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Biomedical subjects

N Takahata

Publications and source records attributed to N Takahata.

At least 37 records · Page 2Linked to original sources
PubMed1998

Evidence for presenilin-1 involvement in amyloid angiopathy in the Alzheimer's disease-affected brain.

Presenilin-1 (PS-1) has been identified as the protein encoded by the chromosome 14 locus that, when mutated, leads to familial Alzheimer's disease (FAD). The role PS-1 plays in the pathogenesis of Alzheimer's disease (AD) remains unclear. Using a set of antibodies raised against PS-1 synthetic peptides, polyclonal antibody to amyloid beta protein (Abeta) and end-specific antibodies against Abeta40, and Abeta42, immunohistochemical studies were performed on brain sections obtained from AD cases and controls. The PS-1 antibodies clearly stained amyloid angiopathies in AD-affected brains, but no recognizable immunoreactions were observed in any other vessels free from amyloid involvement in either AD-affected brains or controls. Abeta antibodies and the end-specific antibody against Abeta40 also decorated amyloid angiopathies, showing localization similar to that of PS-1. Western blot analyses predominantly detected protein band polypeptide species of a 50 kDa, band, presumably full-length PS-1 protein with N-terminus antisera, since these antibodies turned out to recognize a 50-kDa full-length band in cell lysate of transfected HeLa cell overexpressing PS-1. In addition, we recognized 30, 27 and 25 kDa proteins in both AD and control brain homogenate with these antibodies. In microvessel fractions extracted from brain homogenates, the 50, and 27 kDa fragments were observed in AD-affected brains but not in those of controls. C-terminus rabbit antisera reacted strongly with the 33 and 27 kDa bands, and additionally detected a small amount of full-length PS-1 protein in extracts from AD and control brains. Our present data indicate that PS-1 might be involved in the pathogenesis of amyloid angiopathy in the AD brain.

Alzheimer Disease↗
PubMed1998

Measurement of receptor-mediated functional activation of G proteins in postmortem human brain membranes.

Guanine nucleotide-binding regulatory proteins (G proteins) play a pivotal role in receptor-mediated transmembrane signal transduction, and have been implicated in modes of action of psychotropic drugs as well as in pathogenesis of psychiatric disorders. In the present investigation, functional activation of G proteins coupled with several receptors, in particular with GABAB receptors, was assessed by agonist-induced stimulation of high-affinity GTPase, an enzyme that is intrinsic to alpha subunit of G protein, in postmortem human frontal cortical membranes. High-affinity GTPase activity was stimulated by GABA as well as (+/-)-baclofen, a selective GABAB receptor agonist, with EC50 values of 60-150 and 10-40 microM, respectively, in a Mg(2+)-dependent manner. The (+/-)-baclofen-stimulated response was antagonized by the selective GABAB receptor antagonist, 2-hydroxy-saclofen, in a competitive manner with a KB value of 59 microM. Although the maximal percent increase above basal value (% Emax) for GABAB receptor-mediated high-affinity GTPase activity was varied from subject to subject, % Emax values for both agonists were highly correlated with each other, and replicable and stable in a given subject, indicating that this measure is trustworthy as an index of functional coupling between receptors and G proteins in future studies at the aim of elucidating possible alteration of receptor/G protein interaction in psychiatric disorders. The % Emax values for GABAB receptor-mediated responses were correlated inversely with brain storage duration, which should be critically considered in postmortem studies. The increases in high-affinity GTPase activity stimulated by several agonists other than GABAB receptor agonists seemed too low to quantify for making a comparison in future studies.

Aged↗
PubMed1998

Evolutionary history and mechanism of the Drosophila cecropin gene family.

Upon bacterial infection, insects secrete a set of synthesized antibacterial proteins into the hemolymph and initiate synergistic destruction of invaders. Cecropin is one such antibacterial protein which is also found in vertebrates. To study the evolutionary history and mechanism of the Cecropin gene family, we determined DNA sequences of one isogenic In(3R)C and six isofemale lines of Drosophila melanogaster as well as one line of D. simulans and of D. yakuba. The phylogenetic analysis of these sequences together with those published for D. virilis and Sarcophaga peregrina reveals frequent gene re-organization. It was also found that silent nucleotide differences within D. melanogaster are quite heterogeneous across the gene region of approximately 3 kilobases and the extent of polymorphism is unusually usually high. These data suggest that the Cecropin gene region of D. melanogaster underwent intragenic recombination as well as introgression from a closely related sibling species, D. simulans.

Animals↗
PubMed1998

Footprints of intragenic recombination at HLA loci.

To evaluate the effect of balancing selection and intragenic recombination (or gene conversion) at six individual HLA loci, synonymous nucleotide diversity in different exon groups is examined within (pi w) and between (pi b) allelic lineages that may be defined by either serological or DNA sequence differences. Both pi values are high in exons which encode for the peptide binding region (PBR) and tend to decrease in other exons. The value of pi w is significantly smaller than that of pib in any exon of any locus. However, even pi w is much greater than nucleotide diversity at non-HLA loci. These observations provide additional strong evidence for the operation of balancing selection in PBR-encoding exons and its indirect effects on polymorphism at linked neighboring regions. It appears that allelic lineages have generally evolved in isolation but the linkage relationships within and between exons are incomplete throughout the long evolutionary history. To quantify intragenic recombination and account for the large discrepancy between the HLA and non-HLA diversity, a population genetics model is analyzed with special reference to the evolution of modern humans. The analysis suggests that the recombination rate between two sites 1000 base pairs apart is about 10(-5) per generation and that the effective size of human populations (equivalent roughly to the number of breeding individuals in a randomly mating population) has dropped from 10(5) to 10(4) in most of the Quaternary. One possibility for this reduction is discussed.

Chromosome Mapping↗
PubMed1998

Polymorphism of the HLA class II loci in Siberian populations.

The populations that colonized Siberia diverged from one another in the Paleolithic and evolved in isolation until today. These populations are therefore a rich source of information about the conditions under which the initial divergence of modern humans occurred. In the present study we used the HLA system, first, to investigate the evolution of the human major histocompatibility complex (MHC) itself, and second, to reveal the relationships among Siberian populations. We determined allelic frequencies at five HLA class II loci (DRB1, DQA1, DQB1, DPA1, and DPB1) in seven Siberian populations (Ket, Evenk, Koryak, Chukchi, Nivkh, Udege, and Siberian Eskimo) by the combination of single-stranded conformational polymorphism and DNA sequencing analysis. We then used the gene frequency data to deduce the HLA class II haplotypes and their frequencies. Despite high polymorphism at four of the five loci, no new alleles could be detected. This finding is consistent with a conserved evolution of human class II MHC genes. We found a high number of HLA class II haplotypes in Siberian populations. More haplotypes have been found in Siberia than in any other population. Some of the haplotypes are shared with non-Siberian populations, but most of them are new, and some represent "forbidden" combinations of DQA1 and DQB1 alleles. We suggest that a set of "public" haplotypes was brought to Siberia with the colonizers but that most of the new haplotypes were generated in Siberia by recombination and are part of a haplotype pool that is turning over rapidly. The allelic frequencies at the DRB1 locus divide the Siberian populations into eastern and central Siberian branches; only the former shows a clear genealogical relationship to Amerinds.

Alleles↗
PubMed1998

Calcium mobilization evoked by amyloid beta-protein involves inositol 1,4,5-triphosphate production in human platelets.

We examined the effects of amyloid beta-protein (A beta) on Ca2+ mobilization in human platelets. The addition of A beta fragments 25-35 (A beta 25-35) gradually increased the cytoplasmic free Ca2+ concentration ([Ca2+]i). After the maximum response, [Ca2+]i decreased and then reached a sustained, higher level of [Ca2+]i. Similar effects were also observed with A beta 1-40, whereas 1-28 , 12-28 and 31-35 did not affect the Ca2+ response. In the absence of extracellular Ca2+, A beta 25-35 caused a transient increase in [Ca2+]i, which returned to the resting level. U73122, a phospholipase C inhibitor, completely abolished Ca2+ mobilization induced by thrombin and A beta 25-35. Furthermore, A beta enhanced the production of inositol 1,4,5-trisphosphate (IP3) in platelets. These findings suggest that Ca2+ mobilization induced by A beta 25-35 is due to phospholipase C activation and IP3 production.

Amyloid beta-Peptides↗
PubMed1998

Differential alteration of adenylyl cyclase subtypes I, II, and V/VI in postmortem human brains of heroin addicts.

In animal and culture cell experiments, the upregulation of cAMP-related signal transduction after chronic opioid administration has been hypothesized to be an adaptive change of the molecular mechanism to maintain homeostasis in intracellular signals downstream from opioid receptors. Herein, we have examined the quantitative changes of three adenylyl cyclase (AC) subtypes (I, II, and V/VI) in temporal cortex membranes from brains of heroin addicts and age-matched controls by immunoblotting. The immunoreactivity of AC-I decreased significantly (p < 0.05) in heroin addicts, compared with controls; whereas those of AC-II and AC-V/VI were not changed. The present findings indicate that differential regulation of AC subtypes occurs and that AC-I may play an important role in the signal transduction for opiate-induced tolerance and dependence mechanisms in human brain cortex.

Adenylyl Cyclases↗
PubMed1998

Reduced immunoreactivity of type I adenylyl cyclase in the postmortem brains of alcoholics.

Reduced adenylyl cyclase activity after chronic ethanol exposure has been reported. In this study, we investigated by immunoblotting whether quantitative changes of adenylyl cyclase isoforms (type I, type II, and type V/VI adenylyl cyclases) exist in membrane preparations of the temporal cortex obtained from six alcoholics and six age-matched controls. The immunoreactivity of type I adenylyl cyclase decreased significantly in the temporal cortex of alcoholics when compared with controls (p < 0.05), whereas those of type II and type V/VI adenylyl cyclases showed no changes between the groups. These findings suggest that these isoform-specific afterations in the adenylyl cyclase system may be involved in the pathophysiology of alcoholism.

Adenylyl Cyclases↗
PubMed1998

Differential effects of chronic administration of the antidepressants amitriptyline and rolipram on adenylyl cyclase activity.

Significant increases were observed in guanylylimidodiphosphate- and fluoride-stimulated adenylyl cyclase activities in synaptic membrane preparations from rat cerebral cortex subsequent to the repeated administration of rolipram and amitriptyline. The potency of GppNHp-dependent inhibition of adenylyl cyclase was decreased by chronic treatment with rolipram but not by amitriptyline treatment. These findings suggest that chronically administered rolipram and amitriptyline share the augmentation of adenylyl cyclase activity by different actions on post-receptor signaling.

Adenylyl Cyclases↗
PubMed1998

Selection, convergence, and intragenic recombination in HLA diversity.

To account for high degrees of human leukocyte antigen (HLA) diversity, a method is proposed for detecting intragenic recombination or gene conversion separately from parallel substitutions or convergent evolution. An application of the method to HLA protein sequences suggests that intragenic recombination played important roles in HLA-B and DPB1, some in HLA-A and DRB1, and least in HLA-C and DQB1 diversity. However, the extent of diversity of these molecules does not necessarily correlate with the frequency of intragenic recombination, supporting the view that (balancing) selection is a primary agent of HLA diversity and often leads to convergent evolution. Computer simulation is carried out to examine two models of balancing selection under the coupled effect with mutation, intragenic recombination, and random drift in a diploid population. It is emphasized that break points by intragenic recombination need be specified to account for HLA diversity. Implications of HLA diversity in human evolution are briefly discussed.

Amino Acid Sequence↗
PubMed1997

Amino-terminus truncated apolipoprotein E is the major species in amyloid deposits in Alzheimer's disease-affected brains: a possible role for apolipoprotein E in Alzheimer's disease.

Amyloid deposits in Alzheimer's disease (AD) are composed of amyloid beta protein (A beta) and many other components called amyloid-associated proteins. Apolipoprotein E (apoE) is one of the most important amyloid-associated proteins. The role apoE plays in AD, however, is yet to be determined. In this study, we present the biochemical and histochemical nature of apoE in AD-affected brains using four monoclonal antibodies (mAbs) against apoE and newly established antibodies against the amino-terminal (anti-apoE-N), and carboxyl-terminal regions (anti-apoE-C) of apoE. Competitive ELISA and Western-blot analysis combined with thrombolytic digestion of apoE indicated that our four mAbs recognized at least two different epitopes within a 22-kDa amino-terminal domain of apoE. Using these mAbs and an anti-A beta mAb, double immunostaining showed that the majority of amyloid deposits were stained by both anti-apoE and anti-A beta mAbs, but the minority of them were detected only by either anti-apoE or anti-A beta mAbs. Differences in staining properties between anti-apoE-N and anti-apoE-C were that anti-apoE-C recognized both amyloid deposits and astrocytes similar to anti-apoE mAbs, but anti-apoE-N strongly stained only astrocytes. Preliminary semi-quantitative determinations of apoE in CSF and brain homogenate showed that the amount of apoE increased in AD and Creutzfeldt-Jakob disease brains compared to normal samples. Our immunological data, using antibodies specific for the amino and carboxyl termini of apoE, suggest that apoE may, in some circumstances, initiate plaque formation, and that apoE in amyloid deposits has at least part of its amino termini cleaved out.

Alzheimer Disease↗
PubMed1997

Amyloid precursor protein, A beta and amyloid-associated proteins involved in chloroquine retinopathy in rats--immunopathological studies.

To understand the retinal changes in Alzheimer disease (AD) patients, pathological and immunocytochemical studies were performed on retinal cells in the chloroquine-treated rats at 0, 4, 8, 12, 16, 20, and 24 weeks after the initial injection, using anti-amyloid precursor protein (APP), -amyloid beta protein (A beta), -apolipoprotein E (apoE), -ubiquitin, and -cathepsin D antibodies. Pathological alterations consistent with chloroquine retinopathy were recognized in the ganglion cells of the ganglion cell layer (GCL) and the inner plexiform layer (IPL) 4 weeks after initial chloroquine injection. Rat retinal changes appear to have a direct relationship to the duration of chloroquine administration. Intense immunoreactivities for anti-APP, A beta, apoE (an associated protein), and ubiquitin co-localized in the swollen ganglion cells and Muller cells by 20-24 weeks together with the lysosomal enzyme cathepsin D. The present data indicate that the endosomal/lysosomal pathway plays an important role in the processing of APP in rat retina. This experimental model is considered to be a suitable neural model to understand retinal pathology and the processing of APP in terms of the pathogenesis of AD, whereas chloroquine-induced myopathy is a useful extra neuronal model.

Amyloid↗
PubMed1997

Evolution of the primate lineage leading to modern humans: phylogenetic and demographic inferences from DNA sequences.

To date major divergences that occurred in the primate lineage leading to modern humans and to infer a demographic parameter (effective population size) of the ancestral lineage that existed at each divergence, a maximum likelihood method was applied to autosomal DNA sequence data currently available for pairs of orthologous genes between the human and each of the chimpanzee, gorilla, Old World monkey (OWM), and New World monkey (NWM). A statistical test is carried out to support the assumption that silent substitutions have accumulated in a clock-like fashion over loci between primate taxa or even among sites within a locus. It is shown that the human ancestral lineage became distinct from the NWM 57.5 million years (Myr) ago, the OWM 31 Myr ago, the gorilla 8.0 Myr ago, and the chimpanzee 4.5 Myr ago, and that the effective population size at these divergences was generally much greater than that of modern humans. It is argued that the human ancestral lineage branched off from the NWM and OWM earlier than once thought and that significant demographic changes might have occurred at different evolutionary stages, particularly at the hominid stage.

Animals↗
PubMed1997

Amyloid beta protein and transthyretin, sequestrating protein colocalize in normal human kidney.

The localization of amyloid beta protein (A beta), A beta 40, A beta 42, and transthyretin (TTR) was investigated immunohistochemically in the autopsied human kidney, using polyclonal antibodies against TTR, A beta and C-terminal end-specific antibodies against A beta 40 and 42. Immunoreactivities of A beta and A beta 40 were found both in the proximal and distal tubular epithelial cells. But the immunolocalization of A beta 40 was observed predominantly in the distal tubules whereas that of A beta 42 was predominantly recognized in the proximal tubules. TTR, sequestrating protein for A beta, was present in the proximal tubules. The mechanism by which A beta does not form amyloid in Alzheimer's disease outside the brain remains unknown. The tubular epithelial cells in the kidney may provide a useful system to shed light on this issue.

Amyloid beta-Peptides↗
PubMed1997

Ca2+/CaM-sensitive adenylyl cyclase activity is decreased in the Alzheimer's brain: possible relation to type I adenylyl cyclase.

Immunoreactivities of four subtypes of adenylyl cyclase (AC) (types I, II, IV and V/VI), and basal, forskolin- and Mn(2+)-stimulated AC activities with or without calcium and calmodulin (Ca2+/CaM) were estimated in parietal cortex membranes from cases with dementia of the Alzheimer type (DAT) and age-matched controls. Immunoreactivities of AC-I and AC-II were significantly decreased, but those of AC-IV and AC-V/VI did not change in DAT brains. There was a significant correlation of AC-I immunoreactivity with Ca2+/CaM-sensitive AC activity, but not with the Ca2+/CaM-insensitive activity. Ca2+/CaM-sensitive AC activity was significantly lower in DAT than in the control, indicating that impairment of Ca2+/CaM-sensitive AC-I is clearly involved in the pathophysiology of DAT.

Adenylyl Cyclases↗
PubMed1997

Dimeric tubulin-stimulated adenylyl cyclase activity is augmented after long-term amitriptyline treatment.

We have investigated altered association of tubulin dimers interacting with G proteins and modulating adenylyl cyclase (AC) as a result of long-term amitriptyline (AMT) treatment. Gpp(NH)p-stimulated, but not basal or manganese-stimulated, AC activity was significantly augmented in cortex membranes prepared from rats chronically treated with AMT. The enhancement of AC activity by Gpp(NH)p-liganded tubulin (tubulin-Gpp(NH)p) was significantly higher in chronically AMT-treated rats than in control rats. Moreover, in cortex membranes from controls, tubulin-Gpp(NH)p prepared from chronically AMT-treated rats was more effective to activate AC activity than tubulin-Gpp(NH)p from controls. Immunoblotting and photoaffinity guanine nucleotide binding procedures showed no significant differences in the amount and the function of G proteins between controls and AMT-treated groups. It is suggested that long-term AMT treatment causes alteration in the functional interaction between tubulin and G protein, and this modification may participate in enhanced coupling of Gs to the catalytic subunit of AC induced by the chronic antidepressant treatment.

Adenylyl Cyclases↗
PubMed1997

Potential antidepressant properties of forskolin and a novel water-soluble forskolin (NKH477) in the forced swimming test.

The mechanisms of the antidepressant activity of forskolin and a novel water soluble forskolin analog (NKH477) were studied using the forced swimming method in rats. Forskolin (0.01-0.1 mg/kg) and NKH477 (0.01-0.1 mg/kg) dose-dependently decreased ratings of immobility, with effects similar to those of amitriptyline treatment. The maximum effects of forskolin and NKH477 were observed at 0.01 mg/kg dose which is 150 more times potent than that (15 mg/kg) of amitriptyline. At a high dose (1.0 mg/kg) of forskolin and NKH477, the duration of immobility was returned to control levels. Forskolin and NKH477 did not influence the spontaneous locomotor activity at intraperitoneal injection doses from 0.01 to 1 mg/kg. Furthermore chronic administration with NKH477 at oral dose from 0.5 to 1.5 mg/kg significantly decreases the duration of immobility. These data indicate that both forskolin and NKH477 have strong antidepressive potency, consistent with the hypothesis that elevation of the cAMP cascade system may have an important role in antidepressive effects.

Administration, Oral↗