Faculty Bibliography

beta-Amyloid precursor protein (APP) mutations cause familial Alzheimer's disease with nearly complete penetrance. We found an APP mutation [alanine-673-->valine-673 (A673V)] that causes disease only in the homozygous state, whereas heterozygous carriers were unaffected, consistent with a recessive Mendelian trait of inheritance. The A673V mutation affected APP processing, resulting in enhanced beta-amyloid (Abeta) production and formation of amyloid fibrils in vitro. Co-incubation of mutated and wild-type peptides conferred instability on Abeta aggregates and inhibited amyloidogenesis and neurotoxicity. The highly amyloidogenic effect of the A673V mutation in the homozygous state and its anti-amyloidogenic effect in the heterozygous state account for the autosomal recessive pattern of inheritance and have implications for genetic screening and the potential treatment of Alzheimer's disease

Down's syndrome (DS) in humans is caused by trisomy of chromosome 21 (HSA 21). DS patients have a variety of pathologies, including mental retardation and an unusually high incidence of leukemia or lymphoma such as megakaryocytic leukemia. Individuals with DS develop the characteristic neuropathological hallmarks of Alzheimer's disease (AD) in early adulthood, generally by the fourth decade of life. There are several mouse models of DS that have a segmental trisomy of mouse chromosome 16 (MMU 16) with triplicated genes orthologous to HSA 21. These mice display neurodegeneration similar to DS. Although brain pathology in DS models is known, little information is available about other organs. We studied the extraneural pathology in aged DS mice (Ts65Dn, Ts2 and Ts1Cje aged 8 to 24 months) as well as other mouse models of neurodegeneration, including presenilin (PS), amyloid-beta precursor protein (APP), and tau (hTau and JNPL) transgenic mice. An increased incidence of peripheral amyloidosis, positive for amyloid A (AA) but not amyloid-beta peptide (A beta), was found in APP over-expressing and tauopathic mice as compared to non-transgenic (ntg) littermates or to DS mouse models. A higher incidence of lymphoma was found in the DS models, including Ts1Cje that is trisomic for a small segment of MMU 16 not including the App gene, but not in the APP over-expressing mice, suggesting that high APP expression is not the cause of lymphoma in DS. The occurrence of lymphomas in mouse DS models is of interest in relation to the increased incidence of malignant conditions in human DS

A role for cystatin C (CysC) in the pathogenesis of Alzheimer's disease (AD) has been suggested by the genetic linkage of a CysC gene (CST3) polymorphism with late-onset AD, the co-localization of CysC with amyloid-beta (Abeta) in AD brains, and binding of CysC to soluble Abeta in vitro and in mouse models of AD. This study investigates the binding between Abeta and CysC in the human central nervous system. While CysC binding to soluble Abeta was observed in AD patients and controls, a SDS-resistant CysC/Abeta complex was detected exclusively in brains of neuropathologically normal controls, but not in AD cases. The association of CysC with Abeta in brain from control individuals and in cerebrospinal fluid reveals an interaction of these two polypeptides in their soluble form. The association between Abeta and CysC prevented Abeta accumulation and fibrillogenesis in experimental systems, arguing that CysC plays a protective role in the pathogenesis of AD in humans and explains why decreases in CysC concentration caused by the CST3 polymorphism or by specific presenilin 2 mutations can lead to the development of the disease. Thus, enhancing CysC expression or modulating CysC binding to Abeta have important disease-modifying effects, suggesting a novel therapeutic intervention for AD

Mechanisms of neuronal loss in Alzheimer's disease (AD) are poorly understood. Here we show that apoptosis is a major form of neuronal cell death in PS/APP mice modeling AD-like neurodegeneration. Pyknotic neurons in adult PS/APP mice exhibited apoptotic changes, including DNA fragmentation, caspase-3 activation, and caspase-cleaved alpha-spectrin generation, identical to developmental neuronal apoptosis in wild-type mice. Ultrastructural examination using immunogold cytochemistry confirmed that activated caspase-3-positive neurons also exhibited chromatin margination and condensation, chromatin balls, and nuclear membrane fragmentation. Numbers of apoptotic profiles in both cortex and hippocampus of PS/APP mice compared with age-matched controls were twofold to threefold higher at 6 months of age and eightfold higher at 21 to 26 months of age. Additional neurons undergoing dark cell degeneration exhibited none of these apoptotic features. Activated caspase-3 and caspase-3-cleaved spectrin were abundant in autophagic vacuoles, accumulating in dystrophic neurites of PS/APP mice similar to AD brains. Administration of the cysteine protease inhibitor, leupeptin, promoted accumulation of autophagic vacuoles containing activated caspase-3 in axons of PS/APP mice and, to a lesser extent, in those of wild-type mice, implying that this pro-apoptotic factor is degraded by autophagy. Leupeptin-induced autophagic impairment increased the number of apoptotic neurons in PS/APP mice. Our findings establish apoptosis as a mode of neuronal cell death in aging PS/APP mice and identify the cross talk between autophagy and apoptosis, which influences neuronal survival in AD-related neurodegeneration

Using transgenic mice expressing human cystatin C (encoded by CST3), we show that cystatin C binds soluble amyloid-beta peptide and inhibits cerebral amyloid deposition in amyloid-beta precursor protein (APP) transgenic mice. Cystatin C expression twice that of the endogenous mouse cystatin C was sufficient to substantially diminish amyloid-beta deposition. Thus, cystatin C has a protective role in Alzheimer's disease pathogenesis, and modulation of cystatin C concentrations may have therapeutic implications for the disease

The CST3 Thr25 allele of CST3, which encodes cystatin C, leads to reduced cystatin C secretion and conveys susceptibility to Alzheimer's disease. Here we show that overexpression of human cystatin C in brains of APP-transgenic mice reduces cerebral amyloid-beta deposition and that cystatin C binds amyloid-beta and inhibits its fibril formation. Our results suggest that cystatin C concentrations modulate cerebral amyloidosis risk and provide an opportunity for genetic risk assessment and therapeutic interventions

Studies in continuously cultured cells have established that familial Alzheimer's disease (FAD) mutant presenilin 1 (PS1) delays exit of the amyloid precursor protein (APP) from the trans-Golgi network (TGN). Here we report the first description of PS1-regulated APP trafficking in cerebral neurons in culture and in vivo. Using neurons from transgenic mice or a cell-free APP transport vesicle biogenesis system derived from the TGN of those neurons, we demonstrated that knocking-in an FAD-associated mutant PS1 transgene was associated with delayed kinetics of APP arrival at the cell surface. Apparently, this delay was at least partially attributable to impaired exit of APP from the TGN, which was documented in the cell-free APP transport vesicle biogenesis assay. To extend the study to APP and carboxyl terminal fragment (CTF) trafficking to cerebral neurons in vivo, we performed subcellular fractionation of brains from APP transgenic mice, some of which carried a second transgene encoding an FAD-associated mutant form of PS1. The presence of the FAD mutant PS1 was associated with a slight shift in the subcellular localization of both holoAPP and APP CTFs toward iodixanol density gradient fractions that were enriched in a marker for the TGN. In a parallel set of experiments, we used an APP : furin chimeric protein strategy to test the effect of artificially forcing TGN concentration of an APP : furin chimera that could be a substrate for beta- and gamma-cleavage. This chimeric substrate generated excess Abeta42 when compared with wildtype APP. These data indicate that the presence of an FAD-associated mutant human PS1 transgene is associated with redistribution of the APP and APP CTFs in brain neurons toward TGN-enriched fractions. The chimera experiment suggests that TGN-enrichment of a beta-/gamma-secretase substrate may play an integral role in the action of mutant PS1 to elevate brain levels of Abeta42