ABSTRACT
The accumulation of amyloid-beta into insoluble plaques is a characteristic feature of Alzheimer's disease. Neuronal morphology is distorted by plaques: rather than being essentially straight, they are substantially more curved than those in control tissue, their trajectories become altered, and they are frequently distended or swollen, presumably affecting synaptic transmission. Clearance of plaques by administration of antibodies to amyloid-beta is a promising therapeutic approach to the treatment of Alzheimer's disease, leading to stabilization of dementia by an unknown cellular mechanism. The effect of plaque clearance on plaque-induced neuronal alterations has not been studied previously. Here we show that both plaques and neuritic lesions are reversible in a strikingly short period of time after administration of a single dose of amyloid-beta antibody. Amyloid clearance and recovery of normal neuronal geometries were observed as early as 4 d and lasted at least 32 d after a single treatment. These results demonstrate that, once plaques are cleared, neuronal morphology is self-correcting and that passive antibody treatment has the potential to reverse neuronal damage caused by Alzheimer's disease and, hence, directly impact cognitive decline. Moreover, the rapid normalization of neuritic dystrophy suggests an unexpected degree of plasticity in the adult nervous system.
Subject(s)
Alzheimer Disease/therapy , Amyloid beta-Peptides/immunology , Antibodies/therapeutic use , Neurites/drug effects , Plaque, Amyloid/drug effects , Age Factors , Alzheimer Disease/pathology , Alzheimer Disease/physiopathology , Amyloid beta-Peptides/antagonists & inhibitors , Animals , Cerebral Cortex/drug effects , Cerebral Cortex/immunology , Cerebral Cortex/pathology , Disease Models, Animal , Drug Administration Routes , Humans , Immunization, Passive/methods , Mice , Mice, Inbred C57BL , Mice, Transgenic , Neurites/immunology , Neurites/pathology , Neuronal Plasticity/drug effects , Plaque, Amyloid/immunology , Plaque, Amyloid/pathology , Treatment OutcomeABSTRACT
Amyloid-beta, the primary constituent of senile plaques in Alzheimer's disease, is hypothesized to cause neuronal damage and cognitive failure, but the mechanisms are unknown. Using multiphoton imaging, we show a direct association between amyloid-beta deposits and free radical production in vivo in live, transgenic mouse models of Alzheimer's disease and in analogous ex vivo experiments in human Alzheimer tissue. We applied two fluorogenic compounds, which become fluorescent only after oxidation, before imaging with a near infrared laser. We observed fluorescence associated with dense core plaques, but not diffuse plaques, as determined by subsequent addition of thioflavine S and immunohistochemistry for amyloid-beta. Systemic administration of N-tert-butyl-alpha-phenylnitrone, a free radical spin trap, greatly reduced oxidation of the probes. These data show directly that a subset of amyloid plaques produces free radicals in living, Alzheimer's models and in human Alzheimer tissue. Antioxidant therapy neutralizes these highly reactive molecules and may therefore be of therapeutic value in Alzheimer's disease.
Subject(s)
Alzheimer Disease/diagnosis , Alzheimer Disease/metabolism , Plaque, Amyloid/metabolism , Reactive Oxygen Species/metabolism , Thiazoles/metabolism , Amyloid beta-Peptides/metabolism , Amyloid beta-Protein Precursor/genetics , Amyloid beta-Protein Precursor/metabolism , Animals , Antioxidants/pharmacology , Benzothiazoles , Cerebral Cortex/drug effects , Cerebral Cortex/metabolism , Cyclic N-Oxides , Disease Models, Animal , Fluorescent Dyes/metabolism , Free Radical Scavengers/pharmacology , Free Radicals/metabolism , Infrared Rays , Lasers , Mice , Mice, Transgenic , Microscopy, Fluorescence, Multiphoton , Nitrogen Oxides/pharmacology , Oxidation-Reduction/drug effects , Oxidative Stress/drug effects , Plaque, Amyloid/classification , Reactive Oxygen Species/analysisABSTRACT
The identification of amyloid deposits in living Alzheimer disease (AD) patients is important for both early diagnosis and for monitoring the efficacy of newly developed anti-amyloid therapies. Methoxy-X04 is a derivative of Congo red and Chrysamine-G that contains no acid groups and is therefore smaller and much more lipophilic than Congo red or Chrysamine-G. Methoxy-X04 retains in vitro binding affinity for amyloid beta (Abeta) fibrils (Ki = 26.8 nM) very similar to that of Chrysamine-G (Ki = 25.3 nM). Methoxy-X04 is fluorescent and stains plaques, tangles, and cerebrovascular amyloid in postmortem sections of AD brain with good specificity. Using multiphoton microscopy to obtain high-resolution (1 microm) fluorescent images from the brains of living PSI/APP mice, individual plaques could be distinguished within 30 to 60 min after a single i.v. injection of 5 to 10 mg/kg methoxy-X04. A single i.p. injection of 10 mg/kg methoxy-X04 also produced high contrast images of plaques and cerebrovascular amyloid in PSI/APP mouse brain. Complementary quantitative studies using tracer doses of carbon- 11-labeled methoxy-X04 show that it enters rat brain in amounts that suggest it is a viable candidate as a positron emission tomography (PET) amyloid-imaging agent for in vivo human studies.
Subject(s)
Alkenes/pharmacokinetics , Alzheimer Disease/diagnosis , Amyloid beta-Peptides/analysis , Benzene/pharmacokinetics , Congo Red/analogs & derivatives , Microscopy/methods , Plaque, Amyloid/pathology , Alkenes/chemistry , Alzheimer Disease/pathology , Amyloid beta-Peptides/chemistry , Amyloid beta-Peptides/metabolism , Animals , Benzene/chemistry , Benzene Derivatives , Binding, Competitive/drug effects , Blood-Brain Barrier , Carbon Radioisotopes , Coloring Agents/chemistry , Coloring Agents/pharmacokinetics , Congo Red/chemistry , Disease Models, Animal , Imaging, Three-Dimensional , Male , Mice , Mice, Transgenic , Peptide Fragments/chemistry , Plaque, Amyloid/metabolism , Rats , Rats, Sprague-Dawley , Sensitivity and Specificity , StilbenesABSTRACT
Transgenic (Tg) mouse models overexpressing amyloid precursor protein (APP) develop senile plaques similar to those found in Alzheimer's disease in an age-dependent manner. Recent reports demonstrated that immunotherapy is effective at preventing or removing amyloid-beta deposits in the mouse models. To characterize the mechanisms involved in clearance, we used antibodies of either IgG1 (10d5) or IgG2b (3d6) applied directly to the brains of 18-month-old Tg2576 or 20-month-old PDAPP mice. Both 10d5 and 3d6 led to clearance of 50% of diffuse amyloid deposits in both animal models within 3 d. Fc receptor-mediated clearance has been shown to be important in an ex vivo assay showing antibody-mediated clearance of plaques by microglia. We now show, using in vivo multiphoton microscopy, that FITC-labeled F(ab')2 fragments of 3d6 (which lack the Fc region of the antibody) also led to clearance of 45% of the deposits within 3 d, similar to the results obtained with full-length 3d6 antibody. This result suggests that direct disruption of plaques, in addition to Fc-dependent phagocytosis, is involved in the antibody-mediated clearance of amyloid-beta deposits in vivo. Dense-core deposits that were not cleared were reduced in size by approximately 30% with full-length antibodies and F(ab')2 fragments 3 d after a topical treatment. Together, these results indicate that clearance of amyloid deposits in vivo may involve, in addition to Fc-dependent clearance, a non-Fc-mediated disruption of plaque structure.
Subject(s)
Alzheimer Disease/metabolism , Amyloid beta-Peptides/metabolism , Immunoglobulin Fc Fragments/metabolism , Immunotherapy , Administration, Topical , Alzheimer Disease/immunology , Alzheimer Disease/pathology , Alzheimer Disease/therapy , Amyloid beta-Peptides/immunology , Amyloid beta-Protein Precursor/genetics , Amyloid beta-Protein Precursor/metabolism , Animals , Antibodies/administration & dosage , Benzothiazoles , Brain/metabolism , Brain/pathology , Disease Models, Animal , Fluorescent Dyes , Humans , Immunoglobulin Fab Fragments/pharmacology , Mice , Mice, Transgenic , Microscopy, Fluorescence/methods , Mutation , Phagocytosis/immunology , Plaque, Amyloid/drug effects , Plaque, Amyloid/metabolism , ThiazolesABSTRACT
The synthesis of a new lipophilic thioflavin-T analogue (2-[4' -(methylamino)phenyl]benzothiazole, 6) with high affinity for amyloid is reported. Intravenous injection of [(11)C]-labeled 6 in control mice resulted in high brain uptake. Amyloid deposits were imaged with multiphoton microscopy in the brains of living transgenic mice following the systemic injection of unlabeled 6. [(11)C]6 is a promising amyloid imaging agent for Alzheimer's disease.
