Biomedical subjects

N Yamaguchi

Publications and source records attributed to N Yamaguchi.

At least 73 records · Page 4Linked to original sources

PubMed2001

Abnormal activation of glial cells in the brains of prion protein-deficient mice ectopically expressing prion protein-like protein, PrPLP/Dpl.

BACKGROUND: Some lines of mice homozygous for a disrupted prion protein gene (Prnp), including Ngsk Prnp(0/0) mice, exhibit Purkinje cell degeneration as a consequence of the ectopic overexpression of the downstream gene for prion protein-like protein (PrPLP/Dpl) in the brain, but others, such as Zrch I Prnp(0/0) mice, show neither the neurodegeneration nor the expression of PrPLP/Dpl. In the present study, we found that Ngsk Prnp(0/0), but not Zrch I Prnp(0/0) mice, developed gliosis involving both astrocytes and microglia in the brain. MATERIALS AND METHODS: The brains from wild-type (Prnp(+/+)), Ngsk Prnp(0/0), Zrch I Prnp(0/0), and reconstituted Ngsk Prnp(0/0) mice carrying a mouse PrP transgene, designated Tg(P) Ngsk Prnp(0/0) mice, were subjected into Northern blotting and in situ hybridization using probes of glial fibrillary acidic protein (GFAP) and lysozyme M (LM) specific for astrocytes and microglia, respectively. Immunohistochemistry was also performed on the brain sections using anti-GFAP and anti-F4/80 antibodies. RESULTS: Northern blotting demonstrated upregulated expression of the genes for GFAP and LM in the brains of Ngsk Prnp(0/0), but not in Zrch I Prnp(0/0) mice. A transgene for normal mouse PrP(C) successfully rescued Ngsk Prnp(0/0) mice from the glial activation. In situ hybridization and immunohistochemistry revealed activated astrocytes and microglia mainly in the white matter of both the forebrains and cerebella. In contrast, there was no evidence of neuronal injury except for the Purkinje cell degeneration. Moreover, the glial cell activation was notable well before the onset of the Purkinje cell degeneration. CONCLUSIONS: These findings strongly suggest that ectopic PrPLP/Dpl in the absence of PrP(C) is actively involved in the glial-cell activation in the brain.

Aging↗

PubMed2001

Pituitary adenylate cyclase activating polypeptide enhances glucose-evoked insulin secretion in the canine pancreas in vivo.

OBJECTIVE: To study a local effect of pituitary adenylate cyclase activating polypeptide (PACAP(1-27)) on glucose-evoked insulin release under in vivo conditions. INTERVENTION: Glucose and PACAP(1-27) were locally infused to the pancreas via the superior pancreaticoduodenal artery without interrupting the blood supply. MAIN OUTCOME MEASURES: Plasma insulin and glucose concentrations were determined in samples obtained from the superior pancreaticoduodenal vein and the aorta. Superior pancreaticoduodenal venous blood flow was measured to compute the net output of insulin. RESULTS: PACAP(1-27) (0.005-5 microg) increased the basal insulin secretion by about 15 folds in a dose-dependent manner. Local infusion of either glucose (5%) or PACAP(1-27) (0.05 microg) resulted in a significant increase in the basal insulin output to about 300 microU x min(-1)g(-1), which was highly reproducible upon the second administration of the same dose with an interval of 30 min. When PACAP(1-27) was simultaneously given during glucose infusion, the increased insulin output due to glucose was further enhanced to about 600 microU x min(-1)g(-1). The net increase in PACAP(1-27)-induced insulin output in the presence of glucose was significantly greater than that obtained with PACAP(1-27) alone. There exists a strong and highly significant correlation between changes in glucose level and those in insulin output when both glucose and PACAP(1-27) were administered simultaneously. CONCLUSION: The results indicate that PACAP(1-27) directly enhances the glucose-evoked insulin secretion in the endocrine pancreas in anesthetized dogs. The study suggests that PACAP may play a local facilitating role in insulin secretion in response to glucose loading.

Animals↗

PubMed2000

The immediate early gene 1 product of human cytomegalovirus is sufficient for up-regulation of interleukin-8 gene expression.

We previously observed that human cytomegalovirus (CMV) infection induced a massive production of a chemokine with potent neutrophil chemotactic activity, interleukin-8 (IL-8). Hence, we examined the effect of CMV immediate early (IE) gene products on IL-8 production by the human astrocytoma cell line, U373MG. Transient or stable transfection with a CMV IE1 gene expression vector, but not with a IE2 gene expression vector, significantly augmented IL-8 protein secretion and IL-8 mRNA expression. Luciferase activity was enhanced in U373MG cells when the cells were cotransfected with CMV IE1 and chimeric firefly luciferase reporter genes driven by the transcriptional regulatory region of the human IL-8 gene. Moreover, IE1 gene-mediated enhancement of luciferase activity was abolished by the introduction of mutations into the AP-1 or NF-kappa B factor binding elements in the regulatory region of the IL-8 promoter. Furthermore, electrophoretic mobility shift assays demonstrated that CMV IE1 gene products induced the formation of NF-kappa B or AP-1 complexes. Finally, Western blotting analysis demonstrated that the CMV IE1 gene product increased the amount of NF-kappa B complexes translocated into the nucleus. Collectively, CMV IE1 gene expression may be sufficient to activate AP-1 and NF-kappa B, resulting in IL-8 gene expression.

Base Sequence↗

PubMed2000

cDNA cloning and tissue-specific splicing variants of mouse hippostasin/TLSP (PRSS20).

Two splicing variants of mouse hippostasin/TLSP (PRSS20) were identified and termed brain-type and prostate-type, respectively. Mouse hippostasin/TLSP showed 76.8% identity to the human homologue. Transient expression showed that both translational products were secreted into the conditioned medium. Mouse hippostasin/TLSP was expressed preferentially in the fetal brain and the prostate, but not in the neonatal brain. The brain expressed only brain-type hippostasin/TLSP, while the prostate expressed both types.

Alternative Splicing↗

PubMed2000

Involvement of TWEAK in interferon gamma-stimulated monocyte cytotoxicity.

TWEAK, a new member of the tumor necrosis factor (TNF) family, induces cell death in some tumor cell lines, but its physiological functions are largely unknown. In this study, we investigated the expression and function of TWEAK in human peripheral blood mononuclear cells (PBMCs) by using newly generated anti-human TWEAK mAbs. Although freshly isolated PBMCs expressed no detectable level of TWEAK on their surfaces, a remarkable TWEAK expression was rapidly observed on monocytes upon stimulation with interferon (IFN)-gamma but not with IFN-alpha or lipopolysaccharide. Cytotoxic activity of IFN-gamma-stimulated monocytes against human squamous carcinoma cell line HSC3 was inhibited partially by anti-TWEAK mAb alone and almost completely by combination with anti-TRAIL (TNF-related apoptosis-inducing ligand) mAb. These results revealed a novel pathway of monocyte cytotoxicity against tumor cells that is mediated by TWEAK and potentiated by IFN-gamma.

Antibodies, Monoclonal↗

PubMed2000

A novel mutation of desert hedgehog in a patient with 46,XY partial gonadal dysgenesis accompanied by minifascicular neuropathy.

We describe a patient with 46,XY partial gonadal dysgenesis (PGD) who presented with polyneuropathy. Sural nerve pathology revealed peculiar findings characterized by extensive minifascicular formation within the endoneurium and with a decreased density of myelinated fibers. We found, in the patient, a homozygous missense mutation (ATG-->ACG) at the initiating codon in exon 1 of the desert hedgehog (DHH) gene, which predicts a failure of translation of the gene. The same heterozygous mutation was found in the patient's father. This is the first report of a human DHH gene mutation, and the findings demonstrate that mutation of the DHH gene may cause 46, XY PGD associated with minifascicular neuropathy.

Animals↗

PubMed2000

A fusion inhibitor (FP-21399) for the treatment of human immunodeficiency virus infection: a phase I study.

FP-21399 is a bis(disulfonaphthalene) derivative that prevents human immunodeficiency virus (HIV) infection of uninfected cells by blocking entry of the virus. FP-21399 shows an affinity for lymph nodes. In this phase I study, FP-21399 was administered intravenously over 1 h as a single dose (0.9, 1.7, 2.8, and 4.2 mg/kg) or as a once-weekly infusion (1, 2, and 3 mg/kg) for 4 consecutive weeks to 34 HIV-1 infected patients with CD4(+) cell counts of 50-400 cells/microL. Concomitant antiretroviral therapy was permitted but not required. The most frequent adverse events involved the transient, dose-dependent appearance of drug- or metabolite-related color in the urine and skin. Plasma drug levels were linear with dose. The drug was cleared, with an elimination half-life of 4 h and a terminal half-life of 1.5-2 days; the terminal half-life represented redistribution and clearance from tissues. FP-21399 administered weekly for 4 weeks was well tolerated. Further studies are necessary to define the role of this fusion inhibitor in the treatment of HIV infection.

Adult↗

PubMed2000

A novel isoform of a kallikrein-like protease, TLSP/hippostasin, (PRSS20), is expressed in the human brain and prostate.

cDNAs encoding two splicing variants of a serine protease, termed hippostasin, were isolated by a PCR-based cloning strategy. The difference of 5' nucleotide sequence resulted in the variation in the amino terminal ends of the two, brain and prostate, types of human hippostasin. The longest ORF of the brain-type was 250 amino acids with a putative signal peptide, while that of the prostate-type was 282 amino acids. Homology search using the amino acid sequence revealed that prostate-type hippostasin was identical to TLSP (PRSS20), which is expressed in human primary keratinocytes (1). Transient expression analysis showed that both brain- and prostate-type TLSP/hippostasin were secreted into the conditioned medium as about 40 kDa proteins. Human TLSP/hippostasin showed 47% and 45% identity to trypsinogen II and kallikrein, respectively. In fact, the recombinant human TLSP/hippostasin efficiently cleaved Bz-Phe-Arg-4-methylcoumaryl-7-amide, a kallikrein substrate, and weakly cleaved other substrates for kallikrein and trypsin. Northern blot analysis detected a 1.3 kb band in the whole brain and a 1.4 kb band in the prostate and the lung. In situ hybridization revealed that it was expressed preferentially by the pyramidal neurons in the human hippocampus and secretory epithelial cells in the prostate. These results indicated that TLSP/hippostasin is involved in the functions of the human central nervous system and prostate and that it is a multifunctional protease present in various organs.

Alternative Splicing↗

PubMed2000

Involvement of protein kinase C in taxol-induced polyploidization in a cultured sarcoma cell line.

Taxol was found to inhibit the proliferation and to induce the polyploidization of cultured methylcholanthrene-induced sarcoma cells (Meth-A cells). To investigate whether protein kinase C is involved in taxol-induced polyploidization, phorbol 12-myristate 13-acetate (PMA), which regulates the activity of protein kinase C, was used along with taxol to treat the cells. We found that PMA did not interfere with the proliferation and did not induce polyploidization by itself. However, at low concentration, taxol, which by itself did not induce polyploidization, clearly induced polyploidization in the presence of PMA. To explore the mechanism by which PMA potentiates polyploidization, the levels of the G1 checkpoint-related proteins cyclin E and cdk2, and those of the G2 checkpoint-related proteins cyclin B and cdc2 were determined by flow cytometry. We found that both G1 and G2 checkpoint-related proteins increased during the induction of polyploidization. To verify the relationship between protein kinase C and tubulin polymerization, flow cytometry was used to determine the total content of tubulin protein, and morphological observation was used to examine spindle organization. PMA did not affect the taxol-induced increase in tubulin protein, but markedly potentiated taxol-induced spindle disorganization. These findings suggest that protein kinase C plays an important role in regulating the induction of polyploidization in Meth-A cells.

Animals↗

PubMed2000

Expression of type XVI collagen in cultured skin fibroblasts is related to cell growth arrest.

The expression of type XVI collagen in various phases of cell growth in cultured skin fibroblasts was studied. A marked increase in type XVI collagen mRNA level was found in stationary phases of cell growth (non-adherent and confluent phases), whereas the expression of type I and III collagens was undetectable in the non-adherent phase but became greater in the confluent phase. When suspended cells were further cultured over 72 h (suspension arrest), mRNA level and gene transcription of type XVI collagen were time-dependently increased whereas those of type I collagen remained undetectable. When the confluent cells were further cultured for 72 h under the condition of serum deprivation (serum deprivation arrest), mRNA levels of both type XVI collagen and type I collagen were elevated. The level of type XVI collagen polypeptide in the culture media of suspension-arrested and serum deprivation-arrested cells paralleled the mRNA level of type XVI collagen. The results indicate that expression of type XVI collagen (a member of the fibril-associated collagens with interrupted triple helices), unlike interstitial collagens (type I collagen), is related to cell growth arrest brought about by two different growth inhibiting systems, suspension arrest and serum deprivation arrest.

Cell Adhesion↗

PubMed2000

Suppression of GD1alpha ganglioside-mediated tumor metastasis by liposomalized WHW-peptide.

GD1alpha ganglioside-replica peptides were recently isolated from a phage-displayed random pentadecapeptide library by assaying for inhibition of adhesion of RAW117-H10 lymphosarcoma cells to hepatic sinusoidal microvessel endothelial (HSE) cells. We show here that the Trp-His-Trp (WHW) peptide was identified as a minimal sequence of the GD1alpha-replica peptide WHWRHRIPLQLAAGR. The addition of WHW peptide-attached liposomes displayed efficient inhibition of liver metastasis of RAW117-H10 cells as well as of GD1alpha-mediated adhesion of RAW117-H10 cells to HSE cells in vitro. These results suggest that engineered liposomes for peptide delivery are applicable to treatment for metastasis.

Amino Acid Sequence↗

PubMed2000

Enhanced expression and DNA binding activity of two CCAAT/enhancer-binding protein isoforms, C/EBPbeta and C/EBPdelta, in rheumatoid synovium.

OBJECTIVE: To investigate the activation and expression of CCAAT/enhancer-binding proteins (C/EBP), especially C/EBPbeta and -delta, in rheumatoid synovium, and their pathogenic implications in rheumatoid arthritis (RA). METHODS: The activation of C/EBPbeta and -delta was assessed in synovial tissues from patients with RA by electrophoretic mobility shift assay (EMSA); DNA binding activity of C/EBPs was evaluated by measuring EMSA band density. The expression and distribution of C/EBPbeta and -delta in synovial tissues were examined by immunohistochemistry analysis. As a control, synovial tissues from patients with osteoarthritis (OA) were studied. RESULTS: Enhanced DNA binding activity of C/EBPbeta and -delta, 2 major members of the C/EBP family, was detected in synovial tissues from RA patients, while synovial tissues from the patients with OA showed only faint or marginal activity (mean +/- SEM arbitrary units [AU] RA 23.3 +/- 11.7 in RA versus 4.5 +/- 1.3 in OA; P < 0.05). Moreover, the binding activities of the C/EBP proteins were correlated with both serum C-reactive protein levels (r = 0.62, P < 0.05) and synovial interleukin-6 messenger RNA levels (r = 0.60, P < 0.05). In immunohistochemistry studies, C/EBPbeta and -delta were detected predominantly in the rheumatoid synovial lining cells (both CD14+ and CD14- cells). CONCLUSION: C/EBPbeta and -delta may contribute to the pathology of rheumatoid synovitis.

Aged↗

PubMed2000

Perturbation of mitochondrial structure and function plays a central role in Actinobacillus actinomycetemcomitans leukotoxin-induced apoptosis.

Certain pore-forming bacterial toxins, including the leukotoxin (Ltx) produced by Actinobacillus actinomycetemcomitans, induce apoptosis in susceptible target cells. Although binding to the target cell surface represents the first step in the initiation of this process, the downstream events leading to toxin-induced apoptotic cell death have not been identified. Perturbation of mitochondrial function has been shown to have a major role in regulating progression of apoptosis initiated by exposure to numerous stimuli. Using Ltx as a model, the aim of this study was to evaluate whether induction of apoptosis by pore-forming toxins follows a similar paradigm. After exposure to Ltx, Epstein-Barr virus transformed B cells (JY cell line) exhibited the classical morphological features of apoptosis including decreased cell size, plasma membrane blebbing, selective alterations in plasma membrane permeability and condensation of nuclear DNA. The morphologic changes were accompanied by swelling of the mitochondria, a decrease in mitochondrial transmembrane potential (Psi(m)), hyperproduction of reactive oxygen intermediates (ROIs) and release of cytochrome c from the intermembrane space. Subsequently, we detected activation of the c ysteine asp artate-specific prote ases (caspases)-3 and -9, cleavage of the nuclear DNA repair enzyme, poly(ADP-ribose)polymerase (PARP) and internucleosomal DNA fragmentation. These results indicate that perturbation of mitochondrial structure and function, in concert with activation of specific caspases, initiate the effector phase of Ltx-induced apoptosis.

Aggregatibacter actinomycetemcomitans↗

PubMed2000

Characterization of the human nebulette gene: a polymorphism in an actin-binding motif is associated with nonfamilial idiopathic dilated cardiomyopathy.

Idiopathic dilated cardiomyopathy (IDC) is characterized by a thin-walled heart with systolic dysfunction of unknown etiology. Because abnormalities in genes for cytoskeletal proteins related to Z-disc function have recently been reported to cause IDC, genomic organization of the gene for nebulette, a novel actin-binding Z-disc protein, was determined and its sequence variations were searched for in Japanese patients with IDC and healthy controls. The nebulette gene consists of 28 exons, and four sequence variations leading to amino acid replacement (Gln187His, Met351Val, Asn654Lys, and Thr728Ala) were identified in the patients. These variations were also found in the healthy controls and hence they were polymorphisms and not disease-specific mutations. Frequencies of Gln187His, Met351Val, and Thr728Ala variants were similar in the patients and controls. However, the frequency of homozygotes for Lys at codon 654, a variant at a relatively conserved residue in an actinbinding motif, was significantly increased in nonfamilial IDC patients (n=106) as compared with healthy control subjects (n=331) (7.54% vs 1.21%, OR=6.25, P=0.002, 95% CI=1.92-20.29), while this association was not found in familial IDC patients (n=24). These observations suggest that the nebulette polymorphism in the actin-binding motif was a novel genetic marker of susceptibility to nonfamilial IDC.

Adolescent↗

PubMed2000

Identification of two novel mutations in the SLC25A13 gene and detection of seven mutations in 102 patients with adult-onset type II citrullinemia.

Adult-onset type II citrullinemia (CTLN2) is characterized by a liver-specific deficiency of argininosuccinate synthetase (ASS) protein. We have recently identified the gene responsible for CTLN2, viz., SLC25A13, which encodes a calcium-binding mitochondrial carrier protein, designated citrin, and found five mutations of the SLC25A13 gene in CTLN2 patients. In the present study, we have identified two novel mutations, 1800ins1 and R605X, in SLC25A13 mRNA and the SLC25A13 gene. Diagnostic analysis for the seven mutations in 103 CTLN2 patients diagnosed by biochemical and enzymatic studies has revealed that 102 patients had one or two of the seven mutations and 93 patients were homozygotes or compound heterozygotes. These results indicate that CTLN2 is caused by an abnormality in the SLC25A13 gene, and that our criteria for CTLN2 before DNA diagnosis are correct. Five of 22 patients from consanguineous unions have been shown to be compound heterozygotes, suggesting a high frequency of the mutated genes. The frequency of homozygotes is calculated to be more than 1 in 20,000 from carrier detection (6 in 400 individuals tested) in the Japanese population. We have detected no cross-reactive immune materials in the liver of CTLN2 patients with any of the seven mutations by Western blot analysis with anti-human citrin antibody. From these findings, we hypothesize that CTLN2 is caused by a complete deletion of citrin, although the mechanism of ASS deficiency is still unknown.

Adolescent↗

PubMed2000

Parental representation in eating disorder patients with suicide.

OBJECTIVE: We examined parental, personality, and symptomatological characteristics in relation to suicide attempts among eating disorder patients. METHODS: Fifty-one eating disorder inpatients, divided into two groups according to lifetime suicide attempts, and 107 non-psychiatric subjects were compared on the following variables: Parental Bonding Instrument (PBI), Global Clinical Score (GCS), Eating Disorder Inventory-91 (EDI-91), Eating Attitudes Test-26 (EAT), clinical and personality characteristics, and family backgrounds. RESULTS: Suicidal patients reported significantly higher overprotection by both parents than non-suicidal patients and non-psychiatric subjects. Suicidal patients had a more prevalent history of child abuse, affective instability, unstable self-image, avoidance of abandonment, maladaptive perfectionism, personality disorder, and mood disorder. There were no differences in symptomatological factors or the severity of the eating disorders. CONCLUSION: The results suggest that high overprotection is associated with suicidal behaviour in eating disorder patients. The association between overprotective parenting and personality characteristics, and methods of suicide prevention are discussed briefly.

Adolescent↗

PubMed2000

The healthy human infant tends to sleep in the prone rather than the supine position.

There are few reports about developmental behavior relating to roll over among healthy infants. We assessed the relationship between the placed position on sleeping and altered sleeping position the next morning by roll over among healthy infants. A health check-up clinic distributed a total of 1626 questionnaires to parents whose infant's ages are 1.5 years (or 18 months) old. The age at the first roll over and the change in sleeping position the next morning after they started to roll over, were investigated. The mean age of roll over from the supine to the prone among infants who were placed mainly in the prone sleeping position, at least in the first week of life, was 4.0 months (S.D., 1.1). The mean age of roll over from the supine to the prone among infants who were placed mainly in the supine sleeping position during early neonatal life and thereafter was 4.4 months (S.D., 1.2). The age of the first roll over from supine to prone was significantly younger in infants who were placed mainly in the prone sleeping position during early neonatal life. Among 651 infants who had been placed supine, 34.7% were found prone by roll over the next morning. Among 211 infants who had been placed prone, 14.2% were found supine by roll over the next morning. The number of infants who rolled over from supine to prone position was statistically greater than those from prone to supine. It is likely that the healthy human infant tends to sleep in the prone rather than the supine position. The finding is especially important for the correct assessment of the position in which SIDS cases were found dead.

Humans↗

PubMed2000

Liquid chromatographic determination of FP-21399 in plasma of patients with HIV infection.

A high-performance liquid chromatographic (HPLC) analysis method for the novel anti-HIV drug FP-21399 in human plasma was developed. The method employed the combination of organic solvent extraction and solid phase extraction. Analysis of FP 21399 and two major metabolites was achieved within 18 min using a reverse phase Puresil Cl 8 analytical column (4.6 x 150 mm, 5 microm, Waters) with a mobile phase of water-acetonitrile containing 20 mM triethylamine acetate (apparent pH 7.0). Linear gradient of mobile phase was applied as water-acetonitrile from 78:22 (v/v) to 55:45 over 8 min, and held at this ratio for the next 4 min. An ultraviolet-visible detector was operated at 265 mn from 0 to 8 min and at 600 mn from 8 min and after. The retention time of FP-21399 was 8.8 min and a linear response was observed over the concentration range 0.01 100 microg ml(-1) (r = 0.994). Lower limit of quantitation was found to be 0.01 microg ml(-1). Intra- and inter-assay precision varied in the range of 0.2 to 8% and 1-12%, respectively. The bias ranged from -17-3% for all analyses. A series of clinical plasma specimens were successfully analyzed using this method. The strategies for the method optimization on HIPLC separation and extraction procedure are discussed as well.

Anti-HIV Agents↗