γ‐Secretase binding sites in aged and Alzheimer's disease human cerebrum: the choroid plexus as a putative origin of CSF Aβ

Deposition of β ‐amyloid (Aβ) peptides, cleavage products of β‐amyloid precursor protein (APP) by β‐secretase‐1 (BACE1) and γ‐secretase, is a neuropathological hallmark of Alzheimer's disease (AD). γ‐Secretase inhibition is a therapeutical anti‐Aβ approach, although changes in the enzyme's activity in AD brain are unclear. Cerebrospinal fluid (CSF) Aβ peptides are thought to derive from brain parenchyma and thus may serve as biomarkers for assessing cerebral amyloidosis and anti‐Aβ efficacy. The present study compared active γ‐secretase binding sites with Aβ deposition in aged and AD human cerebrum, and explored the possibility of Aβ production and secretion by the choroid plexus (CP). The specific binding density of [³H]‐L‐685,458, a radiolabeled high‐affinity γ‐secretase inhibitor, in the temporal neocortex and hippocampal formation was similar for AD and control cases with similar ages and post‐mortem delays. The CP in post‐mortem samples exhibited exceptionally high [³H]‐L‐685,458 binding density, with the estimated maximal binding sites (Bmax) reduced in the AD relative to control groups. Surgically resected human CP exhibited APP, BACE1 and presenilin‐1 immunoreactivity, and β‐site APP cleavage enzymatic activity. In primary culture, human CP cells also expressed these amyloidogenic proteins and released Aβ40 and Aβ42 into the medium. Overall, our results suggest that γ‐secretase activity appears unaltered in the cerebrum in AD and is not correlated with regional amyloid plaque pathology. The CP appears to be a previously unrecognised non‐neuronal contributor to CSF Aβ, probably at reduced levels in AD.     

Upregulation of RAGE at the blood‐brain barrier in streptozotocin‐induced diabetic mice

Deposition of amyloid‐β peptide (Aβ) in the brain of diabetes is poorly understood. The receptor for advanced glycation end products (RAGE) at the blood‐brain barrier (BBB) is critical for regulation of Aβ homeostasis in the brain. In this studies, we used streptozotocin‐induced diabetic mice to observe the expression of RAGE at the BBB by Western blot and immunocytochemical analysis, and the in vivo blood‐to‐brain influx transport of ¹²⁵I‐Aβ_1–40 using the permeability surface area product (PS) and brain capillary uptake. In the diabetic mice with hyperglycemia (>16.0 mmol/L) at 6 weeks, RAGE expression at the BBB was significantly upregulated, no significant changes of RAGE levels were found at 1 and 3 weeks after diabetes induction. The data of PS and brain capillary uptake for Aβ showed significant RAGE‐dependent transport of Aβ across the BBB and substantial RAGE‐dependent brain capillary uptake at 6 weeks after diabetes induction. We conclude that the upregulation of RAGE at the BBB contributes to cerebral Aβ deposition in the diabetes. Synapse 63:636–642, 2009. © 2009 Wiley‐Liss, Inc.     

Limitations of the hCMEC/D3 cell line as a model for Aβ clearance by the human blood‐brain barrier

Alzheimer's disease and cerebral amyloid angiopathy are characterized by accumulation of amyloid‐β (Aβ) at the cerebrovasculature due to decreased clearance at the blood‐brain barrier (BBB). However, the exact mechanism of Aβ clearance across this barrier has not been fully elucidated. The hCMEC/D3 cell line has been characterized as a valid model for the BBB. In this study we evaluated the use of this model to study Aβ clearance across the BBB, with an emphasis on brain‐to‐blood directional permeability. Barrier integrity of hCMEC/D3 monolayers was confirmed for large molecules in both the apical to basolateral and the reverse direction. However, permeability for smaller molecules was substantially higher, especially in basolateral to apical direction, and barrier formation for Aβ was completely absent in this direction. In addition, hCMEC/D3 cells failed to develop a high TEER, possibly caused by incomplete formation of tight junctions. We conclude that the hCMEC/D3 model has several limitations to study the cerebral clearance of Aβ. Therefore, the model needs further characterization before this cell system can be generally applied as a model to study cerebral Aβ clearance. © 2016 The Authors Journal of Neuroscience Research Published by Wiley Periodicals, Inc.     

REVIEW: γ‐Secretase Inhibitors for the Treatment of Alzheimer's Disease: The Current State

Aims: Drugs currently used for the treatment of Alzheimer's disease (AD) partially stabilize patients’ symptoms without modifying disease progression. Brain accumulation of oligomeric species of β‐amyloid (Aβ) peptides, the principal components of senile plaques, is believed to play a crucial role in the development of AD. Based on this hypothesis, huge efforts are being spent to identify drugs able to interfere with proteases regulating Aβ formation from amyloid precursor protein (APP). This article briefly reviews the profile of γ‐secretase inhibitors, compounds that inhibit γ‐secretase, the pivotal enzyme that generates Aβ, and that have reached the clinic. Discussion: Several classes of potent γ‐secretase inhibitors have been designed and synthesized. Preclinical studies have indicated that these compounds are able to lower brain Aβ concentrations and, in some cases, reduce Aβ plaque deposition in transgenic mouse models of AD. The most developmentally advanced of these compounds is semagacestat, presently in Phase III clinical trials. In animals, semagacestat reduced Aβ levels in the plasma, cerebrospinal fluid (CSF), and the brain. However, studies have not reported on its cognitive effects. Studies in both healthy volunteers and patients with AD have demonstrated a dose‐dependent inhibition of plasma Aβ levels, and a recent study in healthy subjects demonstrated a robust, dose‐dependent inhibition of newly generated Aβ in the CSF after single oral doses. Conclusions: Unfortunately, γ‐secretase inhibitors may cause intestinal goblet cell hyperplasia, thymus atrophy, decrease in lymphocytes, and alterations in hair color, effects associated with the inhibition of the cleavage of Notch, a protein involved in cell development and differentiation. Nevertheless, at least other two promising γ‐secretase inhibitors are being tested clinically. This class of drugs represents a major hope to slow the rate of decline of AD.     

Conversion of Aβ43 to Aβ40 by the successive action of angiotensin‐converting enzyme 2 and angiotensin‐converting enzyme

The longer and neurotoxic species of amyloid‐β protein (Aβ), Aβ42 and Aβ43, contribute to Aβ accumulation in Alzheimer's disease (AD) pathogenesis and are considered to be the primary cause of the disease. In contrast, the predominant secreted form of Aβ, Aβ40, inhibits amyloid deposition and may have neuroprotective effects. We have reported that angiotensin‐converting enzyme (ACE) converts Aβ42 to Aβ40 and that Aβ43 is the earliest‐depositing Aβ species in the amyloid precursor protein transgenic mouse brain. Here we found that Aβ43 can be converted to Aβ42 and to Aβ40 in mouse brain lysate. We further identified the brain Aβ43‐to‐Aβ42‐converting enzyme as ACE2. The purified human ACE2 converted Aβ43 to Aβ42, and this activity was inhibited by a specific ACE2 inhibitor, DX600. Notably, the combination of ACE2 and ACE could convert Aβ43 to Aβ40. Our results indicate that the longer, neurotoxic forms of Aβ can be converted to the shorter, less toxic or neuroprotective forms of Aβ by ACE2 and ACE. Moreover, we found that ACE2 activity showed a tendency to decrease in the serum of AD patients compared with normal controls, suggesting an association between lower ACE2 activity and AD. Thus, maintaining brain ACE2 and ACE activities may be important for preventing brain amyloid neurotoxicity and deposition in Alzheimer's disease. © 2014 Wiley Periodicals, Inc.     

Binding and uptake of Aβ1‐42 by primary human astrocytes in vitro

Clearance of the amyloid‐β peptide (Aβ) as a remedy for Alzheimer's disease (AD) is a major target in on‐going clinical trials. In vitro studies confirmed that Aβ is taken up by rodent astrocytes, but knowledge on human astrocyte‐mediated Aβ clearance is sparse. Therefore, by means of flow cytometry and confocal laser scanning microscopy (CLSM), we evaluated the binding and internalization of Aβ1‐42 by primary human fetal astrocytes and adult astrocytes, isolated from nondemented subjects (n = 8) and AD subjects (n = 6). Furthermore, we analyzed whether α1‐antichymotrypsin (ACT), which is found in amyloid plaques and can influence Aβ fibrillogenesis, affects the Aβ uptake by human astrocytes. Upon over night exposure of astrocytes to FAM‐labeled Aβ1‐42 (10 μM) preparations, (80.7 ± 17.7)% fetal and (52.9 ± 20.9)% adult Aβ‐positive astrocytes (P = 0.018) were observed. No significant difference was found in Aβ1‐42 uptake between AD and non‐AD astrocytes, and no influence of ApoE genotype on Aβ1‐42 uptake was observed in any group. There was no difference in the percentage of Aβ‐positive cells upon exposure to Aβ1‐42 (10 μM) combined with ACT (1,000:1, 100:1, and 10:1 molar ratio), versus Aβ1‐42 alone. CLSM revealed binding of Aβ1‐42 to the cellular surfaces and cellular internalization of smaller Aβ1‐42 fragments. Under these conditions, there was no increase in cellular release of the proinflammatory chemokine monocyte‐chemoattractant protein 1, as compared with nontreated control astrocytes. Thus, primary human astrocytes derived from different sources can bind and internalize Aβ1‐42, and fetal astrocytes were more efficient in Aβ1‐42 uptake than adult astrocytes. © 2008 Wiley‐Liss, Inc.     

Caspases‐2 and ‐8 are involved in the presenilin1/γ‐secretase‐dependent cleavage of amyloid precursor protein after the induction of apoptosis

The presenilin/γ‐secretase protease cleaves many type‐I membrane proteins, including the amyloid β‐protein (Aβ) precursor (APP). Previous studies have shown that apoptosis induces alterations in Aβ production in a caspase‐dependent manner. Here, we report that staurosporine (STS)‐induced apoptosis induces caspase‐8 and/or‐2‐dependent γ‐secretase activation. Blocking of caspase activity with caspase‐8 inhibitor z‐IETD‐fmk, and caspase‐2 inhibitor z‐VDVAD‐fmk reduced Aβ production by STS in H4 cells expressing the Swedish mutant of APP (HSW) or APP‐C99 (H4‐C99). There was no inhibitory effect of other caspases (‐1, ‐3, ‐5, ‐6, ‐9) on Aβ production by STS. This finding was further supported by evidence that siRNA transfection, depleting caspase‐2 or ‐8 levels, lowered Aβ production in HSW and H4‐C99 cells without affecting expression of APP or γ‐secretase complex. In addition, Aβ production by STS was decreased by JNK inhibitors, SP600125. These results suggest that caspase‐2 and/or ‐8 is involved in presenilin/γ‐secretase activation and Aβ production in apoptosis. © 2010 Wiley‐Liss, Inc.     

Amyloid‐β induces sleep fragmentation that is rescued by fatty acid binding proteins in Drosophila

Disruption of sleep/wake activity in Alzheimer's disease (AD) patients significantly affects their quality of life and that of their caretakers and is a major contributing factor for institutionalization. Levels of amyloid‐β (Aβ) have been shown to be regulated by neuronal activity and to correlate with the sleep/wake cycle. Whether consolidated sleep can be disrupted by Aβ alone is not well understood. We hypothesize that Aβ42 can increase wakefulness and disrupt consolidated sleep. Here we report that flies expressing the human Aβ42 transgene in neurons have significantly reduced consolidated sleep compared with control flies. Fatty acid binding proteins (Fabp) are small hydrophobic ligand carriers that have been clinically implicated in AD. Aβ42 flies that carry a transgene of either the Drosophila Fabp or the mammalian brain‐type Fabp show a significant increase in nighttime sleep and long consolidated sleep bouts, rescuing the Aβ42‐induced sleep disruption. These studies suggest that alterations in Fabp levels and/or activity may be associated with sleep disturbances in AD. Future work to determine the molecular mechanisms that contribute to Fabp‐mediated rescue of Aβ42‐induced sleep loss will be important for the development of therapeutics in the treatment of AD. © 2016 Wiley Periodicals, Inc.     

Characterization of plasma β‐secretase (BACE1) activity and soluble amyloid precursor proteins as potential biomarkers for Alzheimer's disease

Reduction in cerebrospinal fluid (CSF) amyloid β42 (Aβ42) and elevation in total tau and phospho‐thr181 tau consistently differentiate between Alzheimer's disease (AD) and age‐matched control subjects. In contrast, CSF β‐site APP‐cleaving enzyme activity (BACE1) and soluble amyloid precursor proteins α and β (sAPPα and sAPPβ) are without consistent patterns in AD subjects. Plasma sampling is much easier, with fewer side effects, and is readily applied in primary care centers, so we have developed and validated novel plasma BACE activity, sAPPβ, and sAPPα assays and investigated their ability to distinguish AD from age‐matched controls. Plasma BACE activity assay was sensitive and specific, with signal being immunodepleted with a specific BACE1 antibody and inhibited with a BACE1‐specific inhibitor. Plasma sAPPβ and sAPPα assays were specific, with signal diluting linearly, immunodepleted with specific antibodies, and at background levels in APP knockout mice. In rhesus monkeys, BACE1 but not γ‐secretase inhibitor led to significant lowering of plasma sAPPβ with concurrent elevation of plasma sAPPα. AD subjects showed a significant increase in plasma BACE1 activity, sAPPβ, sAPPα, and Aβ42 (P < 0.001) compared with age‐matched controls. In conclusion, plasma BACE activity and sAPP endpoints provide novel investigative biomarkers for AD diagnosis and potential pharmacodynamic biomarkers for secretase inhibitor studies. © 2012 Wiley Periodicals, Inc.     

The production ratios of AICDε51 and Aβ42 by intramembrane proteolysis of βAPP do not always change in parallel

Background: During intramembrane proteolysis of β‐amyloid protein precursor (βAPP) by presenilin (PS)/γ‐secretase, ε‐cleavages at the membrane‐cytoplasmic border precede γ‐cleavages at the middle of the transmembrane domain. Generation ratios of Aβ42, a critical molecule for Alzheimer's disease (AD) pathogenesis, and the major Aβ40 species might be associated with ε48 and ε49 cleavages, respectively. Medicines to downregulate Aβ42 production have been investigated by many pharmaceutical companies. Therefore, the ε‐cleavages, rather than the γ‐cleavage, might be more effective upstream targets for decreasing the relative generation of Aβ42. Thus, one might evaluate compounds by analyzing the generation ratio of the βAPP intracellular domain (AICD) species (ε‐cleavage‐derived), instead of that of Aβ42. Methods: Cell‐free γ‐secretase assays were carried out to observe de novo AICD production. Immunoprecipitation/MALDI‐TOF MS analysis was carried out to detect the N‐termini of AICD species. Aβ and AICD species were measured by ELISA and immunoblotting techniques. Results: Effects on the ε‐cleavage by AD‐associated pathological mutations around the ε‐cleavage sites (i.e., βAPP V642I, L648P and K649N) were analyzed. The V642I and L648P mutations caused an increase in the relative ratio of ε48 cleavage, as expected from previous reports. Cells expressing the K649N mutant, however, underwent a major ε‐cleavage at the ε51 site. These results suggest that ε51, as well as ε48 cleavage, is associated with Aβ42 production. Only AICDε51, though, and not Aβ42 production, dramatically changed with modifications to the cell‐free assay conditions. Interestingly, the increase in the relative ratio of the ε51 cleavage by the K649N mutation was not cancelled by these changes. Conclusion: Our current data show that the generation ratio of AICDε51 and Aβ42 do not always change in parallel. Thus, to identify compounds that decrease the relative ratio of Aβ42 generation, measurement of the relative level of Aβ42‐related AICD species (i.e., AICDε48 and AICDε51) might not be useful. Further studies to reveal how the ε‐cleavage precision is decided are necessary before it will be possible to develop drugs targeting ε‐cleavage as a means for decreasing Aβ42 production.     

Attenuation of β‐amyloid‐induced tauopathy via activation of CK2α/SIRT1: Targeting for cilostazol

β‐Amyloid (Aβ) deposits and hyperphosphorylated tau aggregates are the chief hallmarks in the Alzheimer's disease (AD) brains, but the strategies for controlling these pathological events remain elusive. We hypothesized that CK2‐coupled SIRT1 activation stimulated by cilostazol suppresses tau acetylation (Ac‐tau) and tau phosphorylation (P‐tau) by inhibiting activation of P300 and GSK3β. Aβ was endogenously overproduced in N2a cells expressing human APP Swedish mutation (N2aSwe) by exposure to medium containing 1% fetal bovine serum for 24 hr. Increased Aβ accumulation was accompanied by increased Ac‐tau and P‐tau levels. Concomitantly, these cells showed increased P300 and GSK3β P‐Tyr216 expression; their expressions were significantly reduced by treatment with cilostazol (3–30 μM) and resveratrol (20 μM). Moreover, decreased expression of SIRT1 and its activity by Aβ were significantly reversed by cilostazol as by resveratrol. In addition, cilostazol strongly stimulated CK2α phosphorylation and its activity, and then stimulated SIRT1 phosphorylation. These effects were confirmed by using the pharmacological inhibitors KT5720 (1 μM, PKA inhibitor), TBCA (20 μM, inhibitor of CK2), and sirtinol (20 μM, SIRT1 inhibitor) as well as by SIRT1 gene silencing and overexpression techniques. In conclusion, increased cAMP‐dependent protein kinase‐linked CK2/SIRT1 expression by cilostazol can be a therapeutic strategy to suppress the tau‐related neurodegeneration in the AD brain. © 2013 Wiley Periodicals, Inc.     

Genetic reductions of β‐site amyloid precursor protein‐cleaving enzyme 1 and amyloid‐β ameliorate impairment of conditioned taste aversion memory in 5XFAD Alzheimer’s disease model mice

Although transgenic mouse models of Alzheimer’s disease (AD) recapitulate amyloid‐β (Aβ)‐related pathologies and cognitive impairments, previous studies have mainly evaluated their hippocampus‐dependent memory dysfunctions using behavioral tasks such as the water maze and fear conditioning. However, multiple memory systems become impaired in AD as the disease progresses and it is important to test whether other forms of memory are affected in AD models. This study was designed to use conditioned taste aversion (CTA) and contextual fear conditioning paradigms to compare the phenotypes of hippocampus‐independent and ‐dependent memory functions, respectively, in 5XFAD amyloid precursor protein/presenilin‐1 transgenic mice that harbor five familial AD mutations. Although both types of memory were significantly impaired in 5XFAD mice, the onset of CTA memory deficits (～9 months of age) was delayed compared with that of contextual memory deficits (～6 months of age). Furthermore, 5XFAD mice that were genetically engineered to have reduced levels of β‐site amyloid precursor protein‐cleaving enzyme 1 (BACE1) (BACE1^+/?·5XFAD) exhibited improved CTA memory, which was equivalent to the performance of wild‐type controls. Importantly, elevated levels of cerebral β‐secretase‐cleaved C‐terminal fragment (C99) and Aβ peptides in 5XFAD mice were significantly reduced in BACE1^+/?·5XFAD mice. Furthermore, Aβ deposition in the insular cortex and basolateral amygdala, two brain regions that are critically involved in CTA performance, was also reduced in BACE1^+/?·5XFAD compared with 5XFAD mice. Our findings indicate that the CTA paradigm is useful for evaluating a hippocampus‐independent form of memory defect in AD model mice, which is sensitive to rescue by partial reductions of the β‐secretase BACE1 and consequently of cerebral Aβ.     

Fustin flavonoid attenuates β‐amyloid (1–42)‐induced learning impairment

Natural flavonoids ameliorate amyloid‐β peptide (Aβ)‐induced neurotoxicity. We examined whether the fustin flavonoid affects Aβ‐induced learning impairment in mice. Repeated treatment with fustin significantly attenuated Aβ (1–42)‐induced conditioned fear and passive avoidance behaviors. This effect was comparable to that of EGb761, a standard extract of ginkgo. Fustin treatment significantly prevented decreases in acetylcholine (ACh) levels, choline acetyltransferase (ChAT) activity, and ChAT gene expression induced by Aβ (1–42). Fustin also consistently suppressed increases in acetyl cholinesterase (AChE) activity and AChE gene expression induced by Aβ (1–42). In addition, fustin significantly attenuated Aβ (1–42)‐induced selective decreases in muscarinic M1 receptor gene expression and muscarinic M1 receptor binding activity (as determined by [³H]pirenzepine binding) by modulating extracellular signal‐regulated kinase 1/2 (ERK 1/2) and cAMP response‐element binding protein (CREB) phosphorylation and brain‐derived neurotrophic factor (BDNF) expression. These effects of fustin were reversed by treatment with dicyclomine, a muscarinic M1 receptor antagonist, and SL327, a selective ERK inhibitor, but not by chelerythrine, a pan‐protein kinase C (PKC) inhibitor. Taken together, our results suggest that fustin attenuates Aβ (1–42)‐impaired learning, and that the ERK/CREB/BDNF pathway is important for the M1 receptor‐mediated cognition‐enhancing effects of fustin. © 2009 Wiley‐Liss, Inc.     

Altered levels and distribution of amyloid precursor protein and its processing enzymes in Niemann‐Pick type C1‐deficient mouse brains

Niemann‐Pick type C (NPC) disease is an autosomal recessive neurodegenerative disorder characterized by intracellular accumulation of cholesterol and glycosphingolipids in many tissues including the brain. The disease is caused by mutations of either NPC1 or NPC2 gene and is accompanied by a severe loss of neurons in the cerebellum, but not in the hippocampus. NPC pathology exhibits some similarities with Alzheimer's disease, including increased levels of amyloid β (Aβ)‐related peptides in vulnerable brain regions, but very little is known about the expression of amyloid precursor protein (APP) or APP secretases in NPC disease. In this article, we evaluated age‐related alterations in the level/distribution of APP and its processing enzymes, β‐ and γ‐secretases, in the hippocampus and cerebellum of Npc1^−/− mice, a well‐established model of NPC pathology. Our results show that levels and expression of APP and β‐secretase are elevated in the cerebellum prior to changes in the hippocampus, whereas γ‐secretase components are enhanced in both brain regions at the same time in Npc1^−/− mice. Interestingly, a subset of reactive astrocytes in Npc1^−/− mouse brains expresses high levels of APP as well as β‐ and γ‐secretase components. Additionally, the activity of β‐secretase is enhanced in both the hippocampus and cerebellum of Npc1^−/− mice at all ages, while the level of C‐terminal APP fragments is increased in the cerebellum of 10‐week‐old Npc1^−/− mice. These results, taken together, suggest that increased level and processing of APP may be associated with the development of pathology and/or degenerative events observed in Npc1^−/− mouse brains. © 2010 Wiley‐Liss, Inc.     

Glutamate protects neuromuscular junctions from deleterious effects of β‐amyloid peptide and conversely: An in vitro study in a nerve–muscle coculture

Murine models of Alzheimer's disease with elevated levels of amyloid‐β (Aβ) peptide present motor axon defects and neuronal death. Aβ_1–42 accumulation is observed in motor neurons and spinal cords of sporadic and familial cases of amyotrophic lateral sclerosis (ALS). Motor neurons are highly susceptible to glutamate, which has a role in ALS neuronal degeneration. The current study investigates the link between Aβ and glutamate in this neurodegenerative process. Primary rat nerve and human muscle cocultures were intoxicated with glutamate or Aβ. Neuromuscular junction (NMJ) mean size and neurite length were evaluated. The role of N‐methyl‐D‐aspartate receptor (NMDAR) was investigated by using MK801. Glutamate and Aβ production were evaluated in culture supernatant. The current study shows that NMJs are highly sensitive to Aβ peptide, that the toxic pathway involves glutamate and NMDAR, and that glutamate and Aβ act in an interlinked manner. Some motor diseases (e.g., ALS), therefore, could be considered from a new point of view related to these balance disturbances. © 2014 Wiley Periodicals, Inc.     

Astrocytic Aβ1‐42 uptake is determined by Aβ‐aggregation state and the presence of amyloid‐associated proteins

Intracerebral accumulation of amyloid‐β (Aβ) leading to Aβ plaque formation, is the main hallmark of Alzheimer's disease and might be caused by defective Aβ‐clearance. We previously found primary human astrocytes and microglia able to bind and ingest Aβ1‐42 in vitro, which appeared to be limited by Aβ1‐42 fibril formation. We now confirm that astrocytic Aβ‐uptake depends on size and/or composition of Aβ‐aggregates as astrocytes preferably take up oligomeric Aβ over fibrillar Aβ. Upon exposure to either fluorescence‐labelled Aβ1‐42 oligomers (Aβ_oligo) or fibrils (Aβ_fib), a larger (3.7 times more) proportion of astrocytes ingested oligomers compared to fibrils, as determined by flow cytometry. Aβ‐internalization was verified using confocal microscopy and live‐cell imaging. Neither uptake of Aβ_oligo nor Aβ_fib, triggered proinflammatory activation of the astrocytes, as judged by quantification of interleukin‐6 and monocyte‐chemoattractant protein‐1 release. Amyloid‐associated proteins, including α1‐antichymotrypsin (ACT), serum amyloid P component (SAP), C1q and apolipoproteins E (ApoE) and J (ApoJ) were earlier found to influence Aβ‐aggregation. Here, astrocytic uptake of Aβ_fib increased when added to the cells in combination with SAP and C1q (SAP/C1q), but was unchanged in the presence of ApoE, ApoJ and ACT. Interestingly, ApoJ and ApoE dramatically reduced the number of Aβ_oligo‐positive astrocytes, whereas SAP/C1q slightly reduced Aβ_oligo uptake. Thus, amyloid‐associated proteins, especially ApoJ and ApoE, can alter Aβ‐uptake in vitro and hence may influence Aβ clearance and plaque formation in vivo. © 2010 Wiley‐Liss, Inc.     

Hereditary cerebral hemorrhage with amyloidosis‐Dutch type

The amyloid β‐protein (Aβ) E22Q mutation of the rare disorder hereditary cerebral hemorrhage with amyloidosis‐Dutch type (HCHWA‐D) causes severe cerebral amyloid angiopathy (CAA) with hemorrhagic strokes of mid‐life onset and dementia. The mutation does not affect total Aβ production but may alter the Aβ1–42:Aβ1–40 ratio, and affect the proteolytic degradation of Aβ and its transport across the blood–brain barrier. Aβ E22Q aggregates faster into more stable amyloid‐like fibrils than wild‐type Aβ. Non‐fibrillar Aβ(x‐42) deposits precede the appearance of fibrils and the deposition of Aβ(x‐40) in the vascular basement membrane. CAA severity tends to increase with age but may vary greatly among patients of comparable ages. Lumenal narrowing of affected blood vessels, leukoencephalopathy, CAA‐associated vasculopathies, and perivascular astrocytosis, microgliosis, and neuritic degeneration complicate the development of HCHWA‐D CAA. Parenchymal Aβ deposition is also enhanced in the HCHWA‐D brain with non‐fibrillar membrane‐bound Aβ(x‐42) deposits evolving into relatively fibrillar diffuse plaques variously associated with reactive astrocytes, activated microglia, and degenerating neurites. Plaque density tends to decrease with age. Neurofibrillary degeneration is absent or limited. HCHWA‐D dementia is associated with CAA severity independently of Braak stage, age, and plaque density. Particularly, microaneurysms may contribute to the development of (small) hemorrhages/infarcts and the latter to cognitive decline in affected subjects. However, the relative importance of cerebral hemorrhages/infarcts, white matter damage and/or other CAA‐ or Aβ‐related factors for cognitive deterioration in HCHWA‐D remains to be determined.     

Regionally specific changes in the hippocampal circuitry accompany progression of cerebrospinal fluid biomarkers in preclinical Alzheimer's disease

Neuropathological and in vivo brain imaging studies agree that the cornu ammonis 1 and subiculum subfields of the hippocampus are most vulnerable to atrophy in the prodromal phases of Alzheimer's disease (AD). However, there has been limited investigation of the structural integrity of the components of the hippocampal circuit, including subfields and extra‐hippocampal white matter structure, in relation to the progression of well‐accepted cerebrospinal fluid (CSF) biomarkers of AD, amyloid‐β 1‐42 (Aβ) and total‐tau (tau). We investigated these relationships in 88 aging asymptomatic individuals with a parental or multiple‐sibling familial history of AD. Apolipoprotein (APOE) ?4 risk allele carriers were identified, and all participants underwent cognitive testing, structural magnetic resonance imaging, and lumbar puncture for CSF assays of tau, phosphorylated‐tau (p‐tau) and Aβ. Individuals with a reduction in CSF Aβ levels (an indicator of amyloid accretion into neuritic plaques) as well as evident tau pathology (believed to be linked to neurodegeneration) exhibited lower subiculum volume, lower fornix microstructural integrity, and a trend towards lower cognitive score than individuals who showed only reduction in CSF Aβ. In contrast, persons with normal levels of tau showed an increase in structural MR markers in relation to declining levels of CSF Aβ. These results suggest that hippocampal subfield volume and extra‐hippocampal white matter microstructure demonstrate a complex pattern where an initial volume increase is followed by decline among asymptomatic individuals who, in some instances, may be a decade or more away from onset of cognitive or functional impairment.     

Immunohistochemical characterization of γ-secretase activating protein expression in Alzheimer's disease brains

J. Satoh, H. Tabunoki, T. Ishida, Y. Saito and K. Arima (2012) Neuropathology and Applied Neurobiology 38, 132–141 Immunohistochemical characterization of γ‐secretase activating protein expression in Alzheimer's disease brains Aims: A recent study showed that γ‐secretase activating protein (GSAP), derived from a C‐terminal fragment of pigeon homolog (PION), increases amyloid‐β (Aβ) production by interacting with presenilin‐1 (PS1) and the β‐secretase‐cleaved C‐terminal fragment of amyloid precursor protein (APP‐CTF). In the study, knockdown of GSAP reduces production of Aβ and plaque formation in the brain of APPswe and PS1ΔE9 double transgenic mice without affecting the Notch‐dependent pathway. Therefore, GSAP is an ideal target for designing γ‐secretase modulators with least side effects in Alzheimer's disease (AD). However, at present, the precise distribution of GSAP in AD brains remains to be characterized. Methods: By immunohistochemistry, we studied GSAP expression in the frontal cortex and the hippocampus of 11 aged AD and 17 age‐matched control cases. Results: GSAP immunoreactivity exhibited distinct morphological features, such as fine granular cytoplasmic deposits, dense nodular and patchy deposits, beads and string‐like deposits, and diffuse dot‐like deposits. In both AD and control brains, a fairly small subset of cerebral cortical and hippocampal neurones expressed fine granular cytoplasmic deposits, while diffuse dot‐like deposits were more frequently found in the neuropil and neuronal processes, particularly enriched in the hippocampal CA2 and CA3 regions. Among GSAP‐immunoreactive deposits, dense nodular and patchy deposits, located in the neuropil and closely associated with PS1 expression and Aβ deposition, indicated the most distinguishing features of AD pathology. Conclusions: Aberrant regulation of GSAP expression plays a key role in acceleration of γ‐cleavage of APP‐CTF and accumulation of Aβ in AD brains.