Faculty Bibliography

Membrane-bound endosomal vesicles play an integral role in multiple cellular events, including protein processing and turnover, and often critically regulate the cell-surface availability of receptors and other plasma membrane proteins in many different cell types. Neurons are no exception, being dependent on endosomal function for housekeeping and synaptic events. Growing evidence suggests a link between neuronal endosomal function and Alzheimer's disease (AD) pathophysiology. Endosomal abnormalities invariably occur within neurons in AD brains, and endocytic compartments are one likely site for the production of the pathogenic beta-amyloid peptide (Abeta), which accumulates within the brain during the disease and is generated by proteolytic processing of the amyloid precursor protein (APP). The enzymes and events involved in APP processing are appealing targets for therapeutic agents aimed at slowing or reversing the pathogenesis of AD. The neuronal endosome may well prove to be the intracellular site of action for inhibitors of beta-amyloidogenic APP processing. We present here the view that knowledge of the endosomal system in the disease can guide drug discovery of AD therapeutic agents

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 beta-amyloid (Abeta) deposits extracellularly in the presenilin (PS) 1/Abeta precursor protein (APP) mouse model of beta-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 Abeta. Purified AVs contain APP and beta-cleaved APP and are highly enriched in PS1, nicastrin, and PS-dependent gamma-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 Abeta production. Our results, therefore, link beta-amyloidogenic and cell survival pathways through macroautophagy, which is activated and is abnormal in AD

Gamma-secretase depends on presence of presenilins (PS), Nct, Aph-1, and PEN-2 within a core complex. This endoproteolytic activity cleaves within transmembrane domains of amyloid-beta precursor protein (APP) and Notch, and familial Alzheimer's disease (FAD) mutations in PS1 or PS2 genes shift APP cleavage from production of amyloid-beta (Abeta) 40 peptide to greater production of Abeta42. Although studies in PS1/PS2-deficient embryonic cells define overlapping activities for these proteins, in vivo complementation of PS1-deficient animals described here reveals an unexpected spectrum of activities dictated by PS1 and PS2 alleles. Unlike PS1 transgenes, wild-type PS2 transgenes expressed in the mouse CNS support little Abeta40 or Abeta42 production, and FAD PS2 alleles support robust production of only Abeta42. Although wild-type PS2 transgenes failed to rescue Notch-associated skeletal defects in PS1 hypomorphs, a 'gained' competence in this regard was apparent for FAD alleles of PS2. The range of discrete and divergent processing activities in mice reconstituted with different PS genes and alleles argues against gamma-secretase being a single enzyme with intrinsically relaxed substrate and cleavage site specificities. Instead, our studies define functionally distinct gamma-secretase variants. We speculate that extrinsic components, in combination with core complexes, may tailor functional variants of this enzyme to their preferred substrates

Presenilin 1 or presenilin 2, nicastrin, APH-1, and PEN-2 form high molecular weight complexes that play a pivotal role in the cleavage of various Type I transmembrane proteins, including the beta-amyloid precursor protein. The specific function of PEN-2 is unclear. To explore its function and intermolecular interactions, we conducted deletion and mutagenesis studies on a series of conserved residues at the C terminus of PEN-2. These studies suggest that: 1) both the presence and amino acid sequence of the conserved DYLSF domain at the C terminus of PEN-2 (residues 90-94) is critical for binding PEN-2 to other components in the presenilin complex and 2) the overall length of the exposed C terminus is critical for functional gamma-secretase activity

The E693Q mutation in the amyloid beta precursor protein (APP) leads to cerebral amyloid angiopathy (CAA), with recurrent cerebral hemorrhagic strokes and dementia. In contrast to Alzheimer disease (AD), the brains of those affected by hereditary cerebral hemorrhage with amyloidosis-Dutch type (HCHWA-D) show few parenchymal amyloid plaques. We found that neuronal overexpression of human E693Q APP in mice (APPDutch mice) caused extensive CAA, smooth muscle cell degeneration, hemorrhages and neuroinflammation. In contrast, overexpression of human wild-type APP (APPwt mice) resulted in predominantly parenchymal amyloidosis, similar to that seen in AD. In APPDutch mice and HCHWA-D human brain, the ratio of the amyloid-beta40 peptide (Abeta40) to Abeta42 was significantly higher than that seen in APPwt mice or AD human brain. Genetically shifting the ratio of AbetaDutch40/AbetaDutch42 toward AbetaDutch42 by crossing APPDutch mice with transgenic mice producing mutated presenilin-1 redistributed the amyloid pathology from the vasculature to the parenchyma. The understanding that different Abeta species can drive amyloid pathology in different cerebral compartments has implications for current anti-amyloid therapeutic strategies. This HCHWA-D mouse model is the first to develop robust CAA in the absence of parenchymal amyloid, highlighting the key role of neuronally produced Abeta to vascular amyloid pathology and emphasizing the differing roles of Abeta40 and Abeta42 in vascular and parenchymal amyloid pathology

The neuronal lysosomal system is a major degradative pathway, induced by cell stress and closely linked to Alzheimer disease (AD) and other neurodegenerative diseases. Here, we show that mutations of presenilin (PS) 1 and 2, which cause familial early-onset AD (FAD), induce more severe lysosomal system neuropathology in humans than does sporadic AD (SAD). Cathepsin D and B levels were higher in PS-FAD neocortex than in SAD and, unlike neurons in SAD, expressed higher levels of the cation-independent mannose-6-phosphate receptor. Lysosomal pathology was also evident in more populations of neurons in PS-FAD brains, including the less vulnerable neurons in laminae II and IV and affected neurons contained high numbers of hydrolase-positive vesicular compartments with a broader range of abnormal morphology. In transgenic mice expressing mutant amyloid precursor protein (APPswe), introducing mutant PSI significantly upregulated the lysosomal system in neocortical and hippocampal neurons. This upregulation, though milder in severity, resembled that seen in human PS-FAD. Accumulation of hydrolases in dystrophic neurites in senile plaques was particularly strong, suggesting that amyloid deposition may be a stimulus for local mobilization of the lysosomal system. PS1 mice lacking the APPswe transgene also had a mild lysosomal response in some neuronal populations, which was not seen in the APPswe mice. Our findings suggest that presenilin mutations have amyloid-independent effects on the lysosomal system, which are synergistic with the lysosomal system pathology that is associated with beta-amyloid

BACKGROUND: The over-expression of transforming growth factor beta-1(TGF-beta1) has been reported to cause hydrocephalus, glia activation, and vascular amyloidbeta (Abeta) deposition in mouse brains. Since these phenomena partially mimic the cerebral amyloid angiopathy (CAA) concomitant to Alzheimer's disease, the findings in TGF-beta1 over-expressing mice prompted the hypothesis that CAA could be caused or enhanced by the abnormal production of TGF-beta1. This idea was in accordance with the view that chronic inflammation contributes to Alzheimer's disease, and drew attention to the therapeutic potential of anti-inflammatory drugs for the treatment of Abeta-elicited CAA. We thus studied the effect of anti-inflammatory drug administration in TGF-beta1-induced pathology. METHODS: Two-month-old TGF-beta1 mice and littermate controls were orally administered pioglitazone, a peroxisome proliferator-activated receptor-gamma agonist, or ibuprofen, a non steroidal anti-inflammatory agent, for two months. Glia activation was assessed by immunohistochemistry and western blot analysis; Abeta precursor protein (APP) by western blot analysis; Abeta deposition by immunohistochemistry, thioflavin-S staining and ELISA; and hydrocephalus by measurements of ventricle size on autoradiographies of brain sections. Results are expressed as means +/- SD. Data comparisons were carried with the Student's T test when two groups were compared, or ANOVA analysis when more than three groups were analyzed. RESULTS: Animals displayed glia activation, hydrocephalus and a robust thioflavin-S-positive vascular deposition. Unexpectedly, these deposits contained no Abeta or serum amyloid P component, a common constituent of amyloid deposits. The thioflavin-S-positive material thus remains to be identified. Pioglitazone decreased glia activation and basal levels of Abeta42- with no change in APP contents - while it increased hydrocephalus, and had no effect on the thioflavin-S deposits. Ibuprofen mimicked the reduction of glia activation caused by pioglitazone and the lack of effect on the thioflavin-S-labeled deposits. CONCLUSIONS: i) TGF-beta1 over-expressing mice may not be an appropriate model of Abeta-elicited CAA; and ii) pioglitazone has paradoxical effects on TGF-beta1-induced pathology suggesting that anti-inflammatory therapy may reduce the damage resulting from active glia, but not from vascular alterations or hydrocephalus. Identification of the thioflavin-S-positive material will facilitate the full appraisal of the clinical implication of the effects of anti-inflammatory drugs, and provide a more thorough understanding of TGF-beta1 actions in brain