Elevated alpha-synuclein mRNA levels in individual UV-laser-microdissected dopaminergic substantia nigra neurons in idiopathic Parkinson's disease.
The presynaptic protein alpha-synuclein is involved in several neurodegenerative diseases, including Parkinson's disease (PD). In rare familial forms of PD, causal mutations (PARK1) as well as multiplications (PARK4) of the alpha-synuclein gene have been identified. In sporadic, idiopathic PD, abnormal accumulation and deposition of alpha-synuclein might also cause degeneration of dopaminergic midbrain neurons, the clinically most relevant neuronal population in PD. Thus, cell-specific quantification of alpha-synuclein expression-levels in dopaminergic neurons from idiopathic PD patients in comparison to controls would provide essential information about contributions of alpha-synuclein to the etiology of PD. However, a number of previous studies addressing this question at the tissue-level yielded varying results regarding alpha-synuclein expression. To increase specificity, we developed a cell-specific approach for mRNA quantification that also took into account the important issue of variable RNA integrities of the individual human postmortem brain samples. We demonstrate that PCR -amplicon size can confound quantitative gene-expression analysis, in particular of partly degraded RNA. By combining optimized UV-laser microdissection- and quantitative RT-PCR-techniques with suitable PCR assays, we detected significantly elevated alpha-synuclein mRNA levels in individual, surviving neuromelanin- and tyrosine hydroxylase-positive substantia nigra dopaminergic neurons from idiopathic PD brains compared to controls. These results strengthen the pathophysiologic role of transcriptional dysregulation of the alpha-synuclein gene in sporadic PD.
Gründemann
Jan
J
Molecular Neurobiology, Department of Physiology, Philipps-University Marburg, Deutschhausstrasse 2, 35037 Marburg, Germany.
Schlaudraff
Falk
F
Haeckel
Olga
O
Liss
Birgit
B
D000368:Aged; D000818:Animals; D004298:Dopamine / Q000096:biosynthesis; D006801:Humans; D007834:Lasers; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D042282:Microdissection / Q000379:methods*; D009474:Neurons / Q000737:chemistry / Q000378:metabolism*; D010300:Parkinson Disease / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013378:Substantia Nigra / Q000166:cytology / Q000378:metabolism* / Q000473:pathology; D014158:Transcription, Genetic; D014446:Tyrosine 3-Monooxygenase / Q000032:analysis; D014466:Ultraviolet Rays; D015854:Up-Regulation; D051844:alpha-Synuclein / Q000096:biosynthesis / Q000235:genetics*
D012333:RNA, Messenger; D051844:alpha-Synuclein; D014446:Tyrosine 3-Monooxygenase; D004298:Dopamine