Biomedical subjects

N Murata

Publications and source records attributed to N Murata.

At least 55 records · Page 3Linked to original sources

PubMed2000

Quantitative measurement of sphingosine 1-phosphate by radioreceptor-binding assay.

In Chinese hamster ovary cells overexpressing Edg-1, one of the sphingosine 1-phosphate (S1P) receptor subtypes, [(3)H]S1P binding was displaced by unlabeled S1P with IC(50), a half-maximal concentration to inhibit the binding, of about 20 nM. This radioreceptor binding was used for quantitative measurement of S1P. Among the various lipids employed, only sphingosylphosphorylcholine (SPC), other than S1P, practically displaced the binding; however, the potency of SPC was about 100 to 1000 times less than that of S1P. Thus, SPC bound to the S1P receptors inefficiently. Furthermore, before the application of test samples to this assay, S1P was partially purified: the lipid was extracted first into the aqueous phase and separated from other lipids under alkaline conditions, and then reextracted into the chloroform phase under acidic conditions. With this assay, we could specifically and quantitatively measure S1P from 2 to 40 pmol per assay well in biological samples including serum samples and various tissues. This assay also allowed us to measure the change in cellular S1P content in U937 cells after treatment with exogenous sphingosine.

Animals↗

PubMed2000

Inhibition by 9alpha-fluoromedoroxyprogesterone acetate (FMPA) against mammary carcinoma induced by dimethylbenz[a]anthracene in rats and angiogenesis in the rabbit cornea - comparison with medroxyprogesterone acetate (MPA).

Medroxyprogesterone acetate (MPA) is currently used therapeutically in the treatment of mammary and endometrial carcinomas. In order to develop a more potent and useful drug, we synthesized the novel compound, 9alpha-fluoromedoroxyprogesterone acetate (FMPA), by fluorinating MPA, and we also previously reported that FMPA displays more potent anti-angiogenic activity in the chorioallantoic membrane assay than MPA. In the present study, we investigated (1) the effects of FMPA on rat mammary carcinomas induced by dimethylbenz[a]anthracene (DMBA) to determine the anti-tumor activity, (2) the effect on angiogenesis in rabbit corneal assays, and (3) compared these results with those for MPA. FMPA inhibited the growth of mammary carcinomas in a dose-dependent manner (7.5, 30 and 120 mg/kg). Almost complete involution of the carcinomas was observed at doses of 30 and 120 mg/kg. MPA also inhibited the growth of carcinomas at doses of 30 and 120 mg/kg, but no involution of carcinomas was observed even at 120 mg/kg. FMPA significantly and MPA to a lesser degree inhibited carcinogenesis at 120 mg/kg within their treatments. In rabbit corneal assays, FMPA significantly inhibited angiogenesis (IC50 value=0.085 microg/pellet). MPA also significantly inhibited angiogenesis (IC50 value=0.60 microg/pellet). From these results, we conclude that FMPA is potentially more effective in the treatment of mammary carcinomas than MPA.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed2000

Absence of p16, p21 and p53 gene alterations in hepatocellular carcinomas induced by N-nitrosodiethylamine or a choline-deficient L-amino acid-defined diet in rats.

To clarify the involvement of tumor suppressor genes in exogenous and endogenous liver carcinogenesis, alterations of p16, p21 and p53 in hepatocellular carcinomas (HCCs) induced by N-nitrosodiethylamine (DEN) and a choline deficient L-amino acid-defined (CDAA) diet in rats were investigated. Male Fischer 344 rats received DEN at 6-week of age followed by partial hepatectomy (PH), with colchicine to induce cell cycle disturbance, and a selection pressure regimen. Sacrifice was after 42 weeks. Other animals continuously received a CDAA diet for 75 weeks and were then killed. Eleven and 15 HCCs were obtained, respectively. Total RNA was extracted from and cDNA was synthesized with reverse transcriptase to allow investigation of mutations in p16, p21 and p53 by polymerase chain reaction single strand conformation polymorphism (PCR-SSCP) analysis. Expression of p16 and p21 mRNA was also analyzed by reverse transcription (RT)-PCR. The results showed no mutations or deletions of p16, p21 and p53 in any of the HCCs induced by DEN or CDAA. Loss or decrease of p16 and p21 expression were also not found, suggesting that p16, p21 and p53 alteration may not be necessary for either exogenous or endogenous liver carcinogenesis in rats.

Animals↗

PubMed2000

Photosystem II fluorescence quenching in the cyanobacterium Synechocystis PCC 6803: involvement of two different mechanisms.

The structural changes associated to non-photochemical quenching in cyanobacteria is still a matter of discussion. The role of phycobilisome and/or photosystem mobility in this mechanism is a point of interest to be elucidated. Changes in photosystem II fluorescence induced by different quality of illumination (state transitions) or by strong light were characterized at different temperatures in wild-type and mutant cells, that lacked polyunsaturated fatty acids, of the cyanobacterium Synechocystis PCC 6803. The amplitude and the rate of state transitions decreased by lowering temperature in both strains. Our results support the hypothesis that a movement of membrane complexes and/or changes in the oligomerization state of these complexes are involved in the mechanism of state transitions. The quenching induced by strong blue light which was not associated to D1 damage and photoinhibition, did not depend on temperature or on the membrane state. Thus, the mechanism involved in the formation of this type of quenching seems to be unrelated to the movement of membrane complexes. Our results strongly support the idea that the mechanism involved in the fluorescence quenching induced by light 2 is different from that involved in strong blue light induced quenching.

Cyanobacteria↗

PubMed2000

The pathway for perception and transduction of low-temperature signals in Synechocystis.

Low temperature is an important environmental factor that has effects on all living organisms. Various low-temperature-inducible genes encode products that are essential for acclimation to low temperature, but low-temperature sensors and signal transducers have not been identified. However, systematic disruption of putative genes for histidine kinases and random mutagenesis of almost all the genes in the genome of the cyanobacterium Synechocystis sp. PCC 6803 have allowed us to identify two histidine kinases and a response regulator as components of the pathway for perception and transduction of low-temperature signals. Inactivation, by targeted mutagenesis, of the gene for each of the two histidine kinases and inactivation of the gene for the response regulator depressed the transcription of several lowtemperature-inducible genes.

Bacterial Proteins↗

PubMed2000

yam8(+), a Schizosaccharomyces pombe gene, is a potential homologue of the Saccharomyces cerevisiae MID1 gene encoding a stretch-activated Ca(2+)-permeable channel.

The Saccharomyces cerevisiae MID1 gene encodes a stretch-activated Ca(2+)-permeable channel. In a protein database, we found a Schizosaccharomyces pombe gene whose predicted protein shows 26% identical and 62% similar to the Mid1 channel in amino acid sequence. cDNA derived from this gene, designated yam8(+), was isolated by reverse transcription-polymerase chain reaction (RT-PCR). Further analysis showed that the Yam8 protein consists of 486 amino acids and has 6 hydrophobic segments. The yam8(+) cDNA, placed under the S. cerevisiae TDH3 promoter, partially complemented the mating pheromone-induced death (mid) phenotype of the S. cerevisiae mid1 mutant. The expression of the yam8(+) cDNA in the mid1 mutant cells partially remediated the mid phenotype and resulted in a slight increase in Ca(2+) uptake activity. These findings suggest that Yam8 is a potential homologue of Mid1.

Amino Acid Sequence↗

PubMed2000

Molecular cloning and characterization of a rice dehydroascorbate reductase.

Plant dehydroascorbate reductase (DHAR), which re-reduces oxidized ascorbate to maintain an appropriate level of ascorbate, is very important, but no gene or cDNA for plant DHAR has been cloned yet. Here, we describe a cDNA for a rice glutathione-dependent DHAR (designated DHAR1). A recombinant Dhar1p produced in Escherichia coli was functional. The expression sequence tag database suggests that Dhar1p homologs exist in various plants. Furthermore, the rice Dhar1p has a low similarity to rat DHAR, although the rice enzyme has a considerably higher specific activity than the mammalian one. The mRNA level of DHAR1, the protein level of Dhar1p and the DHAR activity in rice seedlings were elevated by high temperature, suggesting the protection role of DHAR at high temperature.

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

Tumour necrosis factor alpha (TNF-alpha) interferes with Fas-mediated apoptotic cell death on rheumatoid arthritis (RA) synovial cells: a possible mechanism of rheumatoid synovial hyperplasia and a clinical benefit of anti-TNF-alpha therapy for RA.

To investigate the mechanism of rheumatoid synovial hyperplasia (RASH), the influence of tumour necrosis factor alpha (TNF-alpha) on Fas-mediated apoptotic cell death (Fas-ACD) was examined on cultured rheumatoid synovial cells (RASCs). RASCs were obtained from the synovial tissues of eight patients with rheumatoid arthritis (RA) and SCs from eight patients with osteoarthritis (OA) were used as a control. To examine the influence of TNF-alpha on Fas-ACD, SCs were cultured with anti-Fas antibody (CH11) for 16 h in the absence or presence of different doses of recombinant TNF-alpha. ACD was determined by electron microscopic analysis and the percentage of apoptotic cells was calculated by trypan blue staining. In addition, the expression of Fas and Bcl-2 on RASCs was examined by flow cytometry. As a result, RASCs were more susceptible to Fas-ACD in vitro than OASCs. TNF-alpha interfered with Fas-ACD on RASCs in a dose-dependent manner. Moreover, removal of TNF-alpha activity by a neutralizing anti-TNF-alpha antibody (cA2) restored Fas-ACD. Flow cytometric analysis showed no significant changes in either Fas or Bcl-2 expression on RASCs after the culture with TNFalpha. These results suggest the following: (1) Fas-ACD might be diminished in vivo by local excessive TNF-alpha and this might contribute in part to RASH. (2) The inhibition of Fas-ACD on RASCs by TNF-alpha might not be associated with changes in the expression of Fas or Bcl-2. (3) In addition, considering a magnetic resonance imaging (MRI) finding of marked reduction in the RASH after cA2 treatment, blockade of TNF-alpha activity could restore Fas-ACD in RA synovium, implicating a clinical benefit of anti-TNF-alpha therapy for RA.

Antibodies, Monoclonal↗

PubMed2000

The facilitation of peritoneal dissemination of a tumor by laparotomy in a rabbit model.

The effects of conventional laparotomy and laparoscopic surgery on the peritoneal dissemination of VX2 tumors were evaluated in a rabbit model. Either 5 x 10(3) or 5 x 10(4) of VX2 tumor cells were inoculated into the peritoneal cavities of rabbits which were divided into three groups according to the procedure performed, namely, a conventional laparotomy group (group 1), a laparoscopy group (group 2), and a control group (group 3). All the animals were killed 2 weeks after the treatment to examine the spread of the VX2 tumor in the omentum. In the experiment in which 5 x 10(3) tumor cells were inoculated, no significant difference was found in the number of metastases among the three groups. Conversely, after inoculating 5 x 10(4) tumor cells, the mean numbers of metastatic colonies greater than 1 mm in diameter in the greater omentum were 14.4+/-4.7 in group 1, 9.9+/-2.4 in group 2, and 3.3+/-1.1 in group 3, with a significant difference between groups 1 and 3, and between groups 2 and 3, but not between groups 1 and 2. In conclusion, conventional laparotomy facilitated the greatest peritoneal dissemination of a VX2 tumor in this animal model when 5 x 10(4) tumor cells were inoculated. Furthermore, laparoscopic surgery influences tumor growth to almost the same degree as conventional laparotomy.

Animals↗

PubMed2000

Effect of CO(2) pneumoperitoneum on growth of liver micrometastases in a rabbit model.

Little is known about the risk of metachronous liver metastases following laparoscopic resection for gastrointestinal malignancies. The effect of CO(2) pneumoperitoneum on the growth of established liver micrometastases was investigated in a rabbit model. Male Japanese white rabbits weighing 2.8 to 3.3 kg were randomized to three groups (n = 15 per group) 3 days following intraportal inoculation of a tumor suspension containing 5 x 10(4) cells of VX(2) cancer. In the pneumoperitoneum group, insufflation with CO(2) was maintained at a pressure of 10 mmHg for 30 minutes. In the laparotomy group the abdominal cavity remained open through a 45 mm midline incision for 30 minutes; in the control group no treatment other than anesthesia was performed. Cancer nodules on the liver surface were compared among the three groups on day 17. There was no difference in the number of cancer nodules among the groups (p = 0. 72). A significant difference in the total area of cancer nodules (mean +/- SEM) was found only between the pneumoperitoneum group (696.0 +/- 177.0 mm(2)) and the control group (247.2 +/- 60.7 mm(2)) (p < 0.05). The frequency of cancer nodules larger than 3.0 mm in maximal diameter tended to be highest in the pneumoperitoneum group (p = 0.053). These results suggests that CO(2) pneumoperitoneum may promote the growth of established liver micrometastases in this animal model.

Animals↗

PubMed2000

Identification of 142 single nucleotide polymorphisms in 41 candidate genes for rheumatoid arthritis in the Japanese population.

Single-nucleotide polymorphisms (SNPs) can make an important contribution to our understanding of genetic backgrounds that may influence medical conditions and ethnic diversity. We undertook a systematic survey of genomic DNA for SNPs located not only in coding sequences but also in non-coding regions (e.g., introns and 5' flanking regions) of selected genes. Using DNA samples from 48 Japanese patients with rheumatoid arthritis (RA) as templates, we surveyed 41 genes that represent candidates for RA, screening a total of 104 kb of DNA (30 kb of coding sequences and 74 kb of non-coding DNA). Within this 104 kb of genomic sequences we identified 163 polymorphisms (1 per 638 bases on average), of which 142 were single-nucleotide substitutions and the remainder, insertions or deletions. Of the coding SNPs, 52% were non-synonymous substitutions, and non-conservative amino acid changes were observed in a quarter of those. Sixty-nine polymorphisms showed high frequencies for minor alleles (more than 15%) and 20 revealed low frequencies (<5%). Our results indicated a greater average distance between SNPs than others have reported, but this disparity may reflect the type of genes surveyed and/or the relative ethnic homogeneity of our test population.

Alleles↗

PubMed2000

Pneumoperitoneum with carbon dioxide enhances liver metastases of cancer cells implanted into the portal vein in rabbits.

BACKGROUND: Little is known about the role of the CO2 pneumoperitoneum on tumor cells that spread from the portal system into the liver during laparoscopic surgery for gastrointestinal malignancies. Therefore, we designed a study to investigate the effect of CO2 pneumoperitoneum on cancer cells implanted in the portal vein in a rabbit model. METHODS: Immediately after intraportal inoculation of 2.5x10(5) cells of VX2 cancer, the rabbits received either CO2 pneumoperitoneum at a pressure of 10 mm Hg for 30 min (pneumoperitoneum group, n = 14) or laparotomy alone for 30 min (laparotomy group, n = 14). RESULTS: The number (p<0.01) and area of cancer nodules (p = 0.045) on the liver surface on day 17 were greater in the pneumoperitoneum group than in the laparotomy group. The frequency of cancer nodules >3.0 mm in diameter was higher in the pneumoperitoneum group than in the laparotomy group (p<0.001). CONCLUSIONS: Compared with laparotomy, CO2 pneumoperitoneum enhanced the development of liver metastases in this experimental model.

Animals↗

PubMed2000

The influence of different insufflation pressures during carbon dioxide pneumoperitoneum on the development of pulmonary metastasis in a mouse model.

BACKGROUND: The effects of different insufflation pressures on the development of pulmonary metastasis was investigated in a mouse laparoscopy model. METHODS: BALB/C mice intravenously inoculated with colon 26 cells were randomized to one of five treatment groups (10 mice per group): pneumoperitoneum at different pressures of 5, 10 or 15 mmHg; full laparotomy for 60 min; or anesthesia control. Cancer nodules on the lung surface 19 days postoperatively were compared between groups. RESULTS: (a) As compared with the control group, pneumoperitoneum at 10 and 15 mmHg and laparotomy enhanced the growth of pulmonary metastases (p < 0.01). (b) The growth of metastases also was greater in laparotomy group mice than in mice undergoing pneumoperitoneum at 5 and 10 mmHg (p < 0.05). CONCLUSIONS: These results suggest that the effects of different insufflation pressures on the growth of pulmonary metastases are not identical, and that pneumoperitoneum with high pressure may promote pulmonary metastases similar to those with laparotomy.

Analysis of Variance↗

PubMed2000

Influence of trocar placement and CO(2) pneumoperitoneum on port site metastasis following laparoscopic tumor surgery.

BACKGROUND: The mechanisms involved in the development of port site metastasis following laparoscopic tumor surgery remain controversial. Therefore, we decided to investigate the influence of trocar placement and CO(2) pneumoperitoneum on abdominal wound implantation in relation to intraperitoneal tumor growth in a rabbit model. METHODS: Rabbits received either CO(2) pneumoperitoneum with insertion of nine trocars (pneumoperitoneum group, n = 15), insertion of nine trocars alone (nonpneumoperitoneum group, n = 15), or nine abdominal incisions (control group, n = 13) 3 days after intraperitoneal inoculation of VX(2) cancer cells. RESULTS: The frequency of overall wound implantation on day 17 in the pneumoperitoneum group (24.4%) and nonpneumoperitoneum group (27.4%) tended to be higher than that in the control group (15.3%) (p = 0. 06). There was no significant difference among the three groups in the growth of cancer nodules on the omentum. CONCLUSIONS: The presence of a trocar may be a factor contributing to port site metastasis but CO(2) pneumoperitoneum appears not to be a factor.

Animals↗

PubMed2000

Transformation of Arabidopsis with the codA gene for choline oxidase enhances freezing tolerance of plants.

Arabidopsis thaliana was transformed with the codA gene from Arthrobacter globiformis, which encodes choline oxidase, the enzyme that synthesizes glycinebetaine from choline. The transformation enabled the plants to accumulate glycinebetaine in chloroplasts, and significantly enhanced the freezing tolerance of plants. Furthermore, the photosynthetic machinery of transformed plants was more tolerant to freezing stress than that of wild-type plants. Exogenous application of glycinebetaine also increased the freezing tolerance of wild-type plants, suggesting that the presence of glycinebetaine in transformed plants had enhanced their ability to tolerate freezing stress. Northern blotting analysis revealed that the enhancement of freezing tolerance was not related to the expression of four cold-regulated genes. These results suggest that engineering of the biosynthesis of glycinebetaine by transformation with the codA gene might be an effective method for enhancing the freezing tolerance of plants.

Adaptation, Physiological↗

PubMed2000

Clinical experience with ceramics in total hip replacement.

As part of a search for better articulation in total hip prostheses, the decrease in the thickness of the socket in different total hip prostheses was measured in vivo. The wear rates of (1) RCH 1000 (molecular weight, 10(6)) socket gamma-irradiated with 100 Mrad articulating with a crude COP (stainless steel containing 20% cobalt and 0.01% phosphorous) metal femoral head; (2) RCH 1000 socket nonirradiated articulating with a crude COP femoral head; (3) RCH 1000 socket irradiated with 100 Mrad articulating with an alumina femoral head; (4) ultra high molecular weight polyethylene (molecular weight, 5-6 x 10(6)) socket articulating with an alumina femoral head; and (5) ultrahigh molecular weight polyethylene socket articulating with a stainless steel femoral head (T-28) were 0.06, 0.30, 0.06, 0.1 and 0.25 mm/year, respectively, in the authors' clinical cases. Alumina femoral heads were effective in decreasing wear of the polyethylene socket. However, the wear rates of gamma-irradiated sockets articulating with alumina and with metal femoral heads wear very low and were not different from each other. Regarding the relationship between wear rate and the thickness of the ultra high molecular weight polyethylene socket articulating with a 28 mm alumina femoral head, on radiographs, average wear rates of socket thicknesses of 7, 8, 9, 10 and 11 mm were 0.14, 0.15, 0.12, 0.06, and 0.08 mm/year, respectively. On measuring retrieved prostheses, average wear rates of 7, 8, 9 and 11 mm thickness sockets were 0.2, 0.19, 0.14, and 0.1 mm/year, respectively. The wear of sockets has been proven to be minimal in alumina femoral heads articulating with ultrahigh molecular weight polyethylene sockets thicker than 10 mm.

Arthroplasty, Replacement, Hip↗

PubMed2000

Inactivation of photosystems I and II in response to osmotic stress in Synechococcus. Contribution of water channels.

The effects of osmotic stress due to sorbitol on the photosynthetic machinery were investigated in the cyanobacterium Synechococcus R-2. Incubation of cells in 1.0 M sorbitol inactivated photosystems I and II and decreased the intracellular solute space by 50%. These effects of sorbitol were reversible: Photosynthetic activity and cytoplasmic volume returned to the original values after removal of the osmotic stress. A blocker of water channels prevented the osmotic-stress-induced inactivation and shrinkage of the intracellular space. It also prevented the recovery of photosynthetic activity and cytoplasmic volume when applied just before release from osmotic stress. Inhibition of protein synthesis by lincomycin had no significant effects on the inactivation and recovery processes, an observation that suggests that protein synthesis was not involved in these processes. Our results suggest that osmotic stress decreased the amount of water in the cytoplasm via the efflux of water through water channels (aquaporins), with resultant increases in intracellular concentrations of ions and a decrease in photosynthetic activity.

Aquaporins↗