O1-06-08: Caspase-6 activation disrupts the human neuronal cytoskeleton: Strong implication for an early cause of neuronal disruption in mild cognitively impaired and Alzheimer's disease individuals

Abstract

The cystenyl Caspase-6 protease is highly active in the neurofibrillary tangles, neuropil threads and neuritic plaques of sporadic Alzheimer disease (AD) brains. The active Caspase-6 occurs in neurons that display neurodegeneration but do not have apoptotic morphology. Active Caspase-6 is present in pre-tangles and occurs pre-clinically in aged individuals that have normal but lower cognitive scores than their counterparts, suggesting that Caspase-6 could be an instigator of neurodegeneration. The objective of this study is to determine the effect of active Caspase-6 in the neurons of aged and AD individuals. Cytoskeleton proteins were assessed as substrates of active Caspase-6 in vitro. Caspase-6 was selectively activated by serum deprivation of human primary neurons in the absence or presence of Caspase-6 inhibitors. Cytoskeleton integrity was assessed by immunohistochemistry and western blotting against tubulin, MAP2, neurofilament, and Tau protein. The presence of Caspase-6-cleaved proteins in AD was assessed by immunohistochemistry with neoepitope antisera. Tubulin, MAP2, and Tau, but not Neurofilament proteins, are cleaved by Caspase-6. Whereas tubulin and MAP2 are homogenously distributed in the neurites of primary human neurons in normal culture conditions, they take a beaded appearance in serum-deprived conditions. Tau protein also appears beaded in serum deprivation and additionally redistributes around the nuclei. In contrast, the neurofilaments remain intact with serum deprivation. Caspase-6 inhibitors prevent the cytoskeleton beading and Tau redistribution. Tubulin and Tau cleaved by Caspase-6 are present in AD brains. These results show that the activation of Caspase-6 in human neurons disrupts the integrity of the cytoskeleton. Because of the importance of the cytoskeleton in proper vesicular and protein trafficking, it is likely that the activation of Caspase-6 not only disrupts the architecture but the function of neurons. Together, these results suggest that the activation of Caspase-6 disrupts cytoskeletal integrity and neuronal function in AD.