Differential effects of proteases involved in intracellular degradation of amyloid β-protein between detergent-soluble and -insoluble pools in CHO-695 cells

The deposition of amyloid beta-protein (Abeta or betaA4) is a key feature of Alzheimer's disease. Most studies have focused on the generation of Abeta, but little is known about the degradation of Abeta. Recent reports suggest that insulin-degrading enzyme (IDE) and neutral endopeptidase (NEP) are involved in the extracellular degradation of A. To date, however, far less is known about the degradation of intracellular Abeta. To elucidate the protease(s) responsible for the degradation of intracellular Abeta, we investigated the effect of various protease inhibitors on Abeta 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 Abeta 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 Abeta as well as extracellular Abeta40 and Abeta42. Moreover, intracellular Abeta 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 Abeta in the detergent-insoluble fraction markedly and partially degraded extracellular Abeta40 and Abeta42, but not intracellular soluble Abeta. Thiorphan, an inhibitor of NEP, accumulated, albeit to a lesser extent, in insoluble Abeta but not in soluble Abeta. Thus, IDE appears to degrade intracellular Abeta more effectively than does NEP in both the detergent-soluble and -insoluble fractions.