γ-Secretase Dependent Production of Intracellular Domains Is Reduced in Adult Compared to Embryonic Rat Brain Membranes

BACKGROUND: gamma-Secretase is an intramembrane aspartyl protease whose cleavage of the amyloid precursor protein (APP) generates the amyloid beta-peptide (Abeta) and the APP intracellular domain. Abeta is widely believed to have a causative role in Alzheimer's disease pathogenesis, and therefore modulation of gamma-secretase activity has become a therapeutic goal. Besides APP, more than 50 substrates of gamma-secretase with different cellular functions during embryogenesis as well as adulthood have been revealed. Prior to gamma-secretase cleavage, substrates are ectodomain shedded, producing membrane bound C-terminal fragments (CTFs). PRINCIPAL FINDINGS: Here, we investigated gamma-secretase cleavage of five substrates; APP, Notch1, N-cadherin, ephrinB and p75 neurotrophin receptor (p75-NTR) in membranes isolated from embryonic, young or old adult rat brain by analyzing the release of the corresponding intracellular domains (ICDs) or Abeta40 by western blot analysis and ELISA respectively. The highest levels of all ICDs and Abeta were produced by embryonic membranes. In adult rat brain only cleavage of APP and Notch1 could be detected and the Abeta40 and ICD production from these substrates was similar in young and old adult rat brain. The CTF levels of Notch1, N-cadherin, ephrinB and p75-NTR were also clearly decreased in the adult brain compared to embryonic brain, whereas the APP CTF levels were only slightly decreased. CONCLUSIONS: In summary our data suggests that gamma-secretase dependent ICD production is down-regulated in the adult brain compared to embryonic brain. In addition, the present approach may be useful for evaluating the specificity of gamma-secretase inhibitors

Macroautophagy—a novel β-amyloid peptide-generating pathway activated in Alzheimer's disease

Macroautophagy, which is a lysosomal pathway for the turnover of organelles and long-lived proteins, is a key determinant of cell survival and longevity. In this study, we show that neuronal macroautophagy is induced early in Alzheimer's disease (AD) and before β-amyloid (Aβ) deposits extracellularly in the presenilin (PS) 1/Aβ precursor protein (APP) mouse model of β-amyloidosis. Subsequently, autophagosomes and late autophagic vacuoles (AVs) accumulate markedly in dystrophic dendrites, implying an impaired maturation of AVs to lysosomes. Immunolabeling identifies AVs in the brain as a major reservoir of intracellular Aβ. Purified AVs contain APP and β-cleaved APP and are highly enriched in PS1, nicastrin, and PS-dependent γ-secretase activity. Inducing or inhibiting macroautophagy in neuronal and nonneuronal cells by modulating mammalian target of rapamycin kinase elicits parallel changes in AV proliferation and Aβ production. Our results, therefore, link β-amyloidogenic and cell survival pathways through macroautophagy, which is activated and is abnormal in AD

Gastric carcinoma in young Hong Kong Chinese

Gastric carcinomas usually occur in older people. Those occurring in the young are uncommon. The pathological and clinical features of gastric carcinomas were reviewed in 42 Chinese patients who were 35 years of age and younger. The data were obtained from the record files of the University Department of Pathology, Queen Mary Hospital for the period 1976-85. The patients comprised 4% of the total cases of gastric carcinomas in that period. These patients (age range: 20-35 years, mean: 30 years) showed a male to female ratio of 1:2.5 which differs from the usual male preponderance seen in gastric carcinoma. Among the 27 cases with known staging, 22 (81.5%) were stage III or IV. Twenty-five cases had an ulcerative appearance. All were adenocarcinomas and the majority (83.3%) were poorly differentiated. Associated dysplasia was found in 35 (83.3%) cases, although only 14 of these were in association with poorly differentiated adenocarcinomas. Intestinal metaplasia was found in 13 cases and, when present, involved less than 30% of the mucosa. Only two cases were of type III metaplasia. The findings show that gastric carcinoma in young Chinese tended to occur more frequently in females, presented at late stages, showed poor glandular differentiation, was frequently associated with gastric dysplasia and had minimal association with intestinal metaplasia.link_to_subscribed_fulltex

Live Cell FRET Imaging Reveals Amyloid β-Peptide Oligomerization in Hippocampal Neurons

Amyloid β-peptide (Aβ) oligomerization is believed to contribute to the neuronal dysfunction in Alzheimer disease (AD). Despite decades of research, many details of Aβ oligomerization in neurons still need to be revealed. Förster resonance energy transfer (FRET) is a simple but effective way to study molecular interactions. Here, we used a confocal microscope with a sensitive Airyscan detector for FRET detection. By live cell FRET imaging, we detected Aβ42 oligomerization in primary neurons. The neurons were incubated with fluorescently labeled Aβ42 in the cell culture medium for 24 h. Aβ42 were internalized and oligomerized in the lysosomes/late endosomes in a concentration-dependent manner. Both the cellular uptake and intracellular oligomerization of Aβ42 were significantly higher than for Aβ40. These findings provide a better understanding of Aβ42 oligomerization in neurons

Assembling amyloid fibrils from designed structures containing a significant amyloid beta-peptide fragment.

The amyloid plaque, consisting of amyloid beta-peptide (Abeta) fibrils surrounded by dystrophic neurites, is an invariable feature of Alzheimer's disease. The determination of the molecular structure of Abeta fibrils is a significant goal that may lead to the structure-based design of effective therapeutics for Alzheimer's disease. Technical challenges have thus far rendered this goal impossible. In the present study, we develop an alternative methodology. Rather than determining the structure directly, we design conformationally constrained peptides and demonstrate that only certain 'bricks' can aggregate into fibrils morphologically identical to Abeta fibrils. The designed peptides include variants of a decapeptide fragment of Abeta, previously shown to be one of the smallest peptides that (1) includes a pentapeptide sequence necessary for Abeta-Abeta binding and aggregation and (2) can form fibrils indistinguishable from those formed by full-length Abeta. The secondary structure of these bricks is monitored by CD spectroscopy, and electron microscopy is used to study the morphology of the aggregates formed. We then made various residue deletions and substitutions to determine which structural features are essential for fibril formation. From the constraints, statistical analysis of side-chain pair correlations in beta-sheets and experimental data, we deduce a detailed model of the peptide strand alignment in fibrils formed by these bricks. Our results show that the constrained decapeptide dimers rapidly form an intramolecular, antiparallel beta-sheet and polymerize into amyloid fibrils at low concentrations. We suggest that the formation of an exposed beta-sheet (e.g. an Abeta dimer formed by interaction in the decapeptide region) could be a rate-limiting step in fibril formation. A theoretical framework that explains the results is presented in parallel with the data