Biomedical subjects

S Nagata

Publications and source records attributed to S Nagata.

At least 37 records · Page 2Linked to original sources

PubMed2001

Neural recognition molecule NB-2 of the contactin/F3 subgroup in rat: Specificity in neurite outgrowth-promoting activity and restricted expression in the brain regions.

NB-2, a neural cell recognition molecule of the contactin/F3 subgroup, promoted neurite outgrowth of the cerebral cortical neurons but not the hippocampal neurons. NB-2 in rat became apparent after birth at protein level, reaching a maximum at postnatal day 14 in the cerebrum and postnatal day 3 in the cerebellum. NB-2 in the cerebellum declined abruptly thereafter. In situ hybridization demonstrated that NB-2 mRNA was highly expressed in regions implicated in the central auditory pathway, including the cochlear nuclei, superior olive, inferior colliculi, medial geniculate nuclei, and auditory cortex. In addition, a high level of NB-2 expression was observed in the accessory olfactory bulb, thalamic nuclei, facial nucleus, and inferior olive. By immunohistochemistry, intense immunoreactivity against NB-2 was also detected in the auditory pathway. Thus, NB-2 is expressed in highly restricted brain regions, including the auditory system, suggesting that it plays specific roles in the development and/or maturation of the regions.

Aging↗

PubMed2001

Lowering the isoelectric point of the Fv portion of recombinant immunotoxins leads to decreased nonspecific animal toxicity without affecting antitumor activity.

Recombinant immunotoxins are genetically engineered proteins in which the Fv portion of an antibody is fused to a toxin. Our laboratory uses a 38-kDa form of Pseudomonas exotoxin A termed PE38 for this purpose. Clinical studies with immunotoxins targeting CD25 and CD22 have shown that dose-limiting side effects are attributable to liver damage and other inflammatory toxicities. We recently showed that mutating exposed surface neutral residues to acidic residues in the framework region of the Fv portion of an immunotoxin targeting CD25 [anti-Tac(scFv)-PE38] lowered its isoelectric point (pI) and decreased its toxicity in mice without impairing its cytotoxic or antitumor activities. We have now extended these studies and made mutations that change basic residues to neutral or acidic residues. Initially the pI of the mutant Fv (M1) of anti-Tac(scFv)-PE38 was decreased further. Subsequently, mutations were made in two other immunotoxins, SS1(dsFv)-PE38 targeting ovarian cancer and B3(dsFv)-PE38 targeting colon and breast cancers. We have found that all these mutant molecules fully retained specific target cell cytotoxicity and antitumor activity but were considerably less toxic to mice. Therefore, lowering the pI of the Fv may be a general approach to diminish the nonspecific toxicity of recombinant immunotoxins and other Fv fusion proteins without losing antitumor activity.

ADP Ribose Transferases↗

PubMed2001

Requirement of DNase II for definitive erythropoiesis in the mouse fetal liver.

Mature erythrocytes in mammals have no nuclei, although they differentiate from nucleated precursor cells. The mechanism by which enucleation occurs is not well understood. Here we show that deoxyribonuclease II (DNase II) is indispensable for definitive erythropoiesis in mouse fetal liver. No live DNase II-null mice were born, owing to severe anemia. When mutant fetal liver cells were transferred into lethally irradiated wild-type mice, mature red blood cells were generated from the mutant cells, suggesting that DNase II functions in a non-cell-autonomous manner. Histochemical analyses indicated that the critical cellular sources of DNase II are macrophages present at the site of definitive erythropoiesis in the fetal liver. Thus, DNase II in macrophages appears to be responsible for destroying the nuclear DNA expelled from erythroid precursor cells.

Animals↗

PubMed2001

Abdominal ultrasonography reveals the perforation site of duodenal ulcers.

Regardless of whether or not the laparoscopic approach is used, the simple closure using an omental patch is considered to be preferable to other surgical therapies for a duodenal ulcer perforation. We used abdominal ultrasonography (US) to identify the perforation site of the duodenal ulcer. To our knowledge, this is the first report describing how the perforation site of duodenal ulcers can be identified using US. Three patients diagnosed with perforated duodenal ulcers based on abdominal computed tomography and abdominal radiographic findings were scheduled for surgery. Under general anesthesia, US demonstrated free air on the liver and a "fish-eye sign" when the anterior or lateral wall was perforated. We performed a simple closure using an omental patch through a skin incision measuring only 3 cm in diameter above the area showing the fish-eye sign. The postoperative recovery was no different from that seen when a laparoscopic approach is used at our hospital. Even though the laparoscopic procedure is still the surgical modality of choice, the US technique allows for a successful diagnosis without invasive examinations. We believe that most surgeons and/or radiologists should try to detect the fish-eye sign using US as the diagnostic modality of first choice if they suspect a duodenal ulcer perforation. This technique allows us to mark the perforation site accurately before surgery.

Duodenal Ulcer↗

PubMed2001

Nociceptive stimulation increases NO synthase mRNA and vasopressin heteronuclearRNA in the rat paraventricular nucleus.

Nociceptive stimulation causes neuroendocrine responses such as arginine vasopressin (AVP) release and activation of the hypothalamo-pituitary-adrenal (HPA) axis. We examined the effects of nociceptive stimulation on the expression levels of neuronal nitric oxide synthase (nNOS) mRNA, heteronuclear (hn)RNA for AVP and AVP mRNA in the rat paraventricular nucleus (PVN) and supraoptic nucleus (SON), using in situ hybridization histochemistry. For nociceptive stimulation, formalin (5%) or saline was injected subcutaneously (s.c.) into the bilateral hind paws of rats. The expression of the nNOS gene in the PVN was significantly increased 2 and 6 h after s.c. injection of formalin in comparison with that in untreated and saline injected rats. The expression of the nNOS gene in the SON did not change in the untreated, saline- and formalin-injected rats. The AVP hnRNA in the PVN and SON was also significantly increased 15, 30 min and 2 h after s.c. injection of formalin, though AVP mRNA did not change at any time points that we studied. Plasma concentration of AVP was significantly increased 15 min after s.c. injection of formalin. These results suggest that NO in the PVN may be involved in nociceptive stimulation-induced neuroendocrine responses.

Animals↗

PubMed2001

Evaluation of an intestinal pressure-controlled colon delivery capsules prepared by a dipping method.

A new method for preparation of large amounts of empty pressure-controlled colon delivery capsules (PCDCs) by a dipping method has been developed. Empty PCDCs are composed of two polymer membranes. The inner one was a water-insoluble polymer membrane, ethylcellulose (EC). The outer one was an enteric polymer membrane, hydroxypropylmethylcellulose phthalate (HPMCP) or hydroxypropylmethylcellulose acetate succinate (HPMCAS). By consequently dipping into an ethanolic EC solution and an alkalized enteric polymer solution, empty PCDCs were obtained after both the capsule body and cap were adjusted to the size of #2 capsules. With each enteric polymer, two types of empty PCDCs of different thickness were prepared. Fluorescein (FL) was formulated with suppository base, PEG1000, and used as a model drug. FL/PEG1000 suspension was introduced into empty PCDCs which were then sealed with enteric polymer solution. The PCDCs were evaluated by an in vivo experiment using beagle dogs. After oral administration of the test PCDC preparations containing 30 mg of FL, blood samples were obtained from the jugular vein and serum FL levels were measured. The thickness of the EC membrane layer varied in both the capsule body and cap. HPMCAS PCDCs had 62.1+/-5.0 (S.E.) microm (body) and 49.7+/-3.3 microm (cap) with thicker ones and 55.7+/-6.6 microm (body) and 46.8+/-6.2 microm (cap) with thinner ones. HPMCP PCDCs had 28.1+/-3.3 microm (body), 30.9+/-1.0 microm (cap) with thinner ones and 43.1+/-9.8 microm (body), 42.4+/-8.2 microm (cap) with thicker ones. The mean T(i) values, the first appearance time, of FL in the serum of HPMCAS PCDCs were 2.0+/-0.7 h for thicker ones and 3.8+/-0.5 h for thinner ones, while the mean T(i) values of HPMCP PCDCs were 2.0+/-0.0 h for thinner ones and 3.5+/-0.7 h for thicker ones. Since the colon arrival time in beagle dogs was 3.5+/-0.3 h as determined by a sulfasalazine test, thinner HPMCAS PCDCs and thicker HPMCP PCDCs were thought to deliver FL to the colon.

Animals↗

PubMed2001

Enzymatic active site of caspase-activated DNase (CAD) and its inhibition by inhibitor of CAD.

Caspase-activated DNase (CAD) is a deoxyribonuclease that causes DNA fragmentation during apoptosis. In proliferating cells, CAD is complexed with ICAD (inhibitor of CAD) and its DNase activity is suppressed. Here, we established a quantitative assay for CAD DNase that measures the number of 3' hydroxyl groups on the CAD-generated DNA fragments. Chemical modification of histidine residues and substrate protection experiments demonstrated the presence of reactive histidine residues within the active site of the enzyme. Analysis by site-directed mutagenesis suggested that at least four histidine residues in the C-terminal part of the molecule are essential for the catalytic activity of CAD DNase. ICAD did not protect CAD from the chemical modification of the histidine residues, indicating that it does not mask the active site of CAD. In contrast, ICAD blocked the ability of CAD to bind DNA, suggesting that ICAD causes steric or electrostatic hindrance in CAD for substrate DNA. This molecular mechanism for the inhibition of CAD DNase by ICAD is similar to that proposed for colicin endonuclease and its inhibitor, immunity protein.

Amino Acid Sequence↗

PubMed2001

Testicular FasL is expressed by sperm cells.

The testis is the main source of Fas ligand (FasL) mRNA in rodents; it is generally believed that this molecule, expressed on bordering somatic Sertoli cells, bestows an immune-privileged status in the testis by eliminating infiltrating inflammatory Fas-bearing leukocytes. Our results demonstrate that the attribution of testicular expression of FasL to Sertoli cells is erroneous and that FasL transcription instead occurs in meiotic and postmeiotic germ cells, whereas the protein is only displayed on mature spermatozoa. These findings point to a significant role of the Fas system in the biology of mammalian reproduction.

Animals↗

PubMed2001

Frequent mutations of Fas gene in thyroid lymphoma.

Fas (Apo-1/CD95) is a cell-surface receptor involved in cell death signaling through binding of Fas ligand. Mutation of the Fas gene results in accumulation of lymphoid cells and thus might contribute to lymphomagenesis. Thyroid lymphoma (TL) is supposed to arise from active lymphoid cells formed in the preceding autoimmune chronic lymphocytic thyroiditis (CLTH). We examined the open reading frame of Fas cDNA in 11 cases of CLTH and 26 cases of TL. These patients were admitted to the hospital with varying degrees of goiter. All of the CLTH patients were female, with median age of 65 years, and all but five cases of TL were female, with median age of 61 years. Mutations of the Fas gene were detected in 3 (27.3%) of 11 cases of CLTH and 17 (65.4%) of 26 of TL. The Fas mutations comprised 18 frameshift, 3 missense, and 1 nonsense mutation. Frameshift mutations were caused by insertion of 1 bp (A) at nucleotide 1095 in 10 cases and by lack of exon 8 in 8 cases. The insertion of 1 bp (A) at nucleotide 1095 has never been reported in other kinds of malignancies. Thus, this might be unique in TL and CLTH and might be mutational hotspots in these diseases. All mutations occurred in the cytoplasmic region (death domain) known to be involved in the apoptotic signal transduction and thus could be loss-of-function mutations. These findings suggested that accumulation of lymphoid cells in CLTH with Fas mutation provides a basis for development of TL.

Aged↗

PubMed2001

Multiple variants of receptor-type protein tyrosine phosphatase beta are expressed in the central nervous system of Xenopus.

We have isolated cDNA clones encoding the Xenopus homologue of receptor-type protein tyrosine phosphatase beta (RPTPbeta), and identified 13 forms of the mRNA provably generated by alternative splicing. All the conceptual translates have a carbonic anhydrase-like domain, a fibronectin type III-like repeat and a spacer in the extracellular segment. Eleven of them (designated XRPTPbeta.1-XRPTPbeta.11) also have highly conserved two intracellular PTP domains, whereas the other two variants (sXRPTPbeta.1 and sXRPTPbeta.2) have neither transmembrane nor cytoplasmic segment. There are five peptides that can be inserted in various combinations into the spacer region. Northern and Western blot analyses show central nervous system-specific expression of the XRPTPbeta mRNAs and proteins. Chondroitinase ABC treatment of the brain and spinal cord extracts results in separation of six protein bands on the Western blot, in association with a decrease in the size of major bands, indicating that the major XRPTPbeta variants are chondroitin sulfate proteoglycans. The results of these as well as reverse-transcribed polymerase chain reaction analyses suggest that the amounts of different XRPTPbeta variants are regulated in tissue- and developmental stage-specific manners.

Amino Acid Sequence↗

PubMed2001

The membrane-bound but not the soluble form of human Fas ligand is responsible for its inflammatory activity.

The ectopic expression of Fas ligand (FasL/CD95L) in tissues or tumors induces neutrophil infiltration and the destruction of the tissues or the rejection of tumors. It has been suggested that the infiltrated neutrophils are responsible for the latter phenomena. FasL is synthesized as a type II transmembrane protein, and soluble FasL is produced by a proteolytic mechanism from the membrane-bound form. We previously demonstrated that uncleavable membrane-bound FasL of mice induces IL-1 beta release from inflammatory cells, and suggested that the IL-1 beta enhances neutrophil infiltration. However, recent papers reported that human soluble FasL is directly chemoattractive to neutrophils in vitro and proposed that the soluble form of FasL is responsible for its inflammatory activity. Therefore, in this report, we investigated which form is responsible for the inflammatory activities of human FasL. We produced tumor cell lines expressing one or both forms of human FasL. Cells expressing both forms or only the membrane-bound form of FasL induced neutrophil infiltration when transplanted into the peritoneal cavity of syngeneic mice, while cells expressing only the soluble form did not. Purified soluble FasL failed to induce neutrophil infiltration in vivo. IL-1 beta release from inflammatory peritoneal exudate and acceleration of tumor rejection were also mediated by membrane-bound but not soluble FasL. These results indicate that the membrane-bound form of FasL is primarily responsible for its inflammatory activity.

Animals↗

PubMed2001

Treatment parameters for selective occlusion of experimental corneal neovascularization by photodynamic therapy using a water soluble photosensitizer, ATX-S10(Na).

Time dependent change of an accumulation of an amphiphilic photosensitizer, ATX-S10(Na) on rabbit corneal neovascularization (CoNV) was evaluated by angiography using ATX-S10(Na) as a fluorescent dye on three rabbits. The angiography showed that the dye accumulated on CoNV 3-5 hr after dye injection when the dye in the iris was minimum. The results suggested 3-5 hr after might be the optimal time to start photodynamic therapy (PDT) to occlude CoNV selectively without damage to the surrounding normal tissue such as the iris. Then the optimal treatment parameters in PDT using ATX-S10(Na) for selective occlusion of the CoNV were investigated on rabbit eyes. PDT was performed with two different time intervals between dye injection and laser irradiation of a diode laser (670 nm), different laser doses and three different dye doses on 21 animals. PDT performed immediately after dye injection selectively occluded CoNV with laser irradiations from 30.6 to 38.2 J cm(-2)and a 2 mg kg(-1)dose of ATX-S10(Na), as well as with 15.3 J cm(-2)and a 6 mg kg(-1)dose. PDT performed 4 hr after dye injection with 107.0-152.8 J cm(-2)and a 6 mg kg(-1)dose, as well as with 38.2-53.5 J cm(-2)and a 12 mg kg(-1)dose was also effective. Although PDT performed either immediately or 4 hr after ATX-S10(Na) injection selectively occluded CoNV, the width of the optimal range of radiant exposures seemed wider in PDT performed 4 hr after dye injection. It is supposed that this result is associated with the difference of dye accumulation between in CoNV and in normal tissue as shown by the present angiographical findings.

Animals↗

PubMed2001

Action kinetics of a prothoracicostatic peptide from Bombyx mori and its possible signaling pathway.

A prothoracicostatic peptide (PTSP), purified from the brains of Bombyx mori, was found to inhibit ecdysteroidogenesis in the prothoracic glands (PGs) of this insect. This peptide was active at inhibiting ecdysteroidogenesis in the PGs at concentrations higher than 23 nM and glands incubated in 230 nM PTSP in vitro exhibited maximum inhibition of ecdysteroid production. By incubating PGs in vitro at different incubation periods it was observed that the first statistically significant inhibitory effect occurred after 30 min incubation in the presence of PTSP. Transferral of PGs from a medium with PTSP to a medium without PTSP resulted in the resumption of ecdysteroid production. Statistically significant inhibition of ecdysteroid production by PTSP was observed only in day 6 and in day 3 PGs of the 5th instar. The extracts of day 6 glands incubated in the presence of PTSP did not contain elevated amounts of ecdysteroid relative to controls after the incubations, indicating that PTSP does not inhibit the secretion, rather the synthesis, of ecdysteroid in the PGs. The presence of PTSP completely blocked the increased ecdysteroid production via L-type Ca(2+) channel activation by S(-)*Bay K 8644. There was no inhibition of ecdysteroid production by PTSP with glands incubated in Ca(2+)-free medium. The combined results suggest that PTSP regulates ecdysteroid synthesis only during specific stages of the 5th instar through a mechanism that likely involves the blocking of Ca(2+) influx through voltage-sensitive Ca(2+) channels in the PG cells of B. mori.

Animals↗

PubMed2001

Relationships between perceived workload, stress and oxidative DNA damage.

OBJECTIVES: The present study was performed to investigate the relationship between work-related factors, including psychological stress, and the formation of a type of oxidative DNA damage, 8-hydroxydeoxyguanosine (8-OH-dG), in order to examine their possible risk factor for occupational carcinogenesis. METHODS: A total of 54 healthy workers (27 male and 27 female, aged 41.2 +/- 12.5 years) in a company were investigated for 8-OH-dG levels in the peripheral blood leukocytes at the time of a questionnaire survey regarding several factors, such as working hours, workload, fatigue, sleep, psychological stress and the prospect of alleviating it. Subjects were limited to non-smoking and non-drinking workers to exclude the influence of cigarette smoking and alcohol drinking, which have been reported to have associations with the formation of 8-OH-dG. RESULTS: The levels of 8-OH-dG in female subjects were significantly related to the perceived workload (F = 5.56, P = 0.010), the perceived psychological stress (F = 6.15, P = 0.007), and the impossibility of alleviating stress (F = 3.82, P = 0.048). No associations were observed in male subjects. CONCLUSIONS: Psychological stress and perceived over-work appear to be related to the pathogenesis of cancer via the formation of 8-OH-dG, particularly in female workers.

8-Hydroxy-2'-Deoxyguanosine↗

PubMed2001

Can conditioned histamine release occur under urethane anesthesia in guinea pigs?

Many clinical and experimental data have shown that learning can occur under general anesthesia. To clarify this possibility with respect to allergic reactions, particularly asthmatic responses, we first established classical conditioned histamine release in response to a neutral odor by using pairings of the odor and an inhaled antigen for five sessions (Experiment 1) and then investigated whether conditioned histamine release into the plasma, bronchoalveolar lavage fluid (BALF), and lung tissue, which followed such a conditioning procedure, would be produced in urethane-anesthetized guinea pigs in the presence or absence of antigen (Experiment 2). Ovalbumin (OA) was used as the unconditioned stimulus (US) and dimethylsulfide (DMS) served as the conditioned stimulus (CS) in both experiments. In Experiment 1, the plasma histamine levels in the conditioned group increased significantly more than those of the unpaired control group in response to the CS during consciousness. In Experiment 2 in the absence of antigen, however, no significant differences in the histamine levels were found regarding the groups (DMS, triethylamine, saline, or unsensitized) or the time course (before, immediately, 5 min, and 10 min after the inhalations) during anesthesia, except for the finding that the histamine levels in the lung tissue specimens from the DMS group were significantly higher than those from the triethylamine group. In Experiment 2 in the presence of antigen, there was a significant increase in the plasma histamine levels after exposure to the US, irrespective of the presence of the CS, however, no significant difference in the histamine levels was observed between the US and the CS+US groups. These results indicated that a classically CS might not induce asthmatic responses under anesthesia.

Anesthesia, Intravenous↗

PubMed2001

[18F] labeled diacylglycerol analogue as a potential agent to trace myocardial phosphoinositide metabolism.

Phosphoinositide metabolism plays an important role in cardiac pathophysiology. To investigate whether [18F]diacylglycerol could be used to trace myocardial phosphoinositide metabolism, lipids were extracted from rat myocardium after the injection. 1-[8-[18F]fluorooctanoyl]-2-palmitoylglycerol and 1-[8-[18F]fluoropalmitoyl]-2-palmitoylglycerol were predominantly metabolized to phosphatidylethanolamine and triacylglycerol, respectively. The radioactivity incorporated into phosphoinositide metabolism was 51, 44, 32, and 30% 3, 5, 10, and 30 minutes after the injection of 1-[4-[18F]fluorobutyryl]-2-palmitoylglycerol, respectively. 1-[4-[18F]fluorobutyryl]-2-palmitoylglycerol might be a potential tracer to evaluate myocardial phosphoinositide metabolism early after the injection.

Animals↗

PubMed2001

Shigekazu Nagata.

Having originally researched the activities of the potent hematopeotic stimulator of bone-marrow cells, granulocyte colony stimulating factor, Shigekazu Nagata is better known for his work on apoptosis. Here, one of Japan's most renowned biomedical scientists outlines the path that has taken him full circle: from stimulating cells to grow, to finding out how they die, to tying the two processes together. (Interview by David Cyranoski.)

Apoptosis↗

PubMed2001

Fas gene mutations in prostatic intraepithelial neoplasia and concurrent carcinoma: analysis of laser capture microdissected specimens.

Fas (Apo-1/CD95) is a cell-surface receptor involved in cell death signaling through binding of Fas ligand. Mutations of the Fas gene might be involved in proliferative diseases of the prostate by prolongation of programmed cell death of prostatic epithelial cells. Using the laser capture microdissection method, Fas gene mutations were examined on genomic DNA extracted from lesions with high-grade prostatic intraepithelial neoplasia (HGPIN), a possible precursor of prostatic cancer (PCA), and from PCA. A total of 193 lesions, 111 with HGPIN, 55 with PCA, and 27 benign glands, were microdissected from 27 patients with PCA. Polymerase chain reaction-amplified products were directly sequenced. Loss of heterozygosity (LOH) was examined at four sites of known polymorphisms. Fas gene mutations were detected in HGPIN: 4 of 27 (14.8%) cases or 4 of 111 (3.6%) lesions. All were point mutations: three missense and one nonsense in the death domain. Benign proliferative glands adjoining HGPIN and/or PCA, and PCA never showed mutations. LOH was found in 31.3% of PCA and 25% of HGPIN lesions, but was never found in benign glands. Exclusive occurrence of Fas mutations in HGPIN might underlie the development of these lesions. Occasional findings of LOH in HGPIN and PCA suggested that genetic instability might occur during the early phase of prostatic carcinogenesis.

Alleles↗