Accumulation and Aggregation of Amyloid β-Protein in Late Endosomes of Niemann-Pick Type C Cells

Abstract

There is growing evidence suggesting that cholesterol metabolism is linked to susceptibility to Alzheimer's disease by influencing amyloid ␤-protein (A␤) metabolism. However, the precise cellular linkage sites between cholesterol and A␤ have not yet been clarified. To address this issue, we investigated Niemann-Pick type C (NPC) model cells and NPC mutant cells, which showed aberrant cholesterol trafficking. We observed a remarkable A␤ accumulation in late endosomes of both NPC model cells and mutant cells where cholesterol accumulates and a significant accumulation in the NPC mouse brain. This A␤ accumulation was independent of its constitutive secretion and production through an endocytic pathway. In addition, it is characterized by a marked predominance of A␤42 and insolubility in SDS, suggesting the presence of aggregated A␤ in late endosomes. Most importantly, A␤ accumulation is coupled with the cholesterol levels in late endosomes. Thus, late endosomes of NPC cells are a novel pool of aggregated A␤42 as well as cholesterol, suggesting a direct interaction between aggregated A␤ and cholesterol.Formation of senile plaques composed of amyloid ␤-protein (A␤) 1 is one of the hallmarks of Alzheimer's disease (AD) (1). A␤, a small protein of M r ϳ4,000, is cleaved out sequentially with ␤-and ␥-secretases from a large membrane protein called ␤-amyloid precursor protein (APP), and secreted into the extracellular space. There are two major species of A␤ that are defined by their C-terminal lengths; A␤40 terminates at Val-40, and A␤42 terminates at Ala-42. The latter has a much stronger tendency to aggregate into fibrils and accounts for less than 10% of secreted A␤. However, A␤42 is the most predominant form found in senile plaques and is considered the initial species to be deposited in aged and AD brains (2, 3).Three causative genes, APP, presenilin 1 (PS1), and PS2 have been identified in early onset familial AD (1), and their mutations appear to converge on an increased production of A␤42. However, the pathological mechanisms of sporadic AD, explaining more than 90% of AD patients, remain unknown. Interestingly, there is growing evidence for a significant linkage between A␤ and cholesterol metabolism. First, cholesterol loading or depletion affects A␤ generation both in vitro (4,5) and in vivo (6). Second, the levels of total cholesterol and LDL-cholesterol, but not those of HDL-cholesterol, in the serum correlate with the amount of A␤42 in AD brains (7). Third, aggregated A␤ preferentially binds to cholesterol in vitro (8). Fourth, a substantial fraction of intracellular A␤ is localized in the detergent-insoluble membrane domain that is rich in glycosphingolipid and cholesterol (9, 10). Fifth, the presence of an E4 isoform of apolipoprotein E, an essential molecule for cholesterol metabolism, is a strong risk factor for developing AD (11). Sixth, high dietary cholesterol accelerates AD-related pathologies, including A␤ deposition, in a transgenic mouse model (12). However, no detailed mechanisms for clarifying ...

Endothelin-converting enzyme-1 is expressed in human cerebral cortex and protects against Alzheimer's disease

“…39 BACE-1 is also targeted to the endosomal system, 40 and the results of several groups support an important role for this intracellular compartment in Ab production. [41][42][43] In addition, endocytic pathway abnormalities have been shown to precede Ab deposition in the brains of patients affected by sporadic AD. 44 Finally, our findings also offer new therapeutic options in AD.…”

Subcellular Compartment and Molecular Subdomain of β-Amyloid Precursor Protein Relevant to the Aβ42-promoting Effects of Alzheimer Mutant Presenilin 2

“…For the detection of A␤ in RIPA-soluble fractions of cell lysates, samples were initially lysed in RIPA (50 mM Tris-HCl, pH 7.5, 150 mM NaCl, 1% Triton X-100, 1% sodium deoxycholate), and the supernatants after centrifugation at 15,000 ϫ g for 5 min were immunoprecipitated by BAN50 using protein G-agarose and then analyzed by immunoblotting with BA27, BC05, or BAN50, using previously described procedures (44,45,47). Extraction of cell-associated A␤ by formic acid was performed as described (46). Briefly, cell pellets confluently grown in two 10-cm dishes were solubilized by ultrasonication followed by incubation in 100 l of 70% formic acid at room temperature for 30 min.…”

Preliminary Results on Long-Term Potentiation-Like Cortical Plasticity and Cholinergic Dysfunction After Miglustat Treatment in Niemann-Pick Disease Type C

“…NPC is also characterized by the accumulation of beta amyloid (Yamazaki et al 2001) and neurofibrillary tangles (Love et al 1995), which are immunologically and ultrastructurally similar to those seen in Alzheimer's disease (AD) (Auer et al 1995). The similarities between NPC and AD extend to the cholesterol pathway, which has been repeatedly implicated in the pathogenesis of AD (Liu et al 2013).…”