Differentiation and transcription factor gene therapy in experimental parkinson's disease: sonic hedgehog and gli-1, but not Nurr-1, protect nigrostriatal cell bodies from 6-OHDA-induced neurodegeneration

Abstract

We tested the activity of the dopaminergic neuron differentiation factor sonic hedgehog, its downstream transcription factor target Gli-1, and an orphan nuclear receptor, Nurr-1, necessary for the induction of the dopaminergic phenotype of nigrostriatal neurons, in an in vivo model of nigrostriatal neurodegeneration. Our preliminary experiments demonstrated that all three constructs expressed the proper molecules and that these had the predicted biological activities in vitro. We expressed the N-terminal of sonic hedgehog (ShhN) and the Gli-1 and Nurr-1 entire coding regions from highly purified, and quality controlled, replication-defective adenoviral vectors injected into the brains of rats and used the dopaminergic growth factor GDNF as a positive control. The neurotoxin 6-hydroxydopamine was used to lesion the nigrostriatal dopaminergic innervation; RAd-ShhN and RAd-Gli-1 protected dopaminergic neuronal cell bodies in the substantia nigra, but not axonal terminals in the striatum, from 6-OHDA-induced cell death, while RAd-Nurr-1 was ineffective in protecting either cell bodies or axons. RAd-GDNF was able to protect both the dopaminergic cell bodies and the striatal axon terminals. Our results establish for the first time, to the best of our knowledge, that gene transfer of ShhN and one of its target transcription factors can selectively protect dopaminergic nigrostriatal neuronal cell bodies from a specific neurotoxic insult. Selective protection of nigrostriatal dopaminergic cell bodies by the differentiation factor ShhN and the transcription factor Gli-1 was achieved in a neurotoxic model that eliminates more than 70% of the nigral neurons under consideration. Differentiation and transcription factors can thus be used for the treatment of neurodegeneration by gene therapy.

Neuronal expression of the transcription factor Gli1 using the Tα1 α-tubulin promoter is neuroprotective in an experimental model of Parkinson's disease

“…Also, the immunostaining for TH-immunoreactive fibers in the striatum indicated that, as had been described by us recently, 33,34 only GDNF (Figure 6b) was able to partially preserve striatal dopaminergic fibers ( Figure 6). Only in animals injected intrastriatally with RAd-GDNF did we detect a halo of intense THimmunoreactive fibers surrounding the intrastriatal injection site (Figures 6b and 7).…”

“…The vectors RAd-GDNF and RAd-hCMV-Gli1 have been described by us recently, 33,34 as has the vector expressing the control marker transgene b-galactosidase under the control of the hCMV promoter. [41][42][43][44] The neuroprotective paradigm used was adapted to our particular needs from the detailed 6-OHDA rat model of Parkinson's disease 2,3 In our experiments, the RAds were injected stereotaxically into the rat striatum using the same coordinates as used for the retrograde tracer fluorogold (FG) (Figure 4, and described in detail in Hurtado-Lorenzo et al 33,34 ).…”

“…[41][42][43][44] The neuroprotective paradigm used was adapted to our particular needs from the detailed 6-OHDA rat model of Parkinson's disease 2,3 In our experiments, the RAds were injected stereotaxically into the rat striatum using the same coordinates as used for the retrograde tracer fluorogold (FG) (Figure 4, and described in detail in Hurtado-Lorenzo et al 33,34 ). After 1 week, we induced degeneration of the nigrostriatal pathway by injecting 16 mg of 6-OHDA at the same coordinates of FG and virus injection.…”

“…After 1 week, we induced degeneration of the nigrostriatal pathway by injecting 16 mg of 6-OHDA at the same coordinates of FG and virus injection. We used a dose of 0.02 ml of FG in order to reduce the target neuronal population to be examined to a specific population of dopaminergic neurons within the SNpc, which would be exposed both to the RAds and FG, as described by us in detail recently 33,34 (Figure 4).…”

“…[30][31][32] We have recently demonstrated that adenoviral vectors encoding ShhN, or Gli1 under the control of pancellular viral promoters, have a neuroprotective effect on nigrostriatal dopaminergic neurons in an experimental model of dopamine neurodegeneration induced by 6-OHDA. 33,34 Gli1 is a transcription factor and is thus needed intracellularly in order to be able to exert its neuroprotective effects. Having previously demonstrated the specificity 35 and therapeutic effectiveness 36 of cell-type specific promoters within adenoviral vectors, we hypothesized that expression of Gli1 from a neuronal-specific promoter could increase its effectiveness and safety.…”