Final Activity Report Summary - NEUROTRAIN (Training and understanding neurodegenerative diseases)
The EST NEUROTRAIN project aimed to improve our knowledge of neurodegeneration eventually leading to prevention and therapy. During the project period the following outstanding results were achieved:
1. Parkinson's and Huntington's disease (PD, HD)
Cellular replacement therapy has been shown in principle in PD patients and in animal models. Efficient and lasting cell fate generation of dopaminergic progenitors from pluripotent cells is crucial for successfully taking cell replacement therapy into routine clinical use. Inducing this cell fate in embryonic stem cells by overexpression of transcription factors implicated in midbrain specification has shown promising results. Furthermore loss of the kinase PINK1 and the protease HtrA2/Omi was shown to impair mitochondrial function, leading to increased cellular stress levels, possibly contributing to neurodegeneration in PD. This coincided with an altered availability of OPA1, one of the large GTPases responsible for mitochondrial fusion, in cells devoid of HtrA2/Omi. Furthermore the maintenance of mitochondrial morphology was shown to be dependent on functionally active HtrA2/Omi. Moreover, overexpression of Duo was found to attenuate synphilin-1 induced inclusions by transforming cytoplasmic small aggregates into peri-nuclear aggregates. In time-lapse microscopy study, Sph1 inclusions were susceptible to be degraded while Duo was present. Furthermore HD-Duo transgenic mice were generated and certain lines were selected based on their mRNA expression.
2. Retinitis Pigmentosa (RP)
Inherited retinal degeneration (RP) is a major cause of blindness in the developed world. Using different pharmacological tools on in vitro retinal explant cultures a causal involvement of histone deacetylases in rd1 photoreceptor cell death could be shown. Similarly, it was found that genetic knock-out of the PARP1 gene reduced retinal degeneration in the rd1 mouse (model for RP) suggesting an important function of PARP1 during degeneration. Another objective was to identify candidate genes in the RP25 interval and screen for mutations in affected Spanish families. Mutations were identified in a cluster of uncharacterised gene transcripts. Through 5' and 3' RACE PCR analysis, the full length gene was revealed. It is the largest eye specific gene identified to date (over 2.0 MB). Homologs of the RP25 gene to Drosophila eys/eys-shut were identified, leading to the name of EYS and the protein name SPAM. Furthermore, it was found that after subretinal transplantation of neonatal retinal cells in a RP model, activated microglia abnormally expresses sialoadhesin, which could be of significance for the integration and long-term survival of retinal grafts. Further investigations on micro- and macroglial cells in models for RP showed also that upon activation they up-regulate the expression of metallothioneins, potential neuroprotectants.
RP is caused by RhodopsinP23H (RhP23H). Ectopic expression of mutant Rh1P37H (the equivalent of mammalian RhP23H) in Drosophila photoreceptor neurons (PNs) causes retinal degeneration and blindness. Recruitment of endogenous Rh1 into Rh1P37H aggregates was found to lead to a dramatic depletion of mature Rh1 and strongly accelerates PN degeneration. Decreasing the function of VCP was found to prevent degradation of Rh1 aggregates and strongly suppressed retinal degeneration and blindness.
3. Epilepsy
It was demonstrated that neuropeptide Y effectively modulates inhibitory and excitatory synaptic inputs to inhibitory interneurons in the hippocampal network, thereby altering their synaptic excitability and possibly influencing the way they react to epileptic seizures in the circuitry.
4. Role of endoplasmatic reticulum (ER)
Reversible fragmentation of neuronal ER, a previously unknown biological phenomenon, was characterised. Live cell imaging was implemented which ultimately lead to ER analysis in live animals. The phenomenon was proven to exist during massive depolarisation of neurons (as in stroke or epilepsy) and to be a NMDA receptor gated Ca2+ ion dependent process. Data suggest that ER fragmentation / fusion may occur in physiological conditions.
1. Parkinson's and Huntington's disease (PD, HD)
Cellular replacement therapy has been shown in principle in PD patients and in animal models. Efficient and lasting cell fate generation of dopaminergic progenitors from pluripotent cells is crucial for successfully taking cell replacement therapy into routine clinical use. Inducing this cell fate in embryonic stem cells by overexpression of transcription factors implicated in midbrain specification has shown promising results. Furthermore loss of the kinase PINK1 and the protease HtrA2/Omi was shown to impair mitochondrial function, leading to increased cellular stress levels, possibly contributing to neurodegeneration in PD. This coincided with an altered availability of OPA1, one of the large GTPases responsible for mitochondrial fusion, in cells devoid of HtrA2/Omi. Furthermore the maintenance of mitochondrial morphology was shown to be dependent on functionally active HtrA2/Omi. Moreover, overexpression of Duo was found to attenuate synphilin-1 induced inclusions by transforming cytoplasmic small aggregates into peri-nuclear aggregates. In time-lapse microscopy study, Sph1 inclusions were susceptible to be degraded while Duo was present. Furthermore HD-Duo transgenic mice were generated and certain lines were selected based on their mRNA expression.
2. Retinitis Pigmentosa (RP)
Inherited retinal degeneration (RP) is a major cause of blindness in the developed world. Using different pharmacological tools on in vitro retinal explant cultures a causal involvement of histone deacetylases in rd1 photoreceptor cell death could be shown. Similarly, it was found that genetic knock-out of the PARP1 gene reduced retinal degeneration in the rd1 mouse (model for RP) suggesting an important function of PARP1 during degeneration. Another objective was to identify candidate genes in the RP25 interval and screen for mutations in affected Spanish families. Mutations were identified in a cluster of uncharacterised gene transcripts. Through 5' and 3' RACE PCR analysis, the full length gene was revealed. It is the largest eye specific gene identified to date (over 2.0 MB). Homologs of the RP25 gene to Drosophila eys/eys-shut were identified, leading to the name of EYS and the protein name SPAM. Furthermore, it was found that after subretinal transplantation of neonatal retinal cells in a RP model, activated microglia abnormally expresses sialoadhesin, which could be of significance for the integration and long-term survival of retinal grafts. Further investigations on micro- and macroglial cells in models for RP showed also that upon activation they up-regulate the expression of metallothioneins, potential neuroprotectants.
RP is caused by RhodopsinP23H (RhP23H). Ectopic expression of mutant Rh1P37H (the equivalent of mammalian RhP23H) in Drosophila photoreceptor neurons (PNs) causes retinal degeneration and blindness. Recruitment of endogenous Rh1 into Rh1P37H aggregates was found to lead to a dramatic depletion of mature Rh1 and strongly accelerates PN degeneration. Decreasing the function of VCP was found to prevent degradation of Rh1 aggregates and strongly suppressed retinal degeneration and blindness.
3. Epilepsy
It was demonstrated that neuropeptide Y effectively modulates inhibitory and excitatory synaptic inputs to inhibitory interneurons in the hippocampal network, thereby altering their synaptic excitability and possibly influencing the way they react to epileptic seizures in the circuitry.
4. Role of endoplasmatic reticulum (ER)
Reversible fragmentation of neuronal ER, a previously unknown biological phenomenon, was characterised. Live cell imaging was implemented which ultimately lead to ER analysis in live animals. The phenomenon was proven to exist during massive depolarisation of neurons (as in stroke or epilepsy) and to be a NMDA receptor gated Ca2+ ion dependent process. Data suggest that ER fragmentation / fusion may occur in physiological conditions.