Exploration Of Alzheimer-type Amyloid Plaque Pathogenesis And Modulation

PART I Expression of BACE-1 in Alzheimer's diseaseObjective Determine the pattern of BACE-1 in the normal and AD samples.Methods Cortical samples totally of 17 patients with clinically diagnosed as AD or with a history of dementia (>3 years) and 14 nondemented patients were available for the present study. There were 7 rhesus monkeys with different ages (22-34 years-old). Nissl staining, bielschowsky staining, immunohistochemistry and western-blot methods were used for initial neuropathological screening and to make a pathologic diagnosis and experimental grouping. Furthermore, the normal and abnormal pattern of BACE-1 were detected with the immunohistochemistry.Results Those human cases with well preserved cortical cytoarchitecture in Nissl staining were sorted, and the cases selected for the present study met the following conditions:AD cases (n=7) showed both senile plaques and p-Tau pathology, whereas control cases (n=7) were free of either lesion. Aged group (n=3) and normal group (n=4) of monkey were evaluated using the same criterions as human samples. Western blot showed the elevations of some AD-related protein such as p-Tau, APP, BACE-1 and theβ-site cleavage APP C-terminal fragment (β-CTF) in the diseased brains relative to controls. Levels of the different molecules of BACE-1 proteins are elevated in the aged monkey cortices with cerebral plaques relative to plaques-free cases. BACE-1 immunohistochemistry staining showed that the pattern of BACE-1 was similar to the normal pattern in mouse. The strongest and distinct BACE-1 labeling was expressed in the olfactory bulb and hippocampus fibers and diffuse and in background-like neuropil reactivity occurred in the cortex. The elevation of BACE-1-ir plaques-like profile occurred in the AD and aged rhesus monkeys, appearing as different size and various shape. Conclusion Elevation of BACE-1 maybe promote amyloid plaques formation in the AD and old rhesus monkey PART II The relationship among elevation of BACE-1,Aβdeposition and dystrophic neuritesObjective To explore the relationship among elevation of BACE-1, A(3 deposition and dystrophic neuritis in the AD patients and AD-like rhesus monkey and reveal the potential process of amyloid pathogenesis in AD.Methods To investigate the time-space relationship of the BACE-1-ir dystrophic neuritis with some AD-related protein such asβ-amyloid and presenilin-1 in the adjacent tissues with immunohistochemistry. In addition, the nature of the dystrophic neuritis was determined with double staining of BACE-1 with synaptophysin or microtubule-associated protein-2 (MAP-2) immunofluoresence double staining.Results In adjacent sections from AD cases, BACE-1 labeled profiles appear to match site-specifically with plaques in the cortex with relatively mild plaque load. While in AD cortex with heavier amyloid load, BACE-1 labeled profiles appeared to be less numerous than Aβplaques. Double immunofluoresence from aged monkey case showed that elevated BACE-1 localized in the dystrophic axons (BACE-1 co-localized with SYN)and co-localized with some AD related protein such as APP (substrate of BACE-1), PS 1 (γ-secretase of APP) and Aβ. In the swollen axon, elevated BACE-1 can co-exist locally with A(3 completely and there was no any clear extracellular Aβdeposits surrounded. Among the small plaques, Aβ-ir was relatively weak and turned stronger with the continuous swelling and sprouting dystrophic neurites. When the BACE-1-labeled dystrophic neurite arranged in a rosette-like fashion, there were plenty of densed extracelluar deposition existed.Conclusion1. The onset of the Aβoverproduction maybe derived from the dystrophic axons 2. BACE-1 maybe modulate the dystrophic neurites and contribute to the over-load Aβ PART III The experimental exploration about the BACE-1 elevation and amyloid plaques formationObjective To determine reduced neuronal activity promote BACE-1 and the amyloid plaques formation in Tg2576 mouse olfactory centersMethods Adult Tg2576 mice (n=28) were subjected to the unilateral naris-occlusion on 6 month-old, occluded animals were allowed to survive until 7,8,9,12,18,24 months-old. CO staining, immunohistochemistry, western blot and ELISA were used for detecting the effect of the functional deprivation on the expression of BACE-1 and amyloid plaques in Tg2576 mouse olfactory centers.Results Adjacent sections revealed an inverse modulation between BACE-land CO reactivities in the olfactory bulb and piriform cortex. BACE-1 immunoreactivity is clearly stronger while CO reactivity is clearly lower on the deprived side, which meaned the animal model is successful. Elevations of BACE-1 proteins, enzymatic activity and immediate and end products ofβ-cleavage pathway of APP can be seen in the deprived olfactory bulbs relative to counterparts. The adjacent sections of BACE-1 and A(3 immunohistochemistry staining showed that the amyloid pathogenesis in Tg2576 mouse olfactory bulbs and piriform cortex is age-dependency. BACE-1 labeled neuritis and A(3 deposition occurred firstly in the piriform cortex and then in the bulb. Double immunofluoresence staining of BACE-land Aβrevealed that BACE-1 localized in the dystrophic axons (BACE-1 and SYN colocalization) in Tg2576 mouse olfactory bulbs and piriform cortex. Elevation of BACE-1 colocalized with A(3 and the amyloid plaques formation was similar to Part II.ConclusionLower neuronal activity/hypometablolism maybe one of the upstream mechanisms of BACE-1 elevation and amyloid plaques formation...