Objective/Rationale:
GDNF is a potent survival factor for dopamine (DA) neurons, effective in standard neurotoxin models of PD, but recent studies show that GDNF is unable to rescue DA neurons in the more disease-related alpha-synuclein model. In pre-clinical models we have found that this failure is due to a blockade of the GDNF signaling response induced by alpha-synuclein, linked to a down-regulation of the transcription factor Nurr1 and its targets, including the GDNF receptor Ret. Here, we want to investigate whether this signaling blockade occurs also in patients with PD, and to what extent markers of protein degradation and autophagy are affected as well.
Project Description:
In the tissue samples obtained from APDC we will study to what extent Nurr1 and its target genes - those involved in DA synthesis and release and those involved in GDNF signaling - are affected in patients with PD or incidental LDB patients, compared to healthy age-matched controls. In addition we will study to what extent the expression of the autophagic-lysosomal master regulator TFEB and its targets are affected in midbrain DA neurons from patients with PD and incidental LBD, relative to controls. The expression of these factors will be studied with a combination of immunohistochemistry and in situ hybridization histochemistry. Gene expression analyses will be performed in collaboration with Prof Thomas Perlmann at Karolinska Institutet in Stockholm.
Relevance to Diagnosis/Treatment of Parkinson’s Disease:
The failure of GDNF to exert any neuroprotection in alfa-synuclein models raises an important question: does alpha-synuclein-induced blockade of GDNF signaling occur also in patients with PD? And more specifically: does down-regulation of Nurr1 and its targets, including the GDNF receptor Ret, occur also in human PD? Previous studies have shown that Nurr1 expression is reduced in nigral neurons affected by increased levels of alpha-synuclein. However, the expression of other Nurr1 targets, including Ret, has not been yet examined in human postmortem material. In parallel, the analyses performed here will clarify to what extent defective protein degradation seen in PD may be driven by reduced expression of the recently identified master regulator of the autophagic-lysosomal system, TFEB.
Anticipated Outcome:
The results will provide new insights into the cellular defects induced by increased levels of alpha-synuclein, in particular the role of Nurr1 dysfunction and defective protein clearance in alpha-synuclein toxicity the pathogenesis of PD, and provide further support for Nurr1 and TFEB as interesting novel targets for neuroprotective and disease-modifying therapies.
Final Outcome
The principal aim of this project is to investigate the disease-related changes that develop in midbrain dopamine neurons in the brains of patients suffering from either Parkinson´s disease (PD) or the milder form called incidental Lewy-Body disease (ILBD). The access to sections from brains of diseased patients, supplied by the Arizona Parkinson´s Disease Consortium (APDC), has given us the opportunity to apply an advanced analytical technique called laser capture microdissection (LCM) in combination with single-cell analysis of changes in gene expression based on highly sensitive RNA sequencing. This approach, which has not been applied to postmortem PD or ILBD brains before, has the potential to give new insights into the pathophysiology of PD. Initially the project was delayed due to technical problems associated with the application of the laser-capture procedure to human postmortem sections, but these problems have now been solved. We have now obtained RNA sequencing data of very high quality from all 18 patients (6 PD, 6 ILBD and 6 controls) that is in the final stage of analysis.