Hiroyuki Honda

Altered Expression of Diabetes-Related Genes in Alzheimer's Disease Brains: The Hisayama Study

Diabetes mellitus (DM) is considered to be a risk factor for dementia including Alzheimers disease (AD). However, the molecular mechanism underlying this risk is not well understood. We examined gene expression profiles in postmortem human brains donated for the Hisayama study. Three-way analysis of variance of microarray data from frontal cortex, temporal cortex, and hippocampus was performed with the presence/absence of AD and vascular dementia, and sex, as factors. Comparative analyses of expression changes in the brains of AD patients and a mouse model of AD were also performed. Relevant changes in gene expression identified by microarray analysis were validated by quantitative real-time reverse-transcription polymerase chain reaction and western blotting. The hippocampi of AD brains showed the most significant alteration in gene expression profile. Genes involved in noninsulin-dependent DM and obesity were significantly altered in both AD brains and the AD mouse model, as were genes related to psychiatric disorders and AD. The alterations in the expression profiles of DM-related genes in AD brains were independent of peripheral DM-related abnormalities. These results indicate that altered expression of genes related to DM in AD brains is a result of AD pathology, which may thereby be exacerbated by peripheral insulin resistance or DM.

The changing prevalence and incidence of dementia over time — current evidence

Dementia is an increasing focus for policymakers, civil organizations and multidisciplinary researchers. The most recent descriptive epidemiological research into dementia is enabling investigation into how the prevalence and incidence are changing over time. To establish clear trends, such comparisons need to be founded on population-based studies that use similar diagnostic and research methods consistently over time. This narrative Review synthesizes the findings from 14 studies that investigated trends in dementia prevalence (nine studies) and incidence (five studies) from Sweden, Spain, the UK, the Netherlands, France, the USA, Japan and Nigeria. Besides the Japanese study, these studies indicate stable or declining prevalence and incidence of dementia, and some provide evidence of sex-specific changes. No single risk or protective factor has been identified that fully explains the observed trends, but major societal changes and improvements in living conditions, education and healthcare might have favourably influenced physical, mental and cognitive health throughout an individuals life course, and could be responsible for a reduced risk of dementia in later life. Analytical epidemiological approaches combined with translational neuroscientific research could provide a unique opportunity to explore the neuropathology that underlies changing occurrence of dementia in the general population.

An autopsied case of sporadic adult-onset amyotrophic lateral sclerosis with FUS-positive basophilic inclusions.

Basophilic inclusions (BIs), which are characterized by their staining properties of being weakly argyrophilic, reactive with Nissl staining, and immunohistochemically negative for tau and transactive response (TAR) DNA‐binding protein 43 (TDP‐43), have been identified in patients with juvenile‐onset amyotrophic lateral sclerosis (ALS) and adult‐onset atypical ALS with ophthalmoplegia, autonomic dysfunction, cerebellar ataxia, or a frontal lobe syndrome. Mutations in the fused in sarcoma gene (FUS) have been reported in cases of familial and sporadic ALS, and FUS immunoreactivity has been demonstrated in basophilic inclusion body disease (BIBD), neuronal intermediate filament inclusion disease (NIFID), and atypical frontotemporal lobar degeneration with ubiquitin‐positive and tau‐negative inclusions (aFTLD‐U). In the present study, we immunohistochemically and ultrastructurally studied an autopsy case of sporadic adult‐onset ALS with numerous BIs. The patient presented with the classical clinical course of ALS since 75 years of age and died at age 79. Postmortem examination revealed that both Betz cells in the motor cortex and motor neurons in the spinal cord were affected. The substantia nigra was spared. Notably, BIs were frequently observed in the motor neurons of the anterior horns, the inferior olivary nuclei, and the basal nuclei of Meynert. BIs were immunopositive for p62, LC3, and FUS, but immunonegative for tau, TDP‐43, and neurofilament. Ultrastructurally, BIs consisted of filamentous or granular structures associated with degenerated organelles with no limiting membrane. There were no Bunina bodies, skein‐like inclusions, or Lewy‐like inclusions. All exons and exon/intron boundaries of the FUS gene were sequenced but no mutations were identified.

Protease-resistant PrP and PrP oligomers in the brain in human prion diseases after intraventricular pentosan polysulfate infusion

Intraventricular infusion of pentosan polysulfate (PPS) as a treatment for various human prion diseases has been applied in Japan. To evaluate the influence of PPS treatment we performed pathological examination and biochemical analyses of PrP molecules in autopsied brains treated with PPS (one case of sporadic Creutzfeldt‐Jakob disease (sCJD, case 1), two cases of dura mater graft‐associated CJD (dCJD, cases 2 and 4), and one case of Gerstmann‐Sträussler‐Scheinker disease (GSS, case 3). Six cases of sCJD without PPS treatment were examined for comparison. Protease‐resistant PrP (PrPres) in the frontal lobe was evaluated by Western blotting after proteinase K digestion. Further, the degree of polymerization of PrP molecules was examined by the size‐exclusion gel chromatography assay. PPS infusions were started 3–10 months after disease onset, but the treatment did not achieve any clinical improvements. Postmortem examinations of the treated cases revealed symmetrical brain lesions, including neuronal loss, spongiform change and gliosis. Noteworthy was GFAP in the cortical astrocytes reduced in all treated cases despite astrogliosis. Immunohistochemistry for PrP revealed abnormal synaptic deposits in all treated cases and further plaque‐type PrP deposition in case 3 of GSS and case 4 of dCJD. Western blotting showed relatively low ratios of PrPres in case 2 of dCJD and case 3 of GSS, while in the treated sCJD (case 1), the ratio of PrPres was comparable with untreated cases. The indices of oligomeric PrP were reduced in one sCJD (case 1) and one dCJD (case 2). Although intraventricular PPS infusion might modify the accumulation of PrP oligomers in the brains of patients with prion diseases, the therapeutic effects are still uncertain.

Loss of hnRNPA1 in ALS spinal cord motor neurons with TDP‐43‐positive inclusions

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by loss of motor neurons and appearance of skein‐like inclusions. The inclusions are composed of trans‐activation response (TAR) DNA‐binding protein 43 (TDP‐43), a member of the heterogeneous nuclear ribonucleoprotein (hnRNP) family. hnRNPA1 and hnRNPA2/B1 are hnRNPs that interact with the C‐terminus of TDP‐43. Using immunohistochemistry, we investigated the association between TDP‐43 and hnRNPA1 in ALS spinal motor neurons. We examined spinal cords of seven ALS cases and six muscular dystrophy cases (used as controls) for the presence of TDP‐43 and hnRNPA1 protein. In the control cases, hnRNPA1 immunoreactivity in motor neurons was intense in the nucleus and weak in the cytoplasm where it showed a fine granular appearance. In the ALS cases, hnRNPA1 immunoreactivity in motor neurons was reduced in the nuclei of neurons with skein‐like inclusions but was not detected in the skein‐like inclusions. The marked loss of hnRNPA1 in motor neurons with concomitant cytoplasmic aggregation of TDP‐43 may represent a severe disturbance of mRNA processing, suggesting a key role in progressive neuronal death in ALS.

Prion protein oligomers in Creutzfeldt-Jakob disease detected by gel-filtration centrifuge columns.

Prion diseases are diagnosed by the detection of accumulation of abnormal prion protein (PrP) using immunohistochemistry or the detection of protease‐resistant abnormal PrP (PrPres). Although the abnormal PrP is neurotoxic by forming aggregates, recent studies suggest that the most infectious units are smaller than the amyloid fibrils. In the present study, we developed a simplified method by applying size‐exclusion gel‐filtration chromatography to examine PrP oligomers without proteinase K digestion in Creutzfeldt‐Jakob disease (CJD) samples, and evaluated the correlation between disease severity and the polymerization degree of PrP. Brain homogenates of human CJD and non‐CJD cases were applied to the gel‐filtration spin columns, and fractionated PrP molecules in each fraction were detected by western blot. We observed that PrP oligomers could be detected by the simple gel‐filtration method and distinctly separated from monomeric cellular PrP (PrPc). PrP oligomers were increased according to the disease severity, accompanied by the depletion of PrPc. The separated PrP oligomers were already protease‐resistant in the case with short disease duration. In the cases with quite severe pathology the oligomeric PrP reached a plateau, which may indicate that PrP molecules could mostly develop into amyloid fibrils in the advanced stages. The increase of PrP oligomers correlated with the degree of histopathological changes such as spongiosis and gliosis. The decrease of monomeric PrPc was unexpectedly obvious in the diseased cases. Dynamic changes of both oligomerization of the human PrP and depletion of normal PrPc require further elucidation to develop a greater understanding of the pathogenesis of human prion diseases.

Trends in autopsy-verified dementia prevalence over 29 years of the Hisayama study

We investigated the trends in dementia over the past 29 years in the town of Hisayama, Japan using 1266 autopsy specimens. The Hisayama study is a prospective cohort study of lifestyle‐related diseases that was started in 1961. Clinical examination of dementia was started in 1985 with five detailed cross‐sectional assessments conducted in 1985, 1992, 1998, 2005 and 2012. To examine the trends in dementia, we divided the 1266 autopsy samples into five groups according to the year of death: I (1986–1991, 257 cases), II (1992–1997, 268 cases), III (1998–2004, 318 cases), IV (2005–2011, 296 cases) and V (2012–2014, 127 cases). The prevalence of all‐cause dementia significantly increased over time (28.4% in group I, 22.4% in group II, 32.1% in group III, 30.1% in group IV, 51.2% in group V; P for trend <0.001). A similar trend was observed for Alzheimers disease (AD) (15.2%, 11.9%, 17.3%, 20.6% and 33.1%, respectively; P for trend <0.001). A significant increasing trend was observed in both men and women. A rapid increase in senile dementia of the NFT type (SD‐NFT) in recent years was notable. Vascular dementia was the most common type of dementia in men prior to 2004; however, its prevalence decreased over time. Our study revealed that tauopathies, including AD and SD‐NFT, significantly increased in the aged Japanese population over the course of this study. The neuritic plaque pathology of AD was associated with metabolic disorders such as insulin resistance and abnormal lipid metabolism, whereas the risk factors for tau pathology remain unclear. Although aging is considered one of the important risk factors accelerating tau pathology, there could be other risk factors associated with lifestyle diseases.

Elevated expression of fatty acid synthase and nuclear localization of carnitine palmitoyltransferase 1C are common among human gliomas

Fatty acid synthase (FASN) and carnitine palmitoyltransferase 1C (CPT1C), a brain‐specific isoform of the CPT1 family, are upregulated in certain types of cancers, including gliomas. Acetyl‐CoA carboxylase (ACC) catalyzes the carboxylation of acetyl‐CoA to malonyl‐CoA, the rate‐limiting step in fatty acid synthesis, and its phosphorylated form inhibits lipid synthesis. We examined the expression and subcellular localization of these fatty acid metabolism‐related molecules in human gliomas. We performed immunostaining of two glioma cell lines (U373MG and U87MG) and 41 surgical specimens of diffuse gliomas with various histological grades (21 with the isocitrate dehydrogenase 1(IDH1) R132H mutation and 20 without the mutation). In the cultured glioma cells, CPT1C and phosphorylated ACC (p‐ACC) were mainly localized to the nuclei, whereas FASN localized to the cytoplasm. In the surgical specimens, most glioma tissues showed nuclear staining for CPT1C and p‐ACC, and cytoplasmic staining for FASN, regardless of the genetic status of IDH1 and the histological grade. Therefore, elevated cytoplasmic expression of FASN and nuclear localization of CPT1C are common among human diffuse gliomas, which may be regulated by the differential phosphorylation status of ACC in the cellular compartment.

Microsphere formation in a subtype of Creutzfeldt-Jakob disease with a V180I mutation and codon 129 MM polymorphism.

Creutzfeldt–Jakob disease (CJD) with a point mutation of valine to isoleucine at codon 180 in the gene for PrP (V180I CJD) is genetic form of CJD found most frequently in Japan [1]. There are several reports of autopsied V180I CJD cases [2,3], in which the examples with methionine and valine heterozygosity at the polymorphic codon 129 were found at a higher rate compared with the normal population in Japan [4]. Here, we report a patient with numerous amyloid microsphere formations in an unusual subtype of CJD with a V180I MM mutation who underwent a continuous intraventricular infusion of pentosan polysulphate (PPS). Western blot analysis for proteaseresistant PrP (PrP) revealed the characteristic two bands devoid of a diglycoform. Size exclusion gel chromatography showed that diglycoform PrPs were present mainly in a monomeric state, and less in an oligomeric state, and that the diglycoform PrPs were completely degraded by proteinase K (PK) treatment. These PrP-positive structures and PrP molecular species were unique to genetic CJD with V180I and 129 MM. The patient showed geographical disorientation and memory deficits at the age of 64. Diffusion weighted image of an magnetic resonance imaging showed abnormal high-intensity signals in the left cerebral cortices. Analysis of the PRNP gene revealed a V180I mutation with codon 129 MM polymorphism. PPS infusion was administered from 4 months after the onset of symptoms at the maximum dosage of 120 mg/kg/day. The patient progressively worsened and bilateral subdural effusion was seen on the computed tomography scan. Neither myoclonus nor periodic synchronous discharge on EEG was observed. The patient died of pneumonia with a 6-year clinical course. The patient was described as case No. 4 in the previous paper, which showed clinical data of all 11 patients undergoing an intraventricular PPS administration [5]. At autopsy, the brain weighed 484 g and showed severe atrophy with bilateral subdural fluid collection. In the coronal sections, diffuse atrophy of the cerebrum was noted, but the hippocampus, cerebellum and brainstem were relatively preserved from atrophy. For histological examination, haematoxylin and eosin, Congo red, dylon and Klüver–Barrera stains were performed. Immunohistochemistry was performed using the primary antibodies for PrP (mouse monoclonal 3F4, 1:400; Signet, Cambridge, MA, USA, mouse monoclonal 8G8, 1:400; Cayman, Ann Arbor, MI, USA, mouse monoclonal 8B4, 1:400; Santa Cruz Biotechnology, Santa Cruz, CA, USA, mouse monoclonal 8H4, 1:400; Abcam, Cambridge, UK) and beta-amyloid (mouse monoclonal 6F/3D, 1:400; Dako, Glostrup, Denmark). The study protocol was approved by the Human Ethics Review Committee of the Graduate School of Medical Sciences, Kyushu University. The cerebral cortices showed two different pathological patterns according to the depth of the cortical layers. While the outer and middle neocortical layers showed mild histological changes, its inner layers showed more severe neuronal loss, spongiform change and gliosis (Figure 1A). Additionally, immunohistochemistry results revealed many spherical PrP-positive deposits exclusively at the outer to the middle cortical layers (Figure 1B). The spherical deposits were small and round, and we tentatively named them microspheres. Spongiform changes around the microspheres were rather mild (Figure 1C), while synaptic PrP depositions were enhanced at the inner cortical layer with severe spongiform changes (Figure 1D). The hippocampus was relatively preserved from spongiform change but mild synaptic PrP depositions were seen in the molecular layer. Many microspheres were also noted in the layer of the pre-a neurone at the entorhinal cortex and the transentorhinal cortex (Figure S1A,B). The putamen, globus pallidus and thalamus showed mild PrP synaptic depositions. The cerebellar cortex showed mild spongiform changes with microspheres, which were focally observed in the molecular layer (Figure S1C). In the midbrain, numerous microspheres as well as PrP synaptic depositions were noted in the central grey matter (Figure S1D). There was no laterality of histopathological changes or PrP deposition. The distribution of microspheres is shown in Figure S1E. The PrP-positive spherical structures were almost equal in size and had smooth surfaces, while amyloid plaques of

Inflammatory radiculoneuropathy in an ALS4 patient with a novel SETX mutation

Amyotrophic lateral sclerosis 4 (ALS4), a rare form of autosomal dominant juvenile-onset ALS characterised by slow progression and sparing of bulbar and respiratory muscles,1 2 results from missense mutations in the senataxin ( SETX ) gene.3 This study reports the clinical pathology of a male Japanese ALS4 patient carrying a novel mutation in the SETX gene who presented with coexistent inflammatory radiculoneuropathy. Slight delays in early development resulted in the patient beginning to walk at 1.5 years. He was prone to falls and had suffered from pes cavus since childhood. At 35 years of age, the patient presented with difficulty walking. One year later he found it difficult to fully extend his fingers and this was exacerbated over the following 3 months. Nerve conduction studies revealed asymmetric demyelinating patterns (median nerve motor conduction velocity: right 29 m/s, left 53 m/s). Subsequently, the patient developed a neurogenic bladder. Intravenous immunoglobulin therapy (20 g/day for 5 days) resulted in mild improvement in muscle weakness (one point on the manual muscle test) and in the amplitude of compound motor action potentials for the median (right 1.982 mV to 3.362 mV, left 1.342 mV to 2.062 mV) and right ulnar nerve (6.064 mV to 8.296 mV). At 37 years of age, the patient developed progressive distal weakness and sensory disturbances in the right forearm and medial thigh. Following two courses of intravenous methylprednisolone (1 g/day for 3 days; steroid pulse therapy) and subsequent oral prednisolone treatment (50 mg/day with gradual tapering), sensory impairment and muscle weakness both improved. Thereafter, the patient underwent repeated steroid pulse therapy when muscle weakness became worse. At 41 years of …