Genetic alterations of the BRI2 gene: familial British and Danish dementias.

Classic arguments sustaining the importance of amyloid in the pathogenesis of dementia are usually centered on amyloid beta (Abeta) and its role in neuronal loss characteristic of Alzheimer disease, the most common form of human cerebral amyloidosis. Two non-Abeta cerebral amyloidoses, familial British and Danish dementias, share many aspects of Alzheimer disease, including the presence of neurofibrillary tangles, parenchymal pre-amyloid and amyloid deposits, cerebral amyloid angiopathy, and a widespread inflammatory response. Both early-onset conditions are linked to specific mutations in the BRI2 gene, causing the generation of longer-than-normal protein products and the release of 2 de novo created peptides ABri and ADan, the main components of amyloid fibrils in these inherited dementias. Although the molecular mechanisms and signal transduction pathways elicited by the amyloid deposits and their relation to cognitive impairment remain to be clarified, new evidence indicates that, independent of the differences in their primary structures, Abeta, ABri, and ADan subunits are able to form morphologically compatible ion-channel-like structures and elicit single ion-channel currents in reconstituted lipid membranes. These findings reaffirm the notion that non-Abeta amyloidosis constitute suitable alternative models to study the role of amyloid deposition in the mechanism of neuronal cell death.

Chromosome 13 dementias.

The importance of cerebral amyloid deposition in the mechanism of neurodegeneration is still debatable. Classic arguments are usually centered on amyloid beta(Abeta) and its role in the neuronal loss characteristic of Alzheimer's disease, the most common form of human cerebral amyloidosis. Two non-Abeta cerebral amyloidoses, familial British and Danish dementias (FBD and FDD), share many aspects of Alzheimer's disease, including the presence of neurofibrillary tangles, parenchymal preamyloid and amyloid deposits, cerebral amyloid angiopathy and a variety of amyloid-associated proteins and inflammatory components. Both early-onset conditions are linked to specific mutations at or near the stop codon of the chromosome 13 gene BRI2 that cause generation of longer-than-normal protein products. Furin-like processing of these longer precursors releases two de novo-created peptides, ABri and ADan, which deposit as amyloid fibrils in FBD and FDD, respectively. Due to the similar pathology generated by completely unrelated amyloid subunits, FBD and FDD, collectively referred to as chromosome 13 dementias, constitute alternative models for studying the role of amyloid deposition in the mechanism of neuronal cell death.

Brain Abeta amyloidosis in APPsw mice induces accumulation of presenilin-1 and tau.

APPsw transgenic mice (Tg2576) overproducing mutant amyloid beta protein precursor (betaAPP) show substantial brain Abeta amyloidosis and behavioural abnormalities. To clarify the subsequent abnormalities, the disappearance of neurons and synapses and dystrophic neurite formation with accumulated proteins including hyperphosphorylated tau were examined. Tg2576 demonstrated substantial giant core plaques and diffuse plaques. The number of neurons was significantly decreased in the areas containing the amyloid cores compared with all other areas and corresponding areas in non-transgenic littermates in sections visualized by Nissl plus Congo red double staining (p<0.001). The presynaptic protein alpha-synuclein and postsynaptic protein drebrin were also absent in the amyloid cores. betaAPP and presenilin-1 were accumulated in dystrophic neurites in and around the core plaques. Tau phosphorylated at five independent sites was detected in the dystrophic neurites in the amyloid cores. Thus, the giant core plaques replaced normal brain tissues and were associated with subsequent pathological features such as dystrophic neurites and the appearance of hyperphosphorylated tau. These findings suggest a potential role for brain Abeta amyloidosis in the induction of secondary pathological steps leading to mental disturbance in Alzheimer's disease.

The levels of cerebrospinal fluid Abeta40 and Abeta42(43) are regulated age-dependently.

Decreased levels of cerebrospinal fluid (CSF) Abeta42 is a diagnostic marker of Alzheimer's disease. To clarify the biological basis of this marker, the physiological alterations of CSF Abeta40 and Abeta42 by aging were studied. CSF samples from 92 normal subjects between 8 and 89 years old were measured using a specific ELISA for Abeta40 and Abeta42(43). High concentrations of Abeta40 and Abeta42(43) in the young group, under 29 years old, changed to be at low concentrations in the adult group between 30 and 59 years old. Subsequently, the levels increased again with age. Third order regression analysis showed a significant correlation between the levels of Abeta40 and age (Y = - 169 X(3) + 3.1X(2)- 0.02X + 4135; P < 0.034) and between the levels of Abeta42(43) and age (Y = - 46 X(3) + 0.9 X(2)- 0.005X + 992; P < 0.005). The levels of CSF Abeta40 and Abeta42(43) were physiologically regulated to show a U-shaped natural course in normal aging. These findings suggested that the physiological increase of Abeta42(43) over 59 years of age is selectively inhibited in Alzheimer's disease.

Abeta amyloidosis induces the initial stage of tau accumulation in APP(Sw) mice.

To clarify how Abeta deposits induce secondary tauopathy, the presence of phosphorylated tau, glycogen synthase kinase 3alpha (GSK3alpha), GSK3beta, cyclin-dependent kinase 5 (CDK5), mitogen-activated protein kinase (MAPK) and fyn were examined in the Tg2576 brain showing substantial brain Abeta amyloidosis and behavioral abnormalities. Phosphorylated tau at Ser199, Thr231/Ser235, Ser396 and Ser413 accumulated in the dystrophic neurites of senile plaques. The major kinase for tau phosphorylation was GSK3beta. Smaller contributions of GSK3alpha, CDK5 and MAPK were suggested. Thus, brain Abeta amyloidosis has a potential role in the induction of tauopathy leading to the mental disturbances of Alzheimer's disease.

Impaired neurotransmitter systems by Abeta amyloidosis in APPsw transgenic mice overexpressing amyloid beta protein precursor.

APPsw transgenic mice showing substantial features of brain Abeta amyloidosis such as senile plaques and behavioral abnormalities were examined by immunostaining to determine whether Abeta deposits could induce the subsequent disturbance of neurotransmitter systems including somatostatin, substance P and choline acetyltransferase (ChAT), which are prominent in the Alzheimer's disease brain. Somatostatin, substance P and ChAT disappeared in the areas of senile plaque and were accumulated in dystrophic neurites around the amyloid cores. These findings suggest a potential role of brain Abeta amyloidosis in disturbance of the neurotransmitter systems leading to memory disturbance of Alzheimer's disease.

Accumulation of NACP/alpha-synuclein in lewy body disease and multiple system atrophy.

OBJECTIVES: NACP/alpha-synuclein is an aetiological gene product in familial Parkinson's disease. To clarify the pathological role of NACP/alpha-synuclein in sporadic Parkinson's disease and other related disorders including diffuse Lewy body disease (DLBD) and multiple system atrophy (MSA), paraffin sections were examined immunocytochemically using anti-NACP/alpha-synuclein antibodies. METHODS: A total of 58 necropsied brains, from seven patients with Parkinson's disease, five with DLBD, six with MSA, 12 with Alzheimer's disease, one with Down's syndrome, one with amyotrophic lateral sclerosis (ALS), three with ALS and dementia, one with Huntington's disease, two with progressive supranuclear palsy (PSP), one with Pick's disease, one with myotonic dystrophy, and three with late cerebellar cortical atrophy (LCCA), and 15 elderly normal controls were examined. RESULTS: In addition to immunoreactive Lewy bodies, widespread accumulation of NACP/alpha-synuclein was found in neurons and astrocytes from the brainstem and basal ganglia to the cerebral cortices in Parkinson's disease/DLBD. NACP/alpha-synuclein accumulates in oligodendrocytes from the spinal cord, the brain stem to the cerebellar white matter, and inferior olivary neurons in MSA. These widespread accumulations were not seen in other types of dementia or spinocerebellar ataxia. CONCLUSION: Completely different types of NACP/alpha-synuclein accumulation in Parkinson's disease/DLBD and MSA suggest that accumulation is a major step in the pathological cascade of both diseases and provides novel strategies for the development of therapies.

Age-related amyloid beta protein accumulation induces cellular death and macrophage activation in transgenic mice.

In view of the importance of amyloid beta protein accumulation in Alzheimer's disease, this paper examines age-related amyloid beta protein (Abeta) deposition and accompanying cellular changes in a mouse model in vivo. Transgenic mice were studied which expressed a gene encoding 18 residues of signal peptide and 99 residues of the carboxyl-terminal fragment (CTF) of the Abeta precursor, under the control of the cytomegalovirus enhancer/chicken beta-actin promoter. In the pancreas, Abeta accumulated in an age-dependent manner. Abeta deposits appeared as early as 3 weeks of age and increased in size and number from 4 to 16 months of age. The largest Abeta deposits were observed in the transgenic pancreas at 16 and 20 months of age. Haematoxylin and eosin staining, macrophage immunostaining, and electron microscopy showed that the Abeta fibril deposits closely correlated with degeneration of pancreatic acinar cells and macrophage activation. Abeta1-42 and Abetap3E-42 were predominant components of Abeta deposits among amino- and carboxyl-terminal modified Abeta species. These findings suggest that overproduction of Abeta causes age-related accumulation of Abeta fibrils, with accompanying cellular degeneration and macrophage activation in vivo.

Lipoprotein-free amyloidogenic peptides in plasma are elevated in patients with sporadic Alzheimer's disease and Down's syndrome.

About 90% of the soluble amyloid beta (sA beta) that circulates in normal human plasma is associated with lipoprotein particles. In sporadic Alzheimer's disease patients, free sA beta42 but not sA beta40 is increased approximately 2.3-fold compared with age-matched controls, although a more marked elevation (approximately 8-fold for free sA beta40 and about 20-fold for sA beta42) is found in Down's syndrome patients. The data suggest that lipoprotein-sA beta dissociation may contribute to the influx of sA beta into the brain as a result of decreased plasma clearance.

Carboxyl-terminal fragments of presenilin-1 are closely related to cytoskeletal abnormalities in Alzheimer's brains.

To clarify the role of presenilin-1 (PS-1) in the pathology of Alzheimer's disease (AD), we tested four antisera to PS-1. The specific antisera to the N-terminus (HSN-2) and C-terminus (HS-C) of PS-1 detected a 44/40kD holoprotein, a 25kD N-terminal fragment (NTF) and a 16kD C-terminal fragment (CTF) of PS-1 in COS-7 cells. The 25kD NTF and 16kD CTF were observed in human brains, and their amounts were not significantly different between the control and AD brains. The antibody HS-C labeled extensive neurofibrillary tangles, dystrophic neurites and curly fibers in the AD brains. In the paired helical filament (PHF) fraction containing A68 protein from AD brains, a smear pattern of CTFs was revealed. Antisera (HS-L292 and HS-L300) to cleavage sites of PS-1 also revealed immunoreactive neurofibrillary tangles in the AD brain sections and the smear pattern of CTFs of A68 protein fraction. The CTFs of PS-1 accumulate with PHF tau, suggesting a close relationship between PS-1 and cytoskeletal abnormalities in AD brains.

Combination assay of CSF tau, A beta 1-40 and A beta 1-42(43) as a biochemical marker of Alzheimer's disease.

Cerebrospinal fluid samples from a total of 157 subjects consisting of 55 patients with sporadic Alzheimer's disease (AD), 34 normal controls, 23 patients with non-AD dementia, and 45 with other neurological diseases were examined by ELISA of tau, A beta 1-40, and A beta 1-42(43). The AD group had a significantly higher level of tau than the normal control group (P < 0.001), and the diagnostic sensitivity was 31% and specificity was 94%. CSF A beta 1-40 levels did not show any significant differences. Although the level of A beta 1-42(43) was decreased significantly in the AD group compared to the control group (P < 0.005), the overlap of A beta 1-42(43) levels among all groups meant that none of the AD samples exceeded the cut-off value, the mean 2SD of normal control subjects. Reduction of A beta 1-42(43) levels in AD resulted in a significant increase in the ratio of A beta 1-40 to A beta 1-42(43) (A beta ratio) as an improved marker. The diagnostic sensitivity and specificity of A beta ratio were 51% and 82% respectively. The three indexes, using the tau level and A beta ratio (tau or A beta ratio, deviation score and tau x A beta ratio), showed better sensitivity (58%, 67%, 69%) and specificity (82%, 86%, 88%) than previously reported methods. Combination assay for CSF tau, A beta 1-40 and A beta 1-42(43) in CSF is a biological marker of AD and may be useful to biochemically monitor subjects under treatment.

Familial spinocerebellar ataxia with cerebellar atrophy, peripheral neuropathy, and elevated level of serum creatine kinase, gamma-globulin, and alpha-fetoprotein.

Here, we report a familial spinocerebellar ataxia (FSCA), which has clinical features similar to Friedreich's ataxia, an ataxia with isolated vitamin E deficiency, and ataxia telangiectasia. However, the serum levels of creatine kinase, gamma-globulin, and alpha-fetoprotein were elevated, and biochemical and genetic analyses ruled out diagnosis of these three ataxias as well as other FSCAs. Thus, this family is thought to have a new type of FSCA.

Longitudinal study of cerebrospinal fluid levels of tau, A beta1-40, and A beta1-42(43) in Alzheimer's disease: a study in Japan.

To clarify the alterations of tau, amyloid beta protein (A beta) 1-40 and A beta1-42(43) in the cerebrospinal fluid (CSF) that accompany normal aging and the progression of Alzheimer's disease (AD), CSF samples of 93 AD patients, 32 longitudinal subjects among these 93 AD patients, 33 patients with non-AD dementia, 56 with other neurological diseases, and 54 normal control subjects from three independent institutes were analyzed by sensitive enzyme-linked immunosorbent assays. Although the tau levels increased with aging, a significant elevation of tau and a correlation between the tau levels and the clinical progression were observed in the AD patients. A significant decrease of the A beta1-42(43) levels and a significant increase of the ratio of A beta1-40 to A beta1-42(43) were observed in the AD patients. The longitudinal AD study showed continuous low A beta1-42(43) levels and an increase of the ratio of A beta1-40 to A beta1-42(43) before the onset of AD. These findings suggest that CSF tau may increase with the clinical progression of dementia and that the alteration of the CSF level of A beta1-42(43) and the ratio of A beta1-40 to A beta1-42(43) may start at early stages in AD. The assays of CSF tau, A beta1-40, and A beta1-42(43) provided efficient diagnostic sensitivity (71%) and specificity (83%) by using the production of tau levels and the ratio of A beta1-40 to A beta1-42(43), and an improvement in sensitivity (to 91%) was obtained in the longitudinal evaluation.

Accumulation of amyloid beta protein in transgenic mice.

Carboxyl-terminal fragments of beta amyloid precursor protein (betaAPP) were expressed in mice under the transcriptional control of an ubiquitous promoter system, based upon a chicken beta-actin (betaA) promoter combined with cytomegalovirus (CMV) enhancer to obtain a systemic overproduction of amyloid beta protein (Abeta). Three transgene constructs were designed to encode signal peptide and carboxyl-terminal 99 amino acid residues to betaAPP (NOR-beta), methionine and C-terminal 103 amino acid residues of betaAPP (deltaNOR-beta), and methionine and C-terminal 103 amino acid residues with KM-NL substitution of betaAPP (deltaNL-beta). Although the transcriptional mRNA level and post-translational protein level from transgenes showed the same expression pattern, both the expression of Abeta and distribution of Abeta deposits were completely different among these strains. In NOR-beta mice, considerable amounts of Abeta were detected in plasma and Abeta deposits were observed in the pancreas. Brain Abeta deposits and small amounts of plasma Abeta were recognized in deltaNL-beta. These findings indicate that tissue specific processing and transgene constructs are major factors to determine the distribution of Abeta deposits.

Apolipoprotein E accumulates with the progression of A beta deposition in transgenic mice.

To study the role of apolipoprotein E (apoE) in vivo in deposits of amyloid beta protein (A beta), a major component of senile plaque amyloid in the brain of patients with Alzheimer disease, the transgenic mice were examined by apoE immunostaining. The mice were systemically overexpressing signal peptide and 99 amino acid residues of the carboxy-terminal fragment of human amyloid beta protein precursor (betaAPP) under control of the powerful cytomegalovirus enhancer/chicken beta-actin promotor. A beta deposits appeared at 4 months and increased with aging in the acinar cells of the transgenic pancreas. Similarly, apoE deposits appeared in the pancreatic acinar cells at 4 months old. The number and size of apoE deposits increased with aging and correlated with the progression of A beta deposits. Interstitial macrophages labeled by apoE immunostaining appeared at 8 months after birth and their number increased with aging. On serial section of the pancreata of 24-month-old mice, approximately 70% of A beta deposits were labeled with the apoE antiserum. ApoE was detected in the highly insoluble formic acid fraction of the transgenic pancreas by an immunoblot study. The Northern blot study revealed no increase in synthesis of endogenous apoE mRNA. These findings indicate that apoE is closely related to progression of A beta deposits with aging and suggest that A beta deposition in the transgenic pancreas is similar to that in the senile plaque of Alzheimer brains. Therefore, our experimental system using transgenic mice will provide a useful tool to analyze the molecular mechanism of A beta deposition in association with apoE in vivo.

Lysosomal generation of amyloid beta protein species in transgenic mice.

Soluble amyloid beta protein (A beta)1-40 and highly amyloidogenic A beta 1-42/43 were immunocytochemically labeled in lysosomes of acinar cells and macrophages in the pancreas of transgenic mice systemically expressing a C-terminal fragment of the A beta precursor. A beta 1-42/43 and long A beta species extending their C-termini were detected in the detergent-insoluble fraction. Immunoreactivity of cathepsin D was markedly increased in lysosomes filled with A beta fibrils. These findings indicated that A beta 1-40, A beta 1-42, A beta 1-43 and longer A beta species were generated in the lysosomes of the transgenic pancreas, and suggested that the activation of cathepsin D, a candidate gamma-secretase, leads to acceleration of A beta amyloid formation.

Intracellular generation of amyloid beta-protein from amyloid beta-protein precursor fragment by direct cleavage with beta- and gamma-secretase.

Two amyloid beta protein precursor (beta APP) fragments involving Met and 103 amino acids of C-terminus of beta APP (delta NOR-beta) and its KM-NL substitution (delta NL-beta) were expressed in COS-7 cells to clarify the proteolytic cleavages to generate amyloid beta protein (A beta). The 4.5-kD protein, A beta with additional N-terminal amino acids, and 4-kD A beta were directly produced and released from 12.5-kD expression proteins without any production of 11.4-kD C-terminal fragment starting at N-terminus of A beta and 3-kD "p3" A beta derivative. Intracellular 4-kD A beta was also detected. The substitution of KM-NL of beta APP found in Swedish familial Alzheimer's disease (AD) promoted the production of intracellular A beta and its release with no increase in level of 11.4-kD C-terminal fragment. These results suggested the presence of a distinct pathway in which A beta is directly cleaved at both N- and C-termini from beta APP fragment intracellularly to release A beta. Since KM-NL substitution enhanced intracellular A beta generation, this pathway may be associated with amyloidogenesis in AD.