Ronald B. DeMattos

Immunization reverses memory deficits without reducing brain Aβ burden in Alzheimer's disease model

We have previously shown that chronic treatment with the monoclonal antibody m266, which is specific for amyloid β-peptide (Aβ), increases plasma concentrations of Aβ and reduces Aβ burden in the PDAPP transgenic mouse model of Alzheimers disease (AD). We now report that administration of m266 to PDAPP mice can rapidly reverse memory deficits in both an object recognition task and a holeboard learning and memory task, but without altering brain Aβ burden. We also found that an Aβ/antibody complex was present in both the plasma and the cerebrospinal fluid of m266-treated mice. Our data indicate that passive immunization with this anti-Aβ monoclonal antibody can very rapidly reverse memory impairment in certain learning and memory tasks in the PDAPP mouse model of AD, owing perhaps to enhanced peripheral clearance and (or) sequestration of a soluble brain Aβ species.

A gamma-secretase inhibitor decreases amyloid-beta production in the central nervous system

Accumulation of amyloid‐β (Aβ) by overproduction or underclearance in the central nervous system (CNS) is hypothesized to be a necessary event in the pathogenesis of Alzheimers disease. However, previously, there has not been a method to determine drug effects on Aβ production or clearance in the human CNS. The objective of this study was to determine the effects of a γ‐secretase inhibitor on the production of Aβ in the human CNS.

Clusterin contributes to caspase-3-independent brain injury following neonatal hypoxia-ischemia.

Clusterin, also known as apolipoprotein J, is a ubiquitously expressed molecule thought to influence a variety of processes including cell death. In the brain, it accumulates in dying neurons following seizures and hypoxic-ischemic (H-I) injury. Despite this, in vivo evidence that clusterin directly influences cell death is lacking. Following neonatal H-I brain injury in mice (a model of cerebral palsy), there was evidence of apoptotic changes (neuronal caspase-3 activation), as well as accumulation of clusterin in dying neurons. Clusterin-deficient mice had 50% less brain injury following neonatal H-I. Surprisingly, the absence of clusterin had no effect on caspase-3 activation, and clusterin accumulation and caspase-3 activation did not colocalize to the same cells. Studies with cultured cortical neurons demonstrated that exogenous purified astrocyte-secreted clusterin exacerbated oxygen/glucose-deprivation–induced necrotic death. These results indicate that clusterin may be a new therapeutic target to modulate non-caspase-dependent neuronal death following acute brain injury.

Evidence for peripheral clearance of cerebral Aβ protein following chronic, active Aβ immunization in PSAPP mice

Immunization with amyloid-β (Aβ) peptide in mouse models of Alzheimer’s disease has been reported to decrease cerebral Aβ levels and improve behavioral deficits. Several mechanisms have been proposed, including antibody-induced phagocytosis of Aβ by cerebral microglia and increased efflux of Aβ from the brain to the periphery. The latter mechanism was suggested in mice undergoing acute, passive transfer of an Aβ monoclonal antibody. Here, PSAPP transgenic mice were actively immunized by a single intraperitoneal injection of synthetic Aβ followed by chronic intranasal administration of Aβ with the mucosal adjuvant, Escherichia coli heat-labile enterotoxin, LT, twice weekly for 8 weeks. Serum from Aβ-immunized mice had an average of 240 μg/ml of anti-Aβ-specific antibodies; these antibodies had epitope(s) within Aβ1-15 and were of immunoglobulin (Ig) isotypes IgG2b, IgG2a, and IgG1. Immunization led to a 75% decrease in plaque number (P < 0.0001) and a 58% decrease in Aβx–42 levels (P < 0.026) in brain, and gliosis and neuritic dystrophy were diminished. No pathological effects of the immunization were observed in kidney, spleen, or snout. Serum Aβ levels increased 28-fold in immunized mice (53.06 ng/ml) compared to controls (1.87 ng/ml). Most of the Aβ in the serum of the immunized mice was bound to antibodies. We conclude that following active immunization, anti-Aβ antibodies sequester serum Aβ and may increase central nervous system to serum Aβ clearance.

Purification and characterization of astrocyte-secreted apolipoprotein E and J-containing lipoproteins from wild-type and human apoE transgenic mice

The varepsilon4 allele of apolipoprotein E (apoE) is a genetic risk factor for Alzheimers disease (AD). In order to gain a better understanding of the molecular mechanisms by which apoE and possibly other apolipoproteins produced in the central nervous system (CNS) influence AD pathogenesis, we have purified and characterized the two most abundant apolipoproteins produced in the CNS, apoE and apoJ. We purified apoE and apoJ from primary cultures of mouse astrocytes, which were derived from transgenic mice expressing human apoE isoforms in the absence of mouse apoE. Utilizing antibody affinity columns, we were able to purify both human apoE3 and apoE4, as well as mouse apoJ-containing lipoproteins. Astrocyte-secreted human apoE was present in high density-like lipoproteins of three predominant sizes ranging from 8 to 15 nm in diameter. Mouse apoJ was in particles between 10 and 17 nm in diameter with a peak size range of approximately 11 nm. ApoE and apoJ were in distinct lipoproteins. Utilization of quick-freeze, deep-etch electron microscopy revealed the apoE particles were discs while the apoJ particles were smaller and more irregular in appearance. The lipid composition of apoE particles was very different from those containing apoJ. ApoE-particles contained a similar mass of apoE and lipid, with cholesterol and phospholipid being about equal in mass per particle. ApoJ-particles were relatively lipid poor (three parts protein, one part lipid), with phospholipids being much more abundant than cholesterol. Detailed characterization of phospholipid composition by electrospray ionization mass spectrometry analysis revealed ethanolamine glycerophospholipids to be the most abundant phospholipid present in both apoE and apoJ particles. Analysis of cerebrospinal fluid from apoE3 and apoE4 transgenic mice revealed that human and mouse apoE were in particles the same size as those secreted by astrocytes. Further use of physiological preparations of CNS-derived lipoproteins may allow for a detailed understanding of the role of these molecules in the normal brain and in diseases such as AD.

Aβ immunization and anti-Aβ antibodies: potential therapies for the prevention and treatment of Alzheimer's disease

Abstract Amyloid-β (Aβ) is a normally soluble 39–43 amino peptide. Genetic and biochemical data strongly suggest that the conversion of Aβ from soluble to insoluble forms with high β-sheet content and its buildup in the brain is a key step in the pathogenesis of Alzheimer’s disease (AD) and cerebral amyloid angiopathy (CAA). Prevention and/or reversal of this process may serve as a treatment. Methods to prevent or reverse Aβ deposition and its toxic effects would include decreasing its production, preventing its conversion to insoluble forms (e.g. inhibit β-sheet formation) or in changing the dynamics of extracellular brain Aβ, either locally within the brain or by altering net flux of Aβ between the central nervous system (CNS) and plasma compartment. Transgenic mouse models of AD that develop age-dependent Aβ deposition, damage to the neuropil, and behavioral deficits have enabled researchers to test whether different manipulations can influence these AD-like changes. Recently, active immunization with different forms of the Aβ peptide has been shown to decrease brain Aβ deposition and improve cognitive performance in mouse models of AD. Certain peripherally administered anti-Aβ antibodies have similar effects. The mechanism(s) by which anti-Aβ antibodies result in these effects is just beginning to be elucidated. Aβ-related immune therapies in humans are an exciting new area of AD research. Understanding their detailed mechanism(s) of action and their potential usefulness awaits the results of future animal and human studies.