Dopaminergic Cell Loss Induced by Human A30P α-Synuclein Gene Transfer to the Rat Substantia Nigra

Abstract

Somatic cell gene transfer was used to express a mutant form of alpha-synuclein (alpha-syn) that is associated with Parkinson's disease (PD) in the rat substantia nigra (SN), a brain region that, in humans, degenerates during PD. DNA encoding the A30P mutant of human alpha-syn linked to familial PD was incorporated into an adeno-associated virus vector, which was injected into the adult rat midbrain. The cytomegalovirus/chicken beta-actin promoter was used to drive transgene expression. Over a 1-year time course, this treatment produced three significant features relevant to PD: (1) accumulation of alpha-syn in SN neuron perikarya, (2) Lewy-like dystrophic neurites in the SN and the striatum, and (3) a 53% loss of SN dopamine neurons. However, motor dysfunction was not found in either rotational or rotating rod testing. The lack of behavioral deficits, despite the significant cell loss, may reflect pathogenesis similar to that of PD, where greater than 50% losses occur before motor behavior is affected.

AAV2-mediated CLN2 gene transfer to rodent and non-human primate brain results in long-term TPP-I expression compatible with therapy for LINCL

“…In this respect, the choice of vector is critical. In agreement with other reports, [31][32][33][34] we have shown that an AAV2-based vector can maintain expression of the transgene and hence of the therapeutic protein for 41 year. In addition to the choice of vector, the choice of promoter is also critical for long-term high …”

Tau gene transfer, but not alpha-synuclein, induces both progressive dopamine neuron degeneration and rotational behavior in the rat

“…GFP fluorescence was detected in cell bodies in the SN and in neurites in the corpus striatum of animals injected with the GFP vector. Co-localization studies showed that the GFP fluorescence overlapped with TH immunofluorescent labeling in the majority of dopaminergic neurons in the SN pars compacta as we observed previously (Klein et al, 2002a). Co-localization studies with an antibody against parvalbumin, a marker for GABAergic neurons, demonstrated efficient expression of GFP in parvalbumin neurons of the SN pars reticulata as well (not shown).…”

“…The ASN gene transfer also produced loss of TH immunoreactivity in the pars compacta (Fig. 3F) as we observed previously (Klein et al, 2002a).We evaluated the number of SN pars compacta dopaminergic neurons after tau gene transfer by unbiased stereology on sections immunostained for TH (Table 2). GFP gene transfer did not alter the number of dopaminergic neurons compared to uninjected tissues; however, the P301L vector did cause a significant loss of dopaminergic neurons relative to control GFP gene transfer (P < 0.001, one-way ANOVA/Bonferroni's multiple comparison test) at 4 months post-injection.…”

“…The number of SN pars compacta neurons expressing TH immunoreactivity was estimated by unbiased stereology using the MicroBrightfield Inc. (Williston, VT) system as previously described (Klein et al, 2002a). Seven sections evenly spaced throughout the SN pars compacta structure were analyzed for each probe.…”

“…Human embryonic kidney 293 cells were treated for 4 days with ASN AAV vectors either with or without WPRE. By comparing bands on Western blots with the Scion imaging program (Scion Corporation, Frederick, MD), levels of ASN were boosted 10-fold by the WPRE in titer-matched comparisons (data not shown).The method for packaging plasmids in recombinant AAV-2 was previously described (Klein et al, 2002a). Human embryonic kidney 293 cells were transfected with an AAV terminal repeat-containing plasmid in an equimolar ratio with the AAV helper plasmid pDG (Grimm et al, 1998 …”