Activation of the Notch Signaling Pathway and Cellular Localization of Notch Signaling Molecules in the Spinal Cord of SOD1-G93A ALS Model Mice.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by motor neuron loss and gliosis in the spinal cord, brain stem and cortex. The Notch signaling pathway has been reported to be dysfunctional in neurodegenerative diseases, including ALS. However, the exact mechanism is still unclear. Here, we detected Notch signaling activation in proliferating glial cells, Notch inactivation in motor neurons in the spinal cord of the SOD1-G93A model, and dramatic changes of cellular relocalization of Notch pathway signaling molecules, including activated Notch intracellular domain (NICD), Notch ligands (Jagged1 and DLL4) and the target gene Hes1. We found that Notch activation was universal in proliferating astrocytes and that the Notch ligand Jagged1 was uniquely upregulated in proliferating microglia, while DLL4 expression was increased in both activated astrocytes and degenerating oligodendrocytes. Our results indicate that microglia may play an important role in the intercellular receptor-ligand interaction of the Notch signaling pathway and contribute to the pathogenesis of motor neuron loss in ALS mice. Further experiments are required to clarify the exact mechanism responsible for Notch dysfunction in ALS.

Slow Intrathecal Injection of rAAVrh10 Enhances its Transduction of Spinal Cord and Therapeutic Efficacy in a Mutant SOD1 Model of ALS.

Mutant SOD1 causes amyotrophic lateral sclerosis (ALS) by a dominant gain of toxicity. Previous studies have demonstrated therapeutic potential of mutant SOD1-RNAi delivered by intrathecal (IT) injection of recombinant adeno-associated virus (rAAV). However, optimization of delivery is needed to overcome the high degree of variation in the transduction efficiency and therapeutic efficacy. Here, on the basis of our previously defined, efficient IT injection method, we investigated the influence of injection speed on transduction efficiency in the central nervous system (CNS). We demonstrate that slow IT injection results in higher transduction of spinal cord and dorsal root ganglia (DRG) while fast IT injection leads to higher transduction of brain and peripheral organs. To test how these effects influence the outcome of RNAi therapy, we used slow and fast IT injection to deliver rAAVrh10-GFP-amiR-SOD1, a rAAV vector that expresses GFP and an artificial miRNA targeting SOD1, in SOD1-G93A mice. Both slow and fast IT injection produced therapeutic efficacy but the slow injection trended slightly toward a better outcome than the fast injection. These results demonstrate that IT injection speed influences the predominance of gene delivery at different CNS sites and should be taken into consideration in future therapeutic trials involving IT injection.