Biomedical subjects

M Nakajima

Publications and source records attributed to M Nakajima.

At least 163 records · Page 9Linked to original sources

PubMed2000

The effects of sarin-like and soman-like organophosphorus agents on MAPK and JNK in rat brains.

One sarin-like and one soman-like organophosphorus agent [bis(isopropyl methyl)phosphonate, BIMP and bis(pinacolyl methyl)phosphonate, BPMP] were injected intravenously (iv) in rats. An increase in the tyrosine phosphorylation of several proteins in the cytosol fraction of the brain was observed. Activation of c-Jun N-terminal kinase (JNK) and slight activation of mitogen-activated protein kinase (MAPK) in the cytosol were also observed. The activation of these enzymes may be related to the high toxicity of these nerve agents.

Animals↗

PubMed2000

Genomic organization and chromosome localization of the newly identified human heparanase gene.

Heparanase (HPSE), which we have recently isolated, is an endo-beta-D-glucuronidase capable of cleaving heparan sulfate and has been implicated in inflammation and tumor angiogenesis and metastasis. In this report, the genomic organization and chromosome localization of the human heparanase gene is described. Polymerase chain reaction, subcloning and DNA sequencing analysis of a bacterial artificial chromosome (BAC) clone revealed that the 3.7 kb human heparanase cDNA is spread over about 50 kb and contains 14 exons and 13 introns. The heparanase gene is expressed as two mRNA species containing the same open reading frame, HPSE 1a (5 kb) (GenBank Data Library under accession number: AF155510); and HPSE 1b (1.7 kb) (GenBank Data Library under accession number: AF144325), generated by alternative splicing. The HPSE 1a-form contains all 14 exons, whereas in the HPSE 1b-form the first and fourteenth exons (5'- and 3'-untranslated region) have been spliced out. All splice sites conform to the GT-AG rule, except for the splice donor site of intron 13 (which is GA instead of GT), and the splice acceptor of intron 13 (which is GG instead of AG). Fluorescence in situ hybridization and radiation hybrid mapping suggest that the heparanase gene is located on human chromosome 4q22. This report regarding the structure of the human heparanase gene will aid in understanding the genetic contribution of this gene to normal physiology as well as to disease states. A possible involvement of heparanase in neuronal degeneration is discussed.

Alternative Splicing↗

PubMed2000

alpha-eudesmol, a P/Q-type Ca(2+) channel blocker, inhibits neurogenic vasodilation and extravasation following electrical stimulation of trigeminal ganglion.

In this study, we investigated the effect of alpha-eudesmol, which potently inhibits the presynaptic omega-agatoxin IVA-sensitive (P/Q-type) Ca(2+) channel, on neurogenic inflammation following electrical stimulation of rat trigeminal ganglion. Treatment with alpha-eudesmol (0.1-1 mg/kg. i.v.) dose-dependently attenuated neurogenic vasodilation in facial skin monitored by a laser Doppler flowmetry. In addition, alpha-eudesmol (1 mg/kg. i.v.) significantly decreased dural plasma extravasation in analysis using Evans blue as a plasma marker. On the other hand, alpha-eudesmol (1 mg/kg, i.v.) did not affect mean arterial blood pressure in rats. The calcitonin gene-related peptide (CGRP) and substance P (SP) released from activated sensory nerves have recently been suggested to be associated with the neurogenic inflammation. In this study, we also showed that alpha-eudesmol (0.45-45 microM) concentration-dependently inhibits the depolarization-evoked CGRP and SP release from sensory nerve terminals in spinal cord slices. These results indicate that the anti-neurogenic inflammation action of alpha-eudesmol, which does not affect the cardiovascular system, may be due to its presynaptic inhibition of the neuropeptide release from perivascular trigeminal terminals. We also suggest that the omega-agatoxin IVA-sensitive Ca(2+) channel blocker, alpha-eudesmol, may become useful for the treatment of the neurogenic inflammation in the trigemino-vascular system such as migraine.

Animals↗

PubMed2000

Endothelin receptor subtype antagonist activity of S-0139 in various isolated rabbit and canine arteries.

Vascular responses to endothelin peptides have been proposed to be mainly mediated via subtypes of the endothelin receptor, endothelin ET(A1), endothelin ET(B1), and endothelin ET(B2). The antagonist activity of 27-O-3-[2-(3-carboxy-acryloylamino)-5-hydroxyphenyl]acryloyloxy myricerone, sodium salt (S-0139) at these endothelin receptor subtypes was evaluated using isolated rabbit femoral, pulmonary, and mesenteric arteries. S-0139 competitively antagonized the endothelin-1-induced contraction mediated by the endothelin ET(A1) receptor in endothelium-denuded rabbit femoral arteries with a pA(2) value of 8.6+/-0.1. Endothelin ET(B2) receptor-mediated contraction induced by sarafotoxin S6c in endothelium-denuded rabbit pulmonary arteries was also inhibited by S-0139 with a pA(2) value of 5.6+/-0. 1. The pA(2) value of S-0139 for the endothelin ET(B1) receptor, evaluated from the endothelin-3-induced relaxant response in endothelium-intact rabbit mesenteric arteries, was 6.2+/-0.2. In isolated canine basilar, coronary, mesenteric and renal arteries, endothelin-1 caused concentration-dependent contractions with EC(50) values of 0.49+/-0.07, 0.61+/-0.25, 0.92+/-0.21 and 1.18+/-0.24 nM, respectively. S-0139 antagonized the endothelin-1-induced contraction in these arteries with pA(2) values of 8.0+/-0.1, 7. 6+/-0.2, 7.6+/-0.2 and 7.6+/-0.1, respectively. These results suggest that S-0139 is a potent and selective endothelin ET(A1) receptor antagonist, and that the contractions induced by endothelin-1 in canine basilar, coronary, mesenteric and renal arteries are mediated mainly via the endothelin ET(A1) receptor subtype.

Animals↗

PubMed2000

Activation of fibroblast-derived matrix metalloproteinase-2 by colon-cancer cells in non-contact Co-cultures.

Stromal fibroblasts interact with invading cancer cells by secreting and activating matrix metalloproteinases (MMPs). To elucidate the mechanisms involved in the expression and activation patterns of MMPs, human colon-cancer cell lines Caco-2 and LoVo and colon-fibroblast cell line CCD18-Co were co-cultivated in non-contact and contact conditions which mimic in vivo interaction between cancer cells and fibroblasts before and after cancer invasion respectively. Gelatin zymography disclosed that MMP-2 was secreted from the fibroblasts but not from the cancer cells. The quantity of fibroblast-derived MMP-2 in conditioned medium was not significantly changed in either the contact or the non-contact co-cultures when compared with that of individual cultures of CCD18-Co fibroblasts. Cancer cells in non-contact co-cultures, however, enhanced the activation of fibroblast-derived MMP-2. Transcripts of membrane-type matrix metalloproteinase-1 (MT1-MMP), which is thought to be present on the cell surface and to work as a candidate activator of MMP-2, were detected in both cancer cell lines. Plasma membrane extracts of cancer cells also activated MMP-2 in conditioned media in cell-free conditions. This activation of MMP-2 may be caused by MT1-MMP of the cancer cells, since it was inhibited by a series of MMP inhibitors, including ethylenediaminetetraacetic acid (EDTA), the tissue inhibitor of metalloproteinase-2 (TIMP-2), and the MMP inhibitor CGS 27023A, but not by TIMP-1. Our data demonstrate that in non-contact co-cultures colon-cancer cells activate fibroblast-derived MMP-2 on their plasma membranes. These findings should help to elucidate the mechanism involved in the initial destruction of basement membrane by cancer cells.

Basement Membrane↗

PubMed2000

Preparation of Monodispersed Solid Lipid Microspheres Using a Microchannel Emulsification Technique.

Monodispersed solid lipid microspheres consisting of high melting point edible oil were prepared as a novel sophisticated material. To prepare the monodispersed solid lipid microspheres a temperature-controlled microchannel (MC) emulsification process was devised. The prepared microspheres had diameters of approximately 20 µm or more and the standard deviation of the diameters was less than 1 µm. The effects of different surfactants, the pressure and the shape of the MC on the MC emulsification, and the diameter of the prepared droplet were studied. The experimental results are discussed using the new mechanism of droplets formation which is caused by the interfacial tension. Copyright 2000 Academic Press.

Journal Article↗

PubMed2000

Generation of 10,154 expressed sequence tags from a leafy gametophyte of a marine red alga, Porphyra yezoensis.

A total of 10,154 5'-end expressed sequence tags (EST) were established from the normalized and size-selected cDNA libraries of a marine red alga, Porphyra yezoensis. Among the ESTs, 2140 were unique species, and the remaining 8014 were grouped into 1127 species. Database search of the 3267 non-redundant ESTs by BLAST algorithm showed that the sequences of 1080 species (33.1%) have similarity to those of registered genes from various organisms including higher plants, mammals, yeasts, and cyanobacteria, while 2187 (66.9%) are novel. Codon usage analysis in the coding regions of 101 non-redundant EST groups showing significant similarity to known genes indicated the higher GC contents at the third position of codons (79.4%) than the first (62.2%) and the second position (45.0%), suggesting that the genome has been exposed to high GC pressure during evolution. The sequence data of individual ESTs are available at the web site http://www.kazusa.or.jp/en/plant/porphyra/EST/.

Algorithms↗

PubMed2000

Nitrotyrosine in esophageal squamous cell carcinoma and relevance to p53 expression.

We have examined the expression of nitrotyrosine, a marker of peroxynitrite formation, in 55 esophageal cancers by immunohistochemistry. Nitrotyrosine was detected in 21 of 55 (38.2%) esophageal cancers. Comparison of nitrotyrosine expression and the pathological findings showed that there was a significant association between the expression of nitrotyrosine and each of the depth of tumor invasion (P<0.01), occurrence of metastasis (P<0.05), pathological stage (P<0.01), smoking status (P<0.05) and alcohol intake (P<0.05). The survival rate of patients with nitrotyrosine-negative cancer was significantly higher than that of patients with nitrotyrosine-positive cancer (log-rank test, P<0.01). p53 was detected in 29 of 55 (52.7%) esophageal cancers, however, p53 expression did not correlate with nitrotyrosine expression. In conclusion, nitrotyrosine, a product of nitrogen species, is expressed in esophageal squamous cell carcinoma, which suggests that exogenous risk factors, such as tobacco and alcohol, through NO, are associated with carcinogenesis and progression of esophageal squamous cell carcinoma.

Adult↗

PubMed2000

Improved highly sensitive method for determination of nicotine and cotinine in human plasma by high-performance liquid chromatography.

A highly sensitive and reliable method for the determination of nicotine and its metabolite cotinine in human plasma by high-performance liquid chromatography was developed. Nicotine and cotinine were extracted from alkalinized plasma with dichloromethane and the volatility of nicotine was prevented by the addition of conc. HCl to the organic solvent during evaporation. The sensitivity of quantification at 260 nm absorption was improved by using a noise-base clean Uni-3 to 0.2 ng/ml nicotine and 1.0 ng/ml cotinine. The method was validated over linear ranges of 0.2-25.0 ng/ml for nicotine and 1.0-80.0 ng/ml for cotinine. The intra-day precision and accuracy were < or = 15.9% relative standard variation (RSD) and 89.9-103.5% for nicotine and < or = 8.0% RSD and 98.7-103.0% for cotinine. The inter-day precision and accuracy were < or = 17.0% RSD and 94.2-100.9% for nicotine and < or = 8.2% RSD and 98.0-105.1% for cotinine.

Calibration↗

PubMed2000

A forced-flow membrane reactor for transfructosylation using ceramic membrane.

A forced-flow membrane reactor system for transfructosylation was investigated using several ceramic membranes having different pore sizes. beta-Fructofuranosidase from Aspergillus niger ATCC 20611 was immobilized chemically to the inner surface of a ceramic membrane activated by a silane-coupling reagent. Sucrose solution was forced through the ceramic membrane by crossflow filtration while transfructosylation took place. The saccharide composition of the product, which was a mixture of fructooligosaccharides (FOS), was a function of the permeate flux, which was easily controlled by pressure. Using 0.2 micrometer pore size of symmetric ceramic membrane, the volumetric productivity obtained was 3.87 kg m(-3) s(-1), which was 560 times higher than that in a reported batch system, with a short residence time of 11 s. The half-life of the immobilized enzyme in the membrane was estimated to be 35 days by a long-term operation.

Aspergillus niger↗

PubMed2000

Secretory production of recombinant human chymase as an active form in Pichia pastoris.

We succeeded in expressing in a Pichia pastoris (P. pastoris) host a cDNA encoding a mature human chymase (h-chymase) which was secreted directly into the culture medium. Recombinant human heart chymase (rh-chymase) was purified from the culture medium via a single one-step heparin-agarose column chromatography tracing, using succinyl-Ala-Ala-Pro-Phe-para-nitroanilide (Suc-AAPF-pNA) hydrolysing activity. On SDS-polyacrylamide gel electrophoresis (SDS-PAGE), the rh-chymase showed a diffused protein band with molecular weight of 32-37 kDa. After deglycosylation, however, rh-chymase changed to a sharp protein band with molecular weight 28 kDa, which is equal in size to deglycosylated h-chymase. The rh-chymase had an activity to convert one of the natural substrates, angiotensin I, to angiotensin II. Double reciprocal plot analysis revealed that the K(m) value ofrh-chymase against Suc-AAPF-pNA was approximately 5.1 mM, which is close to that of purified h-chymase.

Angiotensin I↗

PubMed2000

Transesterification reaction between medium- and long-chain fatty acid triglycerides using surfactant-modified lipase.

Transesterification between medium-chain fatty acid triglycerides (MCT) and long-chain fatty acid triglycerides (LCT) in a nonsolvent system was investigated using surfactant modified lipase which is a complex of lipase, Rhizopus japonicus and surfactant, sorbitan monostearate. 74% conversion of was obtained after a 48-h reaction period, and the triglyceride composition was well described by the 1, 3-random 2-random stochastic model. The transesterification reaction between MCT and LCT closely followed the simple kinetic model, and the change in MCT and LCT contents could be simulated using one parameter. The effects of the water activity (A(w)) of modified lipase, the water content of the reaction system and the reaction temperature on the reaction rate were studied. A modified lipase A(w) of 0.35 and a water content of the reaction system at 0.09 wt % showed the highest activity. Inactivation did not occur below 60 degrees C, however, the activity decreased at temperatures over 70 degrees C.

Esterification↗

PubMed2000

Flexible synthesis and biological activity of uronic acid-type gem-diamine 1-N-iminosugars: a new family of glycosidase inhibitors.

An efficient and flexible synthetic route to four gem-diamine 1-N-iminosugars of uronic acid-type (D-glucuronic, D-mannuronic, L-iduronic, and L-guluronic acid), a new family of glycosidase inhibitor, from l-galactono-1,4-lactone have been developed in an enantiodivergent fashion through a sequence involving as the key steps (a) the formation of gem-diamine 1-N-iminopyranose ring by the Mitsunobu reaction of an aminal and (b) the introduction of a carboxylic acid group by the Wittig reaction of a ketone, hydroboration and oxidation, and the Sharpless oxidation. D-Glucuronic and D-mannuronic acid-type 1-N-iminosugars, (3S,4R,5R, 6R)- and (3S,4R,5R,6S)-4, 5-dihydroxy-6-trifluoroacetamido-3-piperidinecarboxylic acid, were proven to be potent inhibitors for beta-D-glucuronidase (IC(50) 6.5 x 10(-)(8)M) and to affect human heparanase (endo-beta-glucuronidase).

Animals↗

PubMed2000

Author reply

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Journal Article↗

PubMed2000

Analysis of a local fitness landscape with a model of the rough Mt. Fuji-type landscape: application to prolyl endopeptidase and thermolysin.

A method of analysis of a local fitness landscape for a current biopolymer is presented. Based on the assumption of additivity of mutational effects in the biopolymer, we assigned a site-fitness to each residue at each site. The assigned values of site-fitnesses were obtained by the least-squares method to minimize discrepancies between experimental fitnesses and theoretical ones. As test cases, we analyzed a section of a local landscape for the thermostability of prolyl endopeptidase and that for the enzymatic activity of thermolysin. These sections were proved to be of the rough Mt. Fuji-type with straight theta values of larger than 1.0, where straight theta is defined as the ratio of the "mean slope" to the "degree of roughness" on the fitness surface. Furthermore, we theoretically explained discrepancies between the fitnesses of multiple mutants and those predicted based on strict additivity of the component mutations by using a model of the rough Mt. Fuji-type landscape. According to this model, the discrepancies depend on the local landscape property (such as the straight theta value) and the location of the wild type on the landscape and the mean change in fitness by the component mutations. Our results suggest that this model may provide a good approximation of real sections of local landscapes for current biopolymers phenomenologically.

Amino Acid Sequence↗

PubMed2000

Onset of rigor mortis is earlier in red muscle than in white muscle.

Rigor mortis is thought to be related to falling ATP levels in muscles postmortem. We measured rigor mortis as tension determined isometrically in three rat leg muscles in liquid paraffin kept at 37 degrees C or 25 degrees C--two red muscles, red gastrocnemius (RG) and soleus (SO) and one white muscle, white gastrocnemius (WG). Onset, half and full rigor mortis occurred earlier in RG and SO than in WG both at 37 degrees C and at 25 degrees C even though RG and WG were portions of the same muscle. This suggests that rigor mortis directly reflects the postmortem intramuscular ATP level, which decreases more rapidly in red muscle than in white muscle after death. Rigor mortis was more retarded at 25 degrees C than at 37 degrees C in each type of muscle.

Adenosine Triphosphate↗

PubMed2000

Effect of intermittent administration of human parathyroid hormone (1-34) on the mandibular condyle of ovariectomized rats.

Intermittent administration of human parathyroid hormone (1-34) (PTH) increases bone mass in lumbar vertebrae and long bones of osteoporotic experimental animals. However, whether PTH has the same effect on jaw bones remains unclear. This study determined the effect of intermittent administration of PTH on rat mandibular condyle affected by estrogen deficiency. Fifty 6-month-old rats were either sham operated or ovariectomized, then divided into five groups depending on surgical procedure and hormone administration: sham + vehicle (SV), OVX + vehicle (OV), OVX + PTH 6 micrograms/kg once per week (OP6-1), OVX + PTH 60 micrograms/kg once per week (OP60-1), and OVX + PTH 20 micrograms/kg three times per week (OP20-3). PTH or vehicle was injected intermittently for 6 months in 5 rats of each group either immediately after surgery in a preventive administration experiment, or injected starting 6 months after surgery in a therapeutic administration experiment. The mandibles were excised, and bone morphometry was performed using confocal laser scanning microscopy and soft X-ray images. In both experiments, the bone volume of the OV groups was significantly lower than that of the SV group (P < 0.01); also, depending on dose and frequency, the bone volume of the OP group was higher than that of the OV group, particularly in the OP20-3 group. The value of mineralized surface of the OP groups was significantly higher than that of the OV group (P < 0.01), whereas the value of eroded surface of the OP groups was not significantly higher than that of the OV group. This study indicates that preventive and therapeutic intermittent administration of PTH in ovariectomized rats increase the bone formation in rat mandibular condyle without accelerating bone resorptive activity. This anabolic effect was best induced by the injection mode of 20 micrograms/kg three times per week.

Animals↗

PubMed2000

Comparison between indocyanine green angiography and outcome of surgical removal of choroidal neovascular membrane in age-related macular degeneration

Purpose: To review the outcome of surgical removal of choroidal neovascular membranes in age-related macular degeneration as classified by indocyanine green angiographic findings.Subjects and Method: Surgery was performed in 42 eyes. They were divided into four types by indocyanine green angiographic findings prior to surgery. Type I comprised 29 eyes showing hyperfluorescence throughout the angiographic phases. Type II comprised 3 eyes showing hyperfluorescence during the early phase only. Type III comprised 5 eyes showing hyperfluorescence in the late phase only. Type IV comprised 5 eyes without hyperfluorescence throughout the angiographic phases. The results were evaluated according to the visual acuity expressed as log MAR before and after surgery.Results: Visual acuity improved significantly in Types I, II, and III after surgery. Visual acuity did not improve in Type IV.Conclusion: The findings of indocyanine green angiography are thought to reflect the histological characteristics of the choroidal neovascular membrane. Neovascular membranes of Type IV may contain a smaller number of vessels and abundant fibrous tissue. Eyes of Type IV will have atrophies in the neurosensory retina, retinal pigment epithelium, and choriocapillaris. Surgical removal of the choroidal neovascular membrane in Type IV is not effective in improving visual acuity.

Journal Article↗