Differential Effects of Proteases Involved in Intracellular Degradation of Amyloid β-Protein between Detergent-Soluble and -Insoluble Pools in CHO-695 Cells

Publisher Website
Google Scholar
Abstract
The deposition of amyloid β-protein (Aβ or βA4) is a key feature of Alzheimer's disease. Most studies have focused on the generation of Aβ, but little is known about the degradation of Aβ. Recent reports suggest that insulin-degrading enzyme (IDE) and neutral endopeptidase (NEP) are involved in the extracellular degradation of Aβ. To date, however, far less is known about the degradation of intracellular Aβ. To elucidate the protease(s) responsible for the degradation of intracellular Aβ, we investigated the effect of various protease inhibitors on Aβ in two distinct intracellular pools (i.e., nonionic detergent-soluble and detergent-insoluble pools) in Chinese hamster ovary cells. Treatment with thiol and metal inhibitors resulted in the accumulation of intracellular Aβ and oligomers in detergent-soluble and -insoluble fractions. The overexpression of thiol-metalloprotease IDE resulted in a marked reduction in levels of detergent-soluble intracellular Aβ as well as extracellular Aβ40 and Aβ42. Moreover, intracellular Aβ in the detergent-insoluble fraction extracted with 70% formic acid or 6 M guanidine hydrochloride decreased markedly in the cells overexpressing IDE. In contrast, expression of NEP degraded the Aβ in the detergent-insoluble fraction markedly and partially degraded extracellular Aβ40 and Aβ42, but not intracellular soluble Aβ. Thiorphan, an inhibitor of NEP, accumulated, albeit to a lesser extent, in insoluble Aβ but not in soluble Aβ. Thus, IDE appears to degrade intracellular Aβ more effectively than does NEP in both the detergent-soluble and -insoluble fractions.