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Hoffmann, R. A wiki for the life sciences where authorship matters. Nature Genetics (2008)

A tetracycline-regulated adenovirus encoding dominant-negative caspase-9 is regulated in rat brain and protects against neurotoxin-induced cell death in vitro, but not in vivo.

Caspase-9 is a critical downstream effector molecule involved in apoptosis, a cell death process thought to be involved in the demise of dopamine (DA) neurons in the substantia nigra (SN) affected by Parkinson's disease (PD). In this study, we determined that a tetracycline-regulated adenovirus harboring a dominant-negative form of caspase-9 (Casp9DN) and the marker gene, enhanced green fluorescent protein (EGFP), under the control of a bidirectional promoter could each be regulated in vitro and in vivo by doxycycline. We next observed that Casp9DN gene delivery significantly protected against TNFalpha and cycloheximide-induced chromatin condensation in HeLa cells and prevented chromatin condensation and the appearance of the early apoptotic marker annexin V in 6-hydroxydopamine (6-OHDA) treated MN9D cells, a dopaminergic cell line. Effects of Casp9DN on DA neurons in vivo were also assessed. DA neurons were retrogradely labeled with fluorogold (FG) and transduced with Casp9DN and EGFP or EGFP alone. A progressive lesion of DA neurons was induced by striatal injection of 6-OHDA 1 week later. At 2 weeks post-lesion, a morphometric analysis of FG+ neurons in the SN revealed that the mean cell diameter of FG labeled neurons in the Casp9DN group was 8% and 21% larger than the EGFP and PBS groups, respectively (P <0.05). However, there was no difference among the treatment groups in the number of neurons remaining in the lesioned SN. These results suggest that while inhibiting apoptosis at the level of caspase-9 is protective in vitro, it is not protective against 6-OHDA-induced cell death in vivo.[1]


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