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In vivo Differential Brain Clearance and Catabolism of Monomeric and Oligomeric Alzheimer's A β protein

View Article: PubMed Central - PubMed

ABSTRACT

Amyloid β (Aβ) is the major constituent of the brain deposits found in parenchymal plaques and cerebral blood vessels of patients with Alzheimer's disease (AD). Several lines of investigation support the notion that synaptic pathology, one of the strongest correlates to cognitive impairment, is related to the progressive accumulation of neurotoxic Aβ oligomers. Since the process of oligomerization/fibrillization is concentration-dependent, it is highly reliant on the homeostatic mechanisms that regulate the steady state levels of Aβ influencing the delicate balance between rate of synthesis, dynamics of aggregation, and clearance kinetics. Emerging new data suggest that reduced Aβ clearance, particularly in the aging brain, plays a critical role in the process of amyloid formation and AD pathogenesis. Using well-defined monomeric and low molecular mass oligomeric Aβ1-40 species stereotaxically injected into the brain of C57BL/6 wild-type mice in combination with biochemical and mass spectrometric analyses in CSF, our data clearly demonstrate that Aβ physiologic removal is extremely fast and involves local proteolytic degradation leading to the generation of heterogeneous C-terminally cleaved proteolytic products, while providing clear indication of the detrimental role of oligomerization for brain Aβ efflux. Immunofluorescence confocal microscopy studies provide insight into the cellular pathways involved in the brain removal and cellular uptake of Aβ. The findings indicate that clearance from brain interstitial fluid follows local and systemic paths and that in addition to the blood-brain barrier, local enzymatic degradation and the bulk flow transport through the choroid plexus into the CSF play significant roles. Our studies highlight the diverse factors influencing brain clearance and the participation of various routes of elimination opening up new research opportunities for the understanding of altered mechanisms triggering AD pathology and for the potential design of combined therapeutic strategies.

No MeSH data available.


Related in: MedlinePlus

Brain clearance of radiolabeled Aβ species. (A) Clearance of monomeric (black bars) and oligomeric (gray bars) [125I]Aβ1-40 species removed from the brain at three different time-points. (B) Monomeric (black bars) and oligomeric (gray bars) [125I]Aβ1-40 species cleared to the CSF evaluated at three different time-points. In all cases bars illustrate percentage cleared relative to total injected radioactivity; values represent mean ± SD obtained from inoculation of 5–7 mice (ANOVA and Tukey post-hoc test; ns, not significant; *p < 0.05; **p < 0.01; ***p < 0.001).
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Figure 2: Brain clearance of radiolabeled Aβ species. (A) Clearance of monomeric (black bars) and oligomeric (gray bars) [125I]Aβ1-40 species removed from the brain at three different time-points. (B) Monomeric (black bars) and oligomeric (gray bars) [125I]Aβ1-40 species cleared to the CSF evaluated at three different time-points. In all cases bars illustrate percentage cleared relative to total injected radioactivity; values represent mean ± SD obtained from inoculation of 5–7 mice (ANOVA and Tukey post-hoc test; ns, not significant; *p < 0.05; **p < 0.01; ***p < 0.001).

Mentions: Figure 2A depicts the time-related clearance of monomeric and LMW oligomeric [125I]Aβ1-40 from brain interstitial fluid, estimated based on the radioactivity remaining in the whole brain at 5, 30, and 60 min post-injection compared to the total injected radioactivity. Brain efflux of monomeric Aβ1-40 was fast, with ~25% of the peptide cleared in only 5 min and >60% cleared within 60 min. Retention of LMW oligomers was consistently higher, with only ~15% of the injected material eliminated in the first 5 min and < 40% at 60 min. Comparison of brain clearance levels for monomeric vs. oligomeric forms of [125I]Aβ1-40 evaluated 60 min after intra-cerebral injection clearly demonstrated that peptide oligomerization significantly increased brain retention (p < 0.001). Clearance from interstitial fluid to the CSF was evaluated by quantitating radioactivity in the CSF 5, 30, and 60 min post-Aβ intracerebral injection (Figure 2B). CSF was collected through a cisterna magna puncturing technique without blood contamination, as corroborated by dot blot analysis probed with antibodies immunoreacting with apolipoprotein-B and α2-macrogloblin, blood proteins not present in the CSF (data not shown). Only a fraction of the material removed from brain reached the CSF in a time-dependent manner. In the case of monomeric Aβ1-40, the levels cleared to the CSF increased from ~3% at 5 min to an average of 13% at 60 min. In contrast, LMW oligomeric Aβ1-40 was cleared less efficiently, with only ~2% present in the CSF after 5 min, and ~4% at 60 min. Comparative evaluation of [125I]-Aβ cleared to the CSF at 60 min indicates a statistically significant decrease of about 3-fold in oligomeric Aβ removal compared to the values obtained for the monomeric form of the peptide (p < 0.05) and in line with the results illustrated in Figure 2A for overall brain efflux.


In vivo Differential Brain Clearance and Catabolism of Monomeric and Oligomeric Alzheimer's A β protein
Brain clearance of radiolabeled Aβ species. (A) Clearance of monomeric (black bars) and oligomeric (gray bars) [125I]Aβ1-40 species removed from the brain at three different time-points. (B) Monomeric (black bars) and oligomeric (gray bars) [125I]Aβ1-40 species cleared to the CSF evaluated at three different time-points. In all cases bars illustrate percentage cleared relative to total injected radioactivity; values represent mean ± SD obtained from inoculation of 5–7 mice (ANOVA and Tukey post-hoc test; ns, not significant; *p < 0.05; **p < 0.01; ***p < 0.001).
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Related In: Results  -  Collection

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Figure 2: Brain clearance of radiolabeled Aβ species. (A) Clearance of monomeric (black bars) and oligomeric (gray bars) [125I]Aβ1-40 species removed from the brain at three different time-points. (B) Monomeric (black bars) and oligomeric (gray bars) [125I]Aβ1-40 species cleared to the CSF evaluated at three different time-points. In all cases bars illustrate percentage cleared relative to total injected radioactivity; values represent mean ± SD obtained from inoculation of 5–7 mice (ANOVA and Tukey post-hoc test; ns, not significant; *p < 0.05; **p < 0.01; ***p < 0.001).
Mentions: Figure 2A depicts the time-related clearance of monomeric and LMW oligomeric [125I]Aβ1-40 from brain interstitial fluid, estimated based on the radioactivity remaining in the whole brain at 5, 30, and 60 min post-injection compared to the total injected radioactivity. Brain efflux of monomeric Aβ1-40 was fast, with ~25% of the peptide cleared in only 5 min and >60% cleared within 60 min. Retention of LMW oligomers was consistently higher, with only ~15% of the injected material eliminated in the first 5 min and < 40% at 60 min. Comparison of brain clearance levels for monomeric vs. oligomeric forms of [125I]Aβ1-40 evaluated 60 min after intra-cerebral injection clearly demonstrated that peptide oligomerization significantly increased brain retention (p < 0.001). Clearance from interstitial fluid to the CSF was evaluated by quantitating radioactivity in the CSF 5, 30, and 60 min post-Aβ intracerebral injection (Figure 2B). CSF was collected through a cisterna magna puncturing technique without blood contamination, as corroborated by dot blot analysis probed with antibodies immunoreacting with apolipoprotein-B and α2-macrogloblin, blood proteins not present in the CSF (data not shown). Only a fraction of the material removed from brain reached the CSF in a time-dependent manner. In the case of monomeric Aβ1-40, the levels cleared to the CSF increased from ~3% at 5 min to an average of 13% at 60 min. In contrast, LMW oligomeric Aβ1-40 was cleared less efficiently, with only ~2% present in the CSF after 5 min, and ~4% at 60 min. Comparative evaluation of [125I]-Aβ cleared to the CSF at 60 min indicates a statistically significant decrease of about 3-fold in oligomeric Aβ removal compared to the values obtained for the monomeric form of the peptide (p < 0.05) and in line with the results illustrated in Figure 2A for overall brain efflux.

View Article: PubMed Central - PubMed

ABSTRACT

Amyloid &beta; (A&beta;) is the major constituent of the brain deposits found in parenchymal plaques and cerebral blood vessels of patients with Alzheimer's disease (AD). Several lines of investigation support the notion that synaptic pathology, one of the strongest correlates to cognitive impairment, is related to the progressive accumulation of neurotoxic A&beta; oligomers. Since the process of oligomerization/fibrillization is concentration-dependent, it is highly reliant on the homeostatic mechanisms that regulate the steady state levels of A&beta; influencing the delicate balance between rate of synthesis, dynamics of aggregation, and clearance kinetics. Emerging new data suggest that reduced A&beta; clearance, particularly in the aging brain, plays a critical role in the process of amyloid formation and AD pathogenesis. Using well-defined monomeric and low molecular mass oligomeric A&beta;1-40 species stereotaxically injected into the brain of C57BL/6 wild-type mice in combination with biochemical and mass spectrometric analyses in CSF, our data clearly demonstrate that A&beta; physiologic removal is extremely fast and involves local proteolytic degradation leading to the generation of heterogeneous C-terminally cleaved proteolytic products, while providing clear indication of the detrimental role of oligomerization for brain A&beta; efflux. Immunofluorescence confocal microscopy studies provide insight into the cellular pathways involved in the brain removal and cellular uptake of A&beta;. The findings indicate that clearance from brain interstitial fluid follows local and systemic paths and that in addition to the blood-brain barrier, local enzymatic degradation and the bulk flow transport through the choroid plexus into the CSF play significant roles. Our studies highlight the diverse factors influencing brain clearance and the participation of various routes of elimination opening up new research opportunities for the understanding of altered mechanisms triggering AD pathology and for the potential design of combined therapeutic strategies.

No MeSH data available.


Related in: MedlinePlus