 1456-2774) 61 (Ninth Int.
Workshop Computational Systems Biology),
92-95.
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