Cathepsin B but not cathepsins L or S contributes to the pathogenesis of Unverricht-Lundborg progressive myoclonus epilepsy (EPM1).

The inherited epilepsy Unverricht-Lundborg disease (EPM1) is caused by loss-of-function mutations in the cysteine protease inhibitor, cystatin B. Because cystatin B inhibits a class of lysosomal cysteine proteases called cathepsins, we hypothesized that increased proteolysis by one or more of these cathepsins is likely to be responsible for the seizure, ataxia, and neuronal apoptosis phenotypes characteristic of EPM1. To test this hypothesis and to identify which cysteine cathepsins contribute to EPM1, we have genetically removed three candidate cathepsins from cystatin B-deficient mice and tested for rescue of their EPM1 phenotypes. Whereas removal of cathepsins L or S from cystatin B-deficient mice did not ameliorate any aspect of the EPM1 phenotype, removal of cathepsin B resulted in a 36-89% reduction in the amount of cerebellar granule cell apoptosis depending on mouse age. The incidence of an incompletely penetrant eye phenotype was also reduced upon removal of cathepsin B. Because the apoptosis and eye phenotypes were not abolished completely and the ataxia and seizure phenotypes experienced by cystatin B-deficient animals were not diminished, this suggests that another molecule besides cathepsin B is also responsible for the pathogenesis, or that another molecule can partially compensate for cathepsin B function. These findings establish cathepsin B as a contributor to the apoptotic phenotype of cystatin B-deficient mice and humans with EPM1. They also suggest that the identification of cathepsin B substrates may further reveal the molecular basis for EPM1.

Neuropathological changes in a mouse model of progressive myoclonus epilepsy: cystatin B deficiency and Unverricht-Lundborg disease.

Progressive myoclonus epilepsy of the Unverricht-Lundborg type (EPM1) is a recessively inherited neurodegenerative disease caused by loss-of-function mutations in the gene encoding cystatin B, a cysteine protease inhibitor. Mice with disruptions in this gene display myoclonic seizures, progressive ataxia, and cerebellar pathology closely paralleling EPMI in humans. To provide further insight into our understanding of EPM1, we report pathological findings in brains from cystatin B-deficient mice. In addition to confirming the loss of cerebellar granular cell neurons by apoptosis, we identified additional neuronal apoptosis in young mutant mice (3-4 months old) within the hippocampal formation and entorhinal cortex. In older mutant mice (16-18 months old), there was also gliosis most marked in the presubiculum and parasubiculum of the hippocampal formation, as well as the entorhinal cortex, neocortex, and striatum. Furthermore, widespread white matter gliosis was also noted, which may be a secondary phenomenon. Within the cerebral cortex, cellular atrophy was a prominent finding in the superficial neurons of the prosubiculum. Finally, we show that mutant mice in either a "seizure-prone" or "seizure-resistant" genetic background display similar neuropathological changes. These findings indicate that neuronal atrophy is an important consequence of cystatin-B deficiency independent of seizure events, suggesting a physiological role for this protein in the maintenance of normal neuronal structure.