D. Larry Sparks

Induction of Alzheimer-like β-Amyloid Immunoreactivity in the Brains of Rabbits with Dietary Cholesterol

beta-amyloid and ALZ-50 immunocytochemical reactivity were determined in the brains of rabbits fed either a control or 2% cholesterol diet. Control rabbits demonstrated no accumulation of intracellular immunolabeled beta-amyloid within 3 min after death. In animals fed the experimental diet for 4, 6, and 8 weeks (postmortem interval < 3 min), there was an increasingly mild-to-moderate-to-severe accumulation of intracellular immunolabeled beta-amyloid. Whether or not beta-amyloid is causally linked to processes leading to dementia, it is related in some way to the prime cause of human death; heart disease. Hypercholesterolemic rabbits may provide an animal model to study altered beta-APP metabolism leading to Alzheimer-like beta-amyloid accumulation xe03and extracellular deposition in brain.

Trace amounts of copper in water induce β-amyloid plaques and learning deficits in a rabbit model of Alzheimer's disease

Despite the crucial role played by cholesterol and copper in nutrition and normal brain function, recent evidence indicates that they may both be important factors in the etiology of Alzheimers disease (AD). Here we provide critical evidence for the role of cholesterol and copper in AD by showing that the addition of trace amounts of copper (0.12 ppm) to water given to cholesterol-fed rabbits can induce β-amyloid (Aβ) accumulation, including senile plaque-like structures in the hippocampus and temporal lobe, and can significantly retard the ability of rabbits to learn a difficult trace conditioning task. The Aβ deposits do not affect the ability of rabbits to detect or respond to the training stimuli nor to learn a simpler delay conditioning task. Trace amounts of copper in drinking water may influence clearance of Aβ from the brain at the level of the interface between the blood and cerebrovasculature and combined with high cholesterol may be a key component to the accumulation of Aβ in the brain, having a significant impact on learning and memory. Cholesterol-fed rabbits have at least 12 pathological markers seen in AD, suggesting that the cholesterolfed rabbit is a good animal model for studying AD.

Cortical senile plaques in coronary artery disease, aging and Alzheimer's disease

Mild alterations in cognitive function are present in normal aging and severe cognitive alterations are a hallmark of Alzheimers disease (AD). The cognitive change in AD has been correlated to the characteristic pathologic lesions in the brain, senile plaques (SP) and neurofibrillary tangles. Senile plaques are the most consistent correlative marker in AD. We present preliminary data indicating that abundant SP are found in the brains of nondemented patients dying with or as a result of critical coronary artery disease (cCAD) compared to nonheart disease (non-HD) subjects; 15 of 20 cCAD patients contained SP and only two of 16 non-HD patients contained SP.

Increased incidence of neurofibrillary tangles (NFT) in non-demented individuals with hypertension.

The incidence rates and numerical densities of argryophilic neurofibrillary tangles (NFT) and senile plaques (SP) were determined in non-demented individuals and subjects with Alzheimers disease (AD). The non-AD subjects were grouped according to cardiac status; those individuals with critical coronary artery disease (cCAD), those hypertensive individuals without cCAD (HyperT), and those without heart disease (non-HD). The incidence and densities of SP and NFT were significantly greater in AD than any of the non-demented groups. The prevalence of SP was increased in both HyperT and cCAD compared to non-HD controls, while NFT occurrence was accentuated in non-demented HyperT subjects only. The densities of SP and NFT in HyperT were elevated compared to cCAD or both cCAD and non-HD controls; NFT densities were similar in cCAD and non-HD. NFT density increased with increasing age in only the non-HD and cCAD groups, suggesting a possible relationship between disease process and NFT formation in the AD and HyperT populations.

Alterations of Alzheimer's disease in the cholesterol-fed rabbit, including vascular inflammation. Preliminary observations.

Abstract: We determined the levels of endothelial inflammation using MECA‐32 antibody and a4 nicotinic receptor subunit densities employing [3H]epibatidine binding in the brains of Alzheimers disease (AD) patients, cholesterol‐fed rabbits, and appropriate controls. We also assessed rabbit brain for β‐amyloid levels and immunohistochemical localization, and for evidence of blood‐brain barrier breach using normally‐excluded Evans Blue dye. Dietary cholesterol induced a twofold increase in β‐amyloid concentration in rabbit hippocampal cortex, which may be related to the appearance of β‐amyloid immunoreactivity in the neuropil. Epibatidine binding was significantly decreased in AD superior frontal cortex, but unchanged in the superior frontal cortex of cholesterol‐fed rabbits. Increased vascular MECA‐32 immunoreactivity occurred in AD and cholesterol‐fed rabbit brain. Evans Blue dye could be found in the parenchyma of cholesterol‐fed rabbits only, and appeared as pockets of dye surrounding small blood vessels. The data suggest that vascular inflammation can lead to breach of the blood‐brain barrier, which may produce biochemical derangements in surrounding brain tissue that are conducive to production of β‐amyloid.

Activation of microglia in the brains of humans with heart disease and hypercholesterolemic rabbits

Abstract Activated microglial cells are concentrated in senile plaques characteristic of Alzheimer’s disease. Such accumulations of activated microglia may contribute towards neurodegeneration via production of cytokines and free radicals. Studies suggesting a link between Alzheimer’s disease and heart disease led us to study microglia immunohistochemically, using monoclonal antibody LN-3, in age-matched nondemented humans with and without heart disease. Using a qualitative staging system for assessing morphological changes occurring in microglia, we found higher microglial activation in the brains of subjects with heart disease than in those without it. Lectin histochemical examination of brains from rabbits maintained on a high-cholesterol diet also revealed increased microglial activation and leukocyte infiltration. Collectively our observations from humans and rabbits suggest that hypercholesterolemia and heart disease accelerate brain aging, and that the formation of senile plaques may be the end result of progressive microglial activation that occurs with aging.

Intraneuronal β-amyloid immunoreactivity in the CNS

The high degree of overlap in the neuropathology outcome of Alzheimers disease (AD), Downs Syndrome (DS), and coronary heart disease suggest a possible interrelationship. The pattern of hippocampal and cortical intraneuronal βA4 immunoreactivity is strikingly similar in AD, DS, coronary heart disease, and two separate animal models of coronary heart disease. Cells in fascia dentata and large cortical neurons were βA4 immunedecorated in half the AD and DS subjects studied. Similar neuronal staining occurred in half the age-matched coronary heart disease subjects, but was absent in each nonheart disease control investigated. Analogous accumulations of neuronal βA4 immunoreactivity were induced in rabbit brain by dietary administration of high cholesterol, and this effect could be reversed by regression of the experimental diet. Decreased density (p < 0.05) and cellular staining intensity occurred after 2 weeks of control diet following 8 weeks of high cholesterol. Microgliosis accompanied the accumulation of βA4 immunoreactivity in the cholesterol-fed rabbits and persisted after regression of the diet and decreases in neuronal βA4 immunoreactivity. An identical pattern of neuronal βA4 immunoreactivity was induced in the brains of adolescent pigs after acute ligation of the left anterior descending coronary artery (LAD) compared to surgical and anesthetic controls. The mean number of βA4 immunoreactive neurons was significantly increased (p < 0.05) in the cortex and hippocampus of pigs with a ligated LAD compared to both control groups. Increased density and intensity of neuronal βA4 immunoreactivity induced by ligation of the LAD was commensurate with the severity of the decreased cardiac output in the LAD group, but not in the anesthetic control groups with decreased cardiac output. The incidence of ALZ-50 (A68) immunoreactive neurons also increased in the ligated pigs compared to both control groups. The data suggest a neuronal origin of βA4 immunoreactive peptide(s), which can be cleared from the brain by microglia after severe accumulation is induced. This could indicate that reduced clearance of β-APP metabolic by-products could contribute to a metabolic backlog and redirection of peptide processing by microglia to extracellular deposition. Neuronal accumulation of βA4 immunoreactivity could be due to the effect of circulating factors on brain function in both animals models. It is likely that animal models of coronary heart disease may be useful in disclosing the mechanism of SP formation and induction of ALZ-50 immunoreactivity irrespective of their pathoclinical significance.

Temporal Sequence of Plaque Formation in the Cerebral Cortex of Non-Demented Individuals

One of the hallmarks of Alzheimers disease is the presence of argyrophilic plaques (arg-P) accompanying dementia and other forms of cognitive alterations. In the present investigation 195 non-demented, cognitively normal patients were grouped according to the presence or absence of critical coronary artery disease (cCAD), defined as a 75% or greater stenosis of one of the epicardial arteries. None of the subjects had significant cerebral vascular disease. The parahippocampal gyrus (PHG) and frontal pole were analyzed for the presence of arg-P, A4 deposition, ALZ-50 immunoreactive (IR) neurons and neuropil threads (NT). Individuals with cCAD have a significantly greater incidence of plaques than non-heart disease (non- HD) subjects. Every cCAD subject had ALZ-50 IR neurons in the PHG and a greater incidence of NT as compared to the non-HD subjects. Every subject with plaques also had IR neurons and NT in the PHG. Based on the presumption that early neurodegeneration labeled by ALZ-50 antibody and amyloid deposition are in some way linked, then the sequence of plaque formation is initiated by the presence of ALZ-50 IR neurons followed in order by NT, A4 deposition and diffuse form arg-P.

Relationship of cognitive measures and gray and white matter in Alzheimer's disease

OBJECTIVE To examine the relationship between commonly used screening cognitive measures with gray and white matter integrity in patients with mild to moderate AD. BACKGROUND New neuroimaging techniques, such as voxel-based morphometry (VBM), make it possible to study the relationship between structural brain integrity and cognitive functioning in AD. METHODS Gray and white matter integrity was evaluated using VBM in fifteen patients with mild to moderate AD. ADAS-Cog and MMSE scores were also performed as part of the baseline assessment for a larger clinical trial in the AD patients. Correlations between cognitive measures and VBM were performed. RESULTS Both the ADAS-Cog and the MMSE showed a similar relationship with gray matter degeneration, reflecting greater cognitive impairment with decreased gray matter in the left temporal lobe. However, the MMSE score was much more reflective of underlying white matter changes than ADAS-Cog scores, particularly in frontotemporal region. These findings suggest that the ADAS-Cog and MMSE reflect different aspects of the underlying brain changes observed in AD. The ADAS-Cog was more specific to gray matter integrity whereas the MMSE reflected a more global reduction in both gray and white matter. CONCLUSIONS These results support using neuroimaging markers of neural integrity as an important consideration when evaluating treatment efficacy. Furthermore, whole-brain analyses such as VBM help to evaluate neural systems that are not necessarily targeted by the treatment.

Immunohistochemical Localization of a Cytochrome P-450 Isozyme in Human Nasal Mucosa: Age-Related Trends

Immunoperoxidase staining with an antibody to cytochrome P-450 (NMa) was used to investigate the localization of this isozyme in the human nasal mucosa. Olfactory mucosa was identified by staining of olfactory receptor cells with an antibody to olfactory marker protein. Immunoreactivity to NMa was localized in sustentacular cells in the olfactory epithelium, and in Bowmans gland acinar cells and vascular endothelial cells in the lamina propria. In the respiratory mucosa, ciliated epithelial cells, as well as serous gland acinar cells and vascular endothelial cells in the lamina propria, were immunoreactive for this isozyme. An age-related decrement in the intensity and extent of immunoreactivity within these cells was noted in nasal tissue from patients 60 years of age and over when compared with that of patients under 60 years of age. These results identify sites of xenobiotic metabolism or activation in human nasal mucosa.