Biomedical subjects

K Nakashima

Publications and source records attributed to K Nakashima.

At least 199 records · Page 11Linked to original sources

PubMed1999

Liquid chromatography/mass spectrometric determination of trans-resveratrol in wine using a tandem solid-phase extraction method.

Liquid chromatography/mass spectrometry (LC/MS) with electrospray ionization has been successfully applied to the determination of trans-resveratrol (3,5,4'-trihydroxystilbene) in wine. Of a range of analytical conditions that were tested, optimum results were obtained by the use of reversed-phase high performance liquid chromatography (HPLC) using a mixture of methanol and ammonium acetate as the mobile phase. The negative-ion spectrum of trans-resveratrol showed pseudo-molecular ion, [M - H](-), which was the most abundant ion, and low fragment ions corresponding to the losses of hydroxyl groups of the phenol nucleus. Enhanced selectivity for the separation between trans-resveratrol and endogenous wine constituents was afforded by sample purification with a tandem solid-phase extraction method. The approach permits detection at low concentration of trans-resveratrol. The combination of improved sample pretreatment and an isocratic chromatographic system in conjunction with internal standardization forms the basis of a new assay for the quantitation of trans-resveratrol in wine. Full-scan mass spectra were readily obtained from 8 ng of trans-resveratrol, while a limit of detection of 200 pg (signal-to-noise ratio 3) was attained in the selected ion monitoring mode. The application of LC/MS to the determination of trans-resveratrol in wines is demonstrated by the analysis of red wines. Copyright 1999 John Wiley & Sons, Ltd.

Journal Article↗

PubMed1999

Simple and rapid high-performance liquid chromatography analysis of propentofylline and its main metabolites in serum using a direct injection technique.

A rapid, simple and reliable high-performance liquid chromatography (HPLC) column-switching method with UV detection (270 nm) for the simultaneous determination of propentofylline and its metabolites in human and rat sera was developed. The method involves direct injection of serum onto an HPLC column, which contains a shielded hydrophobic stationary phase for the separation of analytes from proteins in serum, and then loading the analytes onto a short octadecylsilylated silica gel (ODS) column using a switching valve. Propentofylline and its three metabolites in serum were separated from the serum components within 30 min after the injection. The detection limits (S/N = 3) of analytes spiked in human and rat sera ranged from 0.08 to 0.57 nmol/mL, and the net volume of serum used was 20 microL. The relative standard deviations for within- and between-day variations using rat serum were less than 4.3 and 5.6%, respectively. The method was used to determine propentofylline and its main metabolites in rat serum after a single intravenous dose of propentofylline (5 mg/kg).

Animals↗

PubMed1999

Enantiomer-specific high-performance liquid chromatography with fluorescence detection of methamphetamines in abusers' hair and urine.

Enantiomer-specific high-performance liquid chromatography with fluorescence detection using 4-(4,5-diphenyl-1H-imidazol-2-yl)-benzoyl chloride as a fluorescence labeling reagent was applied to determine methamphetamine and its metabolites in abusers' hair and urine. Hair samples were segmentally analyzed based on 1 cm long segments. In four hair samples, only the S(+)-enantiomers of methamphetamine and its N-demethylated metabolite, S(+)-amphetamine were detected. Satisfactory correlation (r = 0.901) between the results of high-performance liquid chromatography-fluorescence and those of gas chromatography-nitrogen phosphorous detection was obtained (n = 19). In an abuser's urine sample, the S(+)- and R(-)-enantiomers of methamphetamine, amphetamine and para-hydroxymethamphetamine were detected. The degree of N-demethylation of S(+)-methamphetamine into the corresponding metabolite of amphetamine was significantly higher than that of the R(-)-enantiomer.

Chromatography, Gas↗

PubMed1999

Combined effects of IL-12 and IL-18 on the clinical course and local cytokine production in murine pulmonary infection with Cryptococcus neoformans.

We reported recently that interleukin (IL)-12 and IL-18 synergistically increased the fungicidal activity of mouse peritoneal exudate cells against Cryptococcus neoformans by inducing the production of interferon (IFN)-gamma by natural killer (NK) cells. To confirm these findings in vivo, we examined the effect of combined treatment using these two cytokines on the course of experimentally induced pulmonary and disseminated cryptococcosis in mice. IL-12 and IL-18 were used at subtherapeutic doses (0.005 and 2 microg/mouse/day, respectively). A single administration of either cytokine was not effective in protecting mice against the infection, while combined treatment significantly prolonged survival time of infected mice and reduced the lung and brain loads of organisms. These protective effects were associated with elevated IFN-gamma and reduced IL-4 levels in bronchoalveolar lavage fluid. Finally, depletion of NK and gammadelta T cells, but not of CD4+ T cells, by administration of specific antibodies, significantly reduced the production of IFN-gamma in lungs by IL-12/IL-18 treatment during the 7 days of infection. Our results demonstrated that IL-12 and IL-18 protected mice against cryptococcal infection in a synergistic manner by enhancing the local production of IFN-gamma by NK and gammadelta T cells in the early phase of infection and by suppressing the production of IL-4 in lungs.

Animals↗

PubMed1999

cDNA cloning and developmental alterations in gene expression of the two Pit-1/GHF-1 transcription factors in the chicken pituitary.

Pit-1/GHF-1 (Pit-1) transcription factors promote the gene expressions for growth hormone (GH), prolactin (PRL), and the beta chain of thyroid-stimulating hormone in vertebrate pituitary glands. The present study analyzed the nature of chicken Pit-1s (cPit-1s) and their developmental expressions in the pituitary. Chicken pituitary expressed two cPit-1 mRNAs encoding cPit-1alpha and cPit-1gamma composed of 335 and 327 amino acid residues, respectively. They possessed different N-terminal regions and the common C-terminal regions containing a POU-specific domain and a POU homeodomain. Northern blot analysis revealed the pituitary-specific expressions of these Pit-1 mRNAs, and the Pit-1alpha mRNA expressions were two to three times higher than those for Pit-1gamma in both cephalic and caudal lobes of the pituitary. The cPit-1alpha and gamma mRNA expressions simultaneously increased after hatching until 4 weeks and then slightly decreased at 5 weeks. Similar gene expression profiles were observed for GH and PRL during the posthatch developmental period.

Amino Acid Sequence↗

PubMed1999

Astrocytic hyaline inclusions contain advanced glycation endproducts in familial amyotrophic lateral sclerosis with superoxide dismutase 1 gene mutation: immunohistochemical and immunoelectron microscopical analyses.

To clarify the neuropathological significance of the deposition of N(epsilon)-carboxymethyl lysine (CML), an advanced glycation endproduct, in astrocytic hyaline inclusions in familial amyotrophic lateral sclerosis (FALS), autopsy specimens from five members of two different families who had the superoxide dismutase 1 (SOD1) gene mutations were analysed. Immunohistochemically, most of the neuronal and astrocytic hyaline inclusions were intensely stained by the antibody against CML. The distributions and intensities of the immunoreactivities for CML and SOD1 were similar in the inclusions in both cell types. Immunoelectron microscopy showed that both inclusions consisted of CML-positive granule-coated fibrils and granular materials. No significant CML or SOD1 immunoreactivity was observed in the neurons and astrocytes of the normal control subjects. Our results suggest that astrocytic hyaline inclusions contain CML and SOD1 in FALS patients with SOD1 gene mutations, and that the formation of CML-modified protein (probably CML-modified SOD1) is related to the cell degeneration.

Adult↗

PubMed1999

Differential effects of troglitazone and D-chiroinositol on glucosamine-induced insulin resistance in vivo in rats.

Troglitazone and D-chiroinositol have been shown to exert antidiabetic effects by either potentiating or mimicking insulin action. We studied whether pretreatment with these compounds can prevent the deleterious effects of glucosamine on insulin action that may play an important role in hyperglycemia-induced insulin resistance. Normal Wistar rats were pretreated with troglitazone (100 mg/kg/d), D-chiroinositol (100 mg/kg/d), or placebo (saline) for 7 days. Glucosamine (50 micromol/kg/min) was then infused for 210 minutes, and a euglycemic glucose clamp was performed during the last 120 minutes. Pretreatment with troglitazone or D-chiroinositol had no effect on fasting plasma glucose or insulin or basal hepatic glucose output (HGO). Under the euglycemic-hyperinsulinemic (956+/-93 pmol/L) clamp condition, HGO in glucosamine-infused placebo-treated rats was not suppressed, but instead was increased over the basal level, indicative of hepatic insulin resistance. In contrast, HGO failed to increase during glucosamine infusion in rats pretreated with troglitazone but was not normally suppressed. This may indicate a partial improvement in the hepatic insulin resistance. D-Chiroinositol pretreatment had no effect on the glucosamine-induced increase in HGO. The glucose disposal rate (GDR) was 25% lower in rats infused with glucosamine versus saline-infused rats (25.5+/-2.5 v 34.1+/-2.0 mg/kg/min), indicative of peripheral insulin resistance. Pretreatment with D-chiroinositol (34.5+/-2.3 mg/kg/min) prevented the glucosamine-induced decrease in the GDR, indicating an improvement in peripheral insulin resistance. Troglitazone (25.2+/-3.3 mg/kg/min) was without effect. In conclusion, (1) in normal control rats, glucosamine infusion induced hepatic and peripheral insulin resistance; (2) D-chiroinositol, but not troglitazone, pretreatment prevented glucosamine-induced peripheral insulin resistance; and (3) troglitazone, but not D-chiroinositol, partially blocked the glucosamine-induced hepatic insulin resistance. D-Chiroinositol may provide a novel pharmacological approach to hexosamine-induced peripheral insulin resistance.

Animals↗

PubMed1999

Rabbit P300-like potential depends on cortical muscarinic receptor activation.

Novel stimuli trigger a positive brain potential P300 or P3a in mammalian animals, which is considered to represent an orienting response. Drugs systemically administered showed that acetylcholine is involved in the generation of P3. However, it is not known in which structure acetylcholine interacts with other neuronal substances for P3 generation. In this study, we injected scopolamine, an acetylcholine antagonist, into the parietal cortex. After the injection the amplitude of P3 from the parietal electrode decreased, while P3 recorded from neighbouring cortex was preserved. The peak latency of it did not show any significant change after the injection. P3 did not show any significant change after injecting normal saline into the same area. The parietal cortex is an essential structure for local P3 generation Different cerebral cortical regions respond to novel stimuli in parallel, and muscarinic receptor activation is necessary for processing the information of novelty detection.

Acoustic Stimulation↗

PubMed1999

Intradermal transfer of caspase-1 (CASP1) DNA into mouse dissects: role of CASP1 in interleukin-1beta associated skin inflammation and apoptotic cell death.

Caspase-1 (CASP1) interleukin-1beta (IL-1beta) converting enzyme (ICE) has been cloned as a specific enzyme which activates the biologically inactive pro-form of IL-1beta into biological active IL-1beta. Based on the significant homology to Ced-3, Caenorhabditis elegans apoptotic gene and, proof of apoptotic activity of ICE in rat fibroblasts, ICE was renamed as CASP1. In contrast to in vitro functions, the in vivo significance of high expression of CASP1 in skin remains to be elucidated. We transferred plasmid DNA encoding murine CASP1 with beta-actin promoter into mouse skin. The CASP1 DNA-injected skin, but not skin injected with control plasmid without CASP1, developed localized erythema with subcutaneous nodules. The nodules were associated with marked inflammatory infiltrates. The apoptotic cells detected by the TUNEL method were distributed in and around the inflammatory foci. The plasma IL-1beta level of CASP1 DNA-injected mouse was elevated compared with that of the control DNA-injected mouse. These inflammatory reactions of CASP1 DNA-injected skin were suppressed by treatment with neutralizing anti-murine IL-1beta antibodies, but the TUNEL positive apoptotic cells were still detected. This study clearly demonstrate dual roles of CASP1 in causing IL-1beta associated granulomatous skin infiltration and inducing apoptotic cell death in vivo.

Animals↗

PubMed1999

Rapid selection of complement-inhibiting protein variants in group A Streptococcus epidemic waves.

Serotype M1 group A Streptococcus strains cause epidemic waves of human infections long thought to be mono- or pauciclonal. The gene encoding an extracellular group A Streptococcus protein (streptococcal inhibitor of complement) that inhibits human complement was sequenced in 1,132 M1 strains recovered from population-based surveillance of infections in Canada, Finland and the United States. Epidemic waves are composed of strains expressing a remarkably heterogeneous array of variants of streptococcal inhibitor of complement that arise very rapidly by natural selection on mucosal surfaces. Thus, our results enhance the understanding of pathogen population dynamics in epidemic waves and infectious disease reemergence.

Animals↗

PubMed1999

HPLC with fluorescence detection of methamphetamine and amphetamine in segmentally analyzed human hair.

A sensitive high-performance liquid chromatographic method with fluorescence detection for determining methamphetamine and its major metabolite, amphetamine, in abusers' hair segments was developed. Methamphetamine and amphetamine in hair samples collected from addicts were extracted into acidified methanol, derivatized with 4-(4,5-diphenyl-1H-imidazol-2-yl)benzoyl chloride, separated isocratically on an ODS column using TRIS-HCl buffer (0.1 mol dm-3, pH 7.0)-methanol (30 + 70 v/v) as the mobile phase and the derivatives were detected fluorimetrically at 440 nm (lambda ex 330 nm). Calibration curves obtained by using control human hair spiked with standard solutions were linear (r > or = 0.999) up to at least 676.1 ng mg-1 for amphetamine and 746.1 ng mg-1 for methamphetamine. The detection limits at a signal-to-noise ratio of 3 were 51.4 and 74.6 pg mg-1 hair for amphetamine and methamphetamine, respectively. Using control hair spiked with standard solutions, the intra- and inter-day relative standard deviations (n = 5) were < or = 8.6% for both the target compounds. The method was successfully applied to the segmental analyses of methamphetamine abusers' hair samples.

Amphetamine↗

PubMed1999

Human dendritic cells express the thrombopoietin receptor, c-Mpl.

Human thrombopoietin (TPO) is a haemopoietic growth factor that is essential for the growth and development of megakaryocytes. However, c-Mpl, the TPO receptor, has been detected in human leukaemic cell lines with a myelomonocytic phenotype. These results raise the possibility that dendritic cells (DC), a putative myeloid lineage cell, may also express c-Mpl and respond to TPO. In haemopoietic stem cell transplantation, DC could induce graft-versus-host disease by its strong antigen-presenting capacity. In this study we have examined the effect of TPO on differentiation and the antigen-presenting capacity of DC. To differentiate DC, cord blood CD34+ cells were cultured in the presence of a cytokine cocktail either in serum-free medium or RPMI1640 containing 10% fetal bovine serum. Flow cytometric analysis and immunocytochemical staining demonstrated that c-Mpl was expressed on DC. Furthermore, the expression of c-Mpl mRNA was detected in DC by RNase protection assay. However, when TPO was added to the culture system there were no significant changes in the differentiation and mixed leucocyte-stimulating capacity of DC. These findings suggest that TPO may be administered following cord blood transplantation without significant augmentation of antigen presentation mediated by DC.

Cell Division↗

PubMed1999

Evaluation of the in-line filters for the intravenous infusion of amphotericin B fluid.

OBJECTIVE: To evaluate the effects of four types of in-line filters on filtration rate, amphotericin B concentration and particulate matter. METHODS: Filtration rates of amphotericin B fluid through in-line filters under maximum gravity flow were examined. The concentrations of amphotericin B and the particulate matter in the non-filtered and filtered amphotericin B fluid at the flow rate of 500 ml/24 h were measured. RESULTS: Filtration through a 1.2 microm or 0.2 microm polyethersulphone (PES) filter under maximum gravity flow took less than 40 min. The 0.2 microm positively charged nylon 66 and 0.2 microm nylon 66 filters took 70 and 100 min, respectively, to filter 500 ml of amphotericin B fluid. The 0.2 microm positively charged nylon 66 filter and the 0.2 microm nylon 66 filter, but not the PES filter (1.2 and 0.2 microm), decreased the concentrations of amphotericin B in the filtered fluid by 100% within 1 h and by 66% within 24 h after the start of filtration, respectively. The particulate count in the non-filtered amphotericin B fluid was 27+/-5 particles/ml, exceeding the limit defined by USP XXIII. The 1.2 microm and 0.2 microm PES filters significantly decreased particulate matter by 83 and 97%, respectively, just after filtration. CONCLUSION: The present results indicate that the 0.2 microm PES filter is optimal for intravenous infusion of amphotericin B fluid to minimize the introduction of particulate matter, microbial contaminants and endotoxin into patients.

Amphotericin B↗

PubMed1999

Positive association between an estrogen receptor gene polymorphism and Parkinson's disease with dementia.

Parkinson's disease (PD) is a common cause of dementia in the elderly. Dementia in Alzheimer's disease (AD) and PD share common biologic and clinical features. The estrogen receptor (ER) gene is one of the susceptibility genes for AD. In order to test the hypothesis that the overlap between AD and PD may have a genetic basis, we determined ER gene polymorphisms in 13 PD patients with dementia (PDD) (age +/- SD; 71.9 +/- 5.5 years), 71 PD patients without dementia (PDND) (68.4 +/- 7.5 years), 86 AD patients (76.8 +/- 8.0 years) and 51 control subjects (CTL) (74.9 +/- 6.9 years). ER genotypes were classified as a P or p allele on the basis of a Pvu II-RFLP, and X and x on the basis of a Xba I-RFLP. The frequency of the P allele in the PDD group as well as the AD group was higher than that in CTL. There was no significant difference in the distribution of the P allele between CTL and PDND. There were no significant differences in the distribution of the X allele among the PDD, PDND and CTL groups, whereas a higher incidence was found in AD. We conclude that the ER gene may be a common susceptibility gene for dementia in PD as well as AD.

Aged↗

PubMed1999

Low interhemispheric and high intrahemispheric EEG coherence in migraine.

Twenty women with migraine were evaluated by EEG coherence analysis. In comparison with controls, the migrainous subjects showed lower interhemispheric coherence values at C3-C4 for the delta band (P=0.0030) and at F3-F4 (P=0.0055) and C3-C4 (P=0.0018) for the beta band frequency (Wilcoxon criterion). In contrast, 22 intrahemispheric coherence pairs were significantly higher in the migraine group than in the control group (P<0.01). The subjects with visual aura showed lower interhemispheric coherence at O1-O2 and T5-T6 for the alpha band than the subjects without the aura. Discriminant analysis revealed that 95% of the subjects with migraine and 85% of the controls were correctly classified into their proper groups. We suggest that EEG coherence analysis is a sensitive parameter in the detection of electrophysiological abnormalities in patients with migraine.

Adolescent↗

PubMed1999

Transcriptional regulation of the CDK inhibitor p16INK4a gene by a novel pRb-associated repressor, RBAR1.

p16, also known as INK4a, CDKN2, or MTS1, plays an important role in the control of the cell cycle progression, and retinoblastoma protein (pRb) is suggested to be involved in the transcriptional regulation of p16. However, it is not fully understood how pRb regulates transcription of the p16 gene. Nuclear proteins prepared only from the pRb-nonfunctional human tumor cells were found to bind to the 5'-flanking sequence of the p16 gene in the presence of Zn2+ by electrophoretic mobility shift assay (EMSA). EMSA using mutagenized 5'-flanking sequences as competitors suggested that the sequence at position -97 to -87 relative to first ATG of the p16 gene was critical for protein binding. Transient reporter assay indicated that the sequence identified by EMSA acted as a silencer element in the pRb-nonfunctional tumor cells, showing the presence of a transcriptional repressor associated with functional pRb (RBAR1).

Cyclin-Dependent Kinase Inhibitor p16↗

PubMed1999

Temporal and spatial profile of apoptotic cell death in transient intracerebral mass lesion of the rat.

Apoptosis is involved in the pathogenesis of cerebral ischemia. Previous studies have confirmed that the brain surrounding an intracerebral hematoma develops ischemia. We investigated the number and distribution of cells exhibiting DNA fragmentation with apoptotic morphology in the transient intracerebral mass lesion to determine whether apoptosis contributed to the lesion progress after intracerebral hemorrhage (ICH). Transient intracerebral mass was created by inflation of a microballoon for 10 min (group A) or 2 h (group B) in the caudoputamen in rats, and brains were examined 1, 3, 6, 24, and 48 h after microballoon deflation. The lesion volume was calculated using parallel coronal sections with cresyl violet staining. Terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine (dUTP)-biotin nick end labeling (TUNEL) was used to detect cells undergoing DNA fragmentation. Immunohistochemistry for Fas antigen was also done to ascertain molecular mechanisms of apoptosis. Histological examination of hematoxylin and eosin-stained sections showed the typical appearance of neuronal necrosis in the caudoputaminal lesion. Lesion volume in the caudoputamen gradually increased as time advanced from 1 to 48 h. Cells stained heavily by TUNEL with apoptotic morphology were detected in the lesion, but not in the inner boundary zone of the lesion. The number of these cells significantly increased from 6 to 24 h in each experimental group (p < 0.05). The cells with positive immunoreactivity for Fas antigen was prominently observed in the lesion at 6 h. The distribution of apoptotic cells and the rapid increase in the number of apoptotic cells after 24 h propose that apoptotic cell death may contribute to lesion core formation but not to gradual development of the lesion.

Analysis of Variance↗

PubMed1999

Anti-elongation factor-1alpha autoantibody in adult atopic dermatitis patients.

Adult atopic dermatitis (AD) patients develop severe facial lesions, which sometimes distribute in sun-exposed areas similar to the rash of systemic lupus erythematosus. To declare autoimmunity in the pathogenesis of AD, we investigated serum antinuclear antibody (ANA) in 256 adult AD patients and identified its ligands. A high titer of ANA was found in 31.3% of AD patients and 75% of the ANA showed a homogenous pattern. Sixty-five percent of ANA(+) sera reacted to a 52 kDa protein (p52) in HeLa cell immunoblots. By screening the HeLa cell cDNA expression library with anti-p52 sera, a clearly positive clone was isolated. The sequence of this cDNA was identical to human elongation factor (hEF)-1alpha. The eluate of IgG bound to hEF-1alpha-glutathione S-transferase (GST) fusion protein recognized a band at 52 kDa in a HeLa cell immunoblot, and stained Hep-2 cell nuclei and cytoplasma as reported in hEF-1alpha distribution. The anti-p52 AD sera recognized the hEF-1alpha-GST fusion protein. The anti-hEF-1alpha antibody-positive AD patients were characterized by higher facial involvement and lower white blood cell counts compared with antibody-negative patients. The present results suggest the possible involvement of autoimmunity in the pathogenesis of adult AD.

Adult↗