Project

Parkinson's disease; Nogo-A; neuronal transplantation; neuronal phenotype; neurotrophic factors; dopaminergic neurons; signal transduction

Lead
Lay summary
Background Parkinson’s disease (PD) is a neurodegenerative disorder of unknown etiology. The disease is characterized by major clinical disturbances such as resting tremor, bradykinesia and rigidity. The symptoms are caused by dopamine depletion in the striatum resulting from a progressive loss of the nigrostriatal dopamine producing (DAergic) neurons. While medication therapy provides adequate alleviation of the symptoms for several years, the long term treatment is complicated by progressive disability and development of severe side effects. Metabolic compromise, excitotoxicity and oxidative stress are considered the main mechanisms that cause neuronal cell death that is both of apoptotic and necrotic nature. Notably, a marked atrophy of dendrites and spines of striatal medium spiny neurons is observed in advanced PD indicating a dysfunctional signal integration in the striatofugal pathway. A therapeutic concept for long-term treatment of PD is the replacement of the degenerated DAergic neurons by transplanting cells releasing dopamine. Recent findings from our and other laboratories suggest that neuroprotective molecules and neurotrophic factors support the survival of neural DAergic cells and the engraftment of the transplant in the host brain. The myelin associated protein Nogo-A is one of the most potent growth inhibitors of the adult CNS. Nogo-A is expressed on the surface of oligodendrocytes and has been described to limit neuronal regeneration and plasticity. Importantly to note, Nogo-A is also expressed in subpopulations of neurons of the adult and developing CNS. Several findings hint to the idea that Nogo-A has also functions in addition to prevention of axonal regeneration and plasticity. So accumulating body of data demonstrated that Nogo-A has also a neuronal differentiation inhibitor function. Given that Nogo-A is expressed in different populations of neurons in the adult CNS and modulates the cell differentiation, further studies are needed to better understand the function and the expression of this protein in health and disease. Aim and Experimental Design At present only few studies have investigated the expression of Nogo-A and NgR1 in the nigrostriatal system. Our preliminary data gathered by means of double immunofluorescence demonstrated that a subpopulation of DAergic cells also express Nogo-A. We do not know, however, on the extent of colocalization of DAergic neurons with Nogo-A and/or Nogo-A receptor (NgR1) and on the DAergic cell subtype. Accordingly, our research group will assess on the expression pattern of Nogo-A and NgR1 in the nigrostriatal system of the rat brain during development and in the PD disease state. Furthermore, the effects of Nogo-A neutralization in rat brain cultures and characterization of the interactions with neurotrophic factors are foreseen. Finally, the potential of Nogo-A neutralization of donor tissue prior to transplantation will be studied on the outcome in a rat model of Parkinson’s disease. Significance We anticipate that the results of these experiments will yield new insights regarding the fundamental pathophysiological features in PD and will also form a cornerstone for improved cell replacement strategies for patients suffering from Parkinson’s disease.

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Last update: 21.02.2013

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1. BACKGROUNDParkinson's disease (PD) is a disabling neurodegenerative disorder of unknown etiology characterized by a predominant and progressive loss of dopaminergic (DAergic) neurons of midbrain. While medication therapy provides adequate alleviation of the symptoms for several years, the long term treatment is complicated by progressive disability and development of severe side effects. A therapeutic concept for long-term treatment of PD is the replacement of the degenerated dopaminergic neurons by transplanting cells releasing dopamine. Clinical experiences have revealed proof of principle but also demonstrated significant limitations which ask for optimization of protocols. The major problems for cell replacement strategies in PD are concerning the appropriate source of donor tissue, the poor survival of transplanted cells and the suboptimal DAergic innervation of the host striatum.2. WORKING HYPOTHESISRecent findings from our and other laboratories suggest that neuroprotective molecules and neurotrophic factors support the survival of neural dopaminergic cells and the engraftment of the transplant in the host brain. Accumulating evidence indicate that Nogo-A plays an essential role in modulating the processes of axonal growth and innervation. Therefore, the combination of the beneficial effects of neurotrophic factors, neuroprotective molecules and Nogo-A inhibition might open new scenarios for PD therapies. In the present research project we will explore if the transplantation approach based on intracerebral application of anti-Nogo-A antibodies and treatment with creatine and/or neurotrophins significantly ameliorate the outcome of current cell transplantations protocols in PD. 3. SPECIFIC AIMSAim I.Assessment of the expression pattern of Nogo-A and NgR1 in the nigrostriatal system of the rat brain during development and in the PD disease state.Aim II.Assessment of the effects of Nogo-A neutralization in fetal rat VM cultures and characterization of the interactions with neurotrophic factors.Aim III.Assessment of the effects of Nogo-A neutralization on VM GFP transplant function in the 6-OHDA rat model of PD. 4. EXPERIMENTAL DESIGN AND/OR METHODS Distribution of Nogo-A and NgR1 in the nigrostriatal system of rat and human brain will be assessed using immunohistochemistry. By means of in vitro models of PD the effects of Nogo-A neutralization will be assessed using immunohistochemistry followed by analyzes of morphology. HPLC will be used to measure levels of dopamine in cultures. Identification of interactions with neurotrophic factors and creatine will include Western blot analyzes, signal transduction assessments and PCR. Using the 6-OHDA rat model of PD, we will explore Nogo-A neutralization on VM GFP transplants function of rats by means of behavioral assessment, immunohistochemistry and Western blot analysis. Phenotype identification of surviving neurons in grafts with or without factor pretreatment will be studied in the same setting using immunohistochemical methods in combination with measures of dopamine levels in the host brain. The possible alterations of host brain cell proliferation and neurogenesis will similarly be investigated in the 6-OHDA rat model of PD by means of BrdU injections and histological analyses.5. EXPECTED VALUE OF THE PROPOSED PROJECTThe proposed experiments are expected to contribute to the development of improved strategies and to increase understanding of events after transplantation in cell replacement approaches in PD. They concern a highly actual field of neurotransplantation research. Relevance to our studies is furthermore given that the used antibodies are currently in clinical evaluation. The impact of our approach likewise holds up for neural transplantation using alternative tissue sources and to other CNS lesions.