Author(s): Knarik Arkun, Ann C Rice and James P Bennett
Objective: Our objective was to evaluate mitochondrial DNA (mtDNA) gene copy numbers in substantia nigra neurons from post-mortem Parkinson’s Disease cases and determine if the presence of Lewy bodies (LBs) altered mtDNA copy numbers or changed apparent deletion of mtDNAs. Methods: We used laser capture micro dissection to isolate neuromelanin-containing cells with or without Lewy bodies from 6 Parkinson’s Disease cases. Anti-alpha-synuclein immunohistochemistry was used to identify Lewy bodies in flash frozen tissue. Cells were collected in triplicate groups of 15 cells (LB (+) or LB (-)) from each case, and DNA was extracted with proteinase K-containing buffer. mtDNA copy number was determined by quantitative PCR for 4 genes distributed around the mitochondrial genome (12S rRNA; ND2; CO3; ND4) and normalized to a copy number standard curve of human mtDNA run on the same plate. Results: We found an average ~4-fold higher mtDNA copy numbers in LB (+) neurons compared to LB (-) neurons from the same cases, but we observed substantial heterogeneity. ND4 appeared to be deleted in all cases. ND2 appeared to be deleted in one case as was CO3 in two cases. Conclusion: Our studies indicate that in most cases of synucleinopathy, LB presence does not reduce, and appears to increase, mtDNA copy numbers in PD nigral neuromelanin-containing neurons.
