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Biomedical subjects

T Sakamoto

Publications and source records attributed to T Sakamoto.

At least 451 records · Page 25Linked to original sources
PubMed1997

Temperature-regulated mRNA accumulation and stabilization for fatty acid desaturase genes in the cyanobacterium Synechococcus sp. strain PCC 7002.

Cyanobacteria acclimate to low-temperature conditions by desaturating their membrane lipids. The desB (omega 3 desaturase) and desC (delta 9 desaturase) genes of Synechococcus sp. strain PCC 7002 were cloned and characterized, and the expression of the desA (delta 12 desaturase), desB and desC genes was studied as a function of temperature. The steady-state mRNA abundance for the desA gene was threefold higher in cells grown at 22 degrees C than in cells grown at 38 degrees C. desB transcripts were not detected at 38 degrees C, but were abundant in cells grown at 22 degrees C. Levels of desC mRNA were similar at both growth temperatures. The mRNA levels of each desaturase gene increased within 15 min of a temperature shift-down to 22 degrees C, and mRNA levels recovered within 15 min after a shift-up to 38 degrees C. The cold-induced accumulation of transcripts from the desA and desB genes was suppressed by the addition of chloramphenicol, but the transient elevation of the desC transcript levels at 22 degrees C was not affected by chloramphenicol. The half-lives of the desA and desB mRNAs were significantly longer in cells grown at 22 degrees C than in cells grown at 38 degrees C, but the desC mRNA had a similar half-life at both temperatures. These studies reveal three patterns of temperature regulation for the desaturase genes, whose expression is tightly controlled by a combination of mRNA synthesis and stabilization. These studies demonstrate that elevation of desaturase mRNA levels is not the rate-limiting event during the low-temperature acclimation of cyanobacteria.

Blotting, Northern↗
PubMed1997

Complete regression of established murine hepatocellular carcinoma by in vivo tumor necrosis factor alpha gene transfer.

BACKGROUND & AIMS: Although tumor necrosis factor (TNF)-alpha possesses a potent antitumor activity, systemic administration of TNF-alpha causes severe side effects. To circumvent this, the efficacy of tumor cell-targeted TNF-alpha gene therapy was investigated. METHODS: Murine hepatocellular carcinoma (HCC) cells were infected with MNSM-Alb e/p-TNF-alpha retroviruses carrying the murine TNF-alpha gene under the transcriptional control of the murine albumin gene promoter, and antitumor effects induced by TNF-alpha gene transfer were examined in vitro and in vivo. RESULTS: Although MNSM-Alb e/p-TNF-alpha retrovirally infected HCC cells showed the same in vitro cell growth as parental HCC cells, they lost their tumorigenicity when implanted in syngeneic mice and induced tumor immunity against parental HCCs. The retrovirally infected HCC cells also significantly inhibited the tumorigenicity of previously implanted parental HCCs. Furthermore, intratumoral administration of MNSM-Alb e/p-TNF-alpha retroviruses showed the antitumor effect against established HCCs, resulting in significantly prolonged survival periods. Most importantly, intratumoral implantation of MNSM-Alb e/p-TNF-alpha retroviral-producing cells completely abrogated established HCCs in mice. CONCLUSIONS: These results indicate the potential efficacy of transferring the TNF-alpha gene via retroviral vectors directly into tumors for gene therapy against HCCs.

Animals↗
PubMed1997

Effect of electric pulses and antiproliferative drugs on cultured bovine retinal pigment epithelial cells.

PURPOSE: The effect of antiproliferative agents with high voltage electric pulses (EP) on the proliferation of retinal pigment epithelial (RPE) cells was investigated. METHODS: Bovine RPE cells were cultured and exposed briefly to an antiproliferative agent: 0.005 to 500 microM of bleomycin (BLM), 0.005 to 500 microM of mitomycin C (MMC) or 0.05 to 5000 microM of 5-fluorouracil (5-FU) with or without high voltage electric pulses (EP: 2,000V/cm, 98 microseconds, 8 pulses). Streptomycin (SM) was used as a negative control. Cell proliferation with or without antiproliferative agents was assessed on day 3. DNA fragmentation was assessed by flow cytometric analysis. RESULTS: Treatment with BLM, MMC or 5-FU inhibited cell proliferation in a dose-dependent manner, with and without EP (p < 0.05). EP enhanced the inhibitory effect of BLM as much as 3,000 times (ID50: BLM without EP; 163.7 microM, BLM with EP; 0.0574 microM, MMC without EP; 132.4 microM, MMC with EP; 26.2 microM, 5-FU without EP; 616.4 microM, 5-FU with EP; 873.8 microM). EP treatment showed an inhibitory effect of BLM on cell proliferation more prominently than BLM alone. The cell numbers of 0.5 microM BLM treatment without EP were 87.4 +/- 11.7% (mean +/- SD), whereas the cell numbers of 0.5 microM BLM with EP were 21.1 +/- 2.16% (p < 0.005). BLM treatment with EP increased the apoptosis like DNA fragmentation in the flow cytometric DNA histogram, showing dominant accumulation in the A0 region, which is the population of DNA fragmentation. (The cell population of the A0 region: control, 3.4%; EP alone, 18.9%; BLM treatment without EP, 32.0%; with EP, 93.8%.) CONCLUSIONS: EP treatment enhances the inhibitory effect of BLM on RPE cell proliferation. The combination of electric pulse and BLM treatment can localize the drug effect and reduce the necessary dose of the antiproliferative agent in comparison to BLM treatment alone.

Animals↗
PubMed1997

Hypericin inhibits choroidal endothelial cell proliferation and cord formation in vitro.

PURPOSE: To evaluate the effect of hypericin on bovine choroidal endothelial cell proliferation and cord formation and on protein kinase C activity. METHODS: The effect of hypericin (0.1-5 microM) on bovine choroidal endothelial cell proliferation was determined by cell number counting and a 3H-thymidine uptake assay in media containing 1, 5 or 10% serum. For the cord formation assay, bovine choroidal endothelial cells were seeded on basement membrane matrix, and the lengths of the capillary-like structures (cords) formed were quantified by image analysis. The effect of hypericin on cord formation was evaluated in the presence of serum or vascular endothelial growth factor. The effect of hypericin on protein kinase C activity was also measured in the presence or absence of light. RESULTS: Hypericin inhibited bovine choroidal endothelial cell proliferation in a dose-dependent manner in the presence of light but not in the dark. Serum dose-dependently masked the inhibition of DNA synthesis by hypericin. Cord formation by bovine choroidal endothelial cells was stimulated by serum or vascular endothelial growth factor and inhibited by hypericin in the presence of light. Protein kinase C activity was completely inhibited by hypericin in the presence of light but only mildly inhibited in the absence of light. CONCLUSIONS: Hypericin inhibits bovine choroidal endothelial cell proliferation and cord formation and choroidal endothelial cell protein kinase C activity. These results suggest that hypericin should be further investigated in animal models for its potential to inhibit subretinal neovascularization.

Animals↗
PubMed1997

Hair analysis for drugs of abuse. XVII. Simultaneous detection of PCP, PCHP, and PCPdiol in human hair for confirmation of PCP use.

The paper reports the simultaneous detection hair of phencyclidine (PCP) and its two major metabolites, 1-(1-phenylcyclohexyl)-4-hydroxypiperidine (PCHP) and trans-1-(1-phenyl-4-hydroxycyclohexyl)-4'-hydroxypiperidine (t-PCPdiol) in human hair. The detection of these metabolites provides definitive evidence that a positive hair analysis result is due to active PCP use and not due to external contamination of the hair specimen. Hair (5 mg) from known PCP users was washed three times with 0.1% sodium dodecyl sulfate for 1 min before analysis. Three extraction methods were compared: methanol-5N HCl (20:1) (Method A), 10% HCl (Method B), and 2N sodium hydroxide digestion (Method C). PCP-d5 and PCHP-d5 were used as internal standards. Extracts were purified by Bond Elut Certify solid-phase extraction procedures. Samples were derivatized with N,O-bis-trimethylsilyl acetamide and analyzed by gas chromatography-mass spectrometry. Compared with Method A, the extraction efficiencies of Methods B and C for PCP were 83-89%; however, the extraction efficiencies of Methods B and C for the two metabolites were only half or less than that of Method A. Method A was therefore selected for the analysis of clinical hair specimens from eight PCP users. The coefficients of variation of this method (n = 5) for PCP at 4 ng/mg and for PCHP and t-PCPdiol at 0.2 ng/mg were 2.13, 6.09, and 9.38%, respectively. In the eight hair specimens, PCP values ranged between 0.33 and 14 ng/mg. PCHP between 0.02 and 0.12 ng/mg, and trans-PCPdiol between 0.09 and 0.45 ng/mg. It was found that t-PCPdiol was the major metabolite in the PCP users' hair specimens, although t-PCPdiol was a minor metabolite in the hair specimens of rats intoxicated with PCP.

Animals↗
PubMed1997

Properties of a hammerhead ribozyme with deletion of stem II.

The properties of a mutant hammerhead ribozyme system, which consists of two RNA oligomer strands and in which stem II is deleted (replaced with a UUUU loop), are described. The effects of temperature, pH, and metal ions on the cleavage reaction were similar to those for the parent ribozyme with stem II. The mutant ribozyme showed a much lower cleavage rate (kcat = 0.04 min-1) in the presence of 10 mM MgCl2, where the parent ribozyme showed full cleavage activity. However, increasing the concentration of MgCl2 from 10 to 100 mM restored the cleavage activity of the mutant ribozyme to the original level (kcat = 0.2 min-1). CD titration experiments with MgCl2 using a noncleavable substrate were carried out. Deletion of stem II resulted in an about 20-fold reduction of the apparent Mg2+ binding affinity when the Mg2+ concentrations of half-saturation are compared. The results were analyzed by curve-fitting analysis and compared with those for the parent ribozyme. The analysis showed that the Mg2+ concentration dependence data in CD and cleavage experiments for the mutant enzyme can be explained by a two Mg2+ ion binding mechanism. These results that stem II is important for maintaining the conformation of the catalytic core suitable for Mg2+ binding.

Base Sequence↗
PubMed1997

Properties of hepatitis delta virus ribozyme, which consists of three RNA oligomer strands.

Properties of a hepatitis delta virus (HDV) RNA ribozyme system, which consists of three RNA oligomer strands (substrate 8-mer; enzyme 16-mer plus 35-mer) and contains a hybrid sequence of genomic and antigenomic RNA cores, are reported. Effects of Mg2+ concentration, divalent metal ion species, pH, and temperature on the cleavage activity were examined. The substrate cleavage activity increased with increasing Mg2+ concentration (0-100 mM). Ca2+ and Mn2+ ions were the most effective divalent cations and Mg2+ was less effective. The cleavage activity increased with increasing pH (5-7.5). The optimum temperature for the cleavage activity was 25-40 degrees C. The Mg2+ concentration, pH and temperature dependencies are different from those reported for the single-strand ribozymes (about 90-mer) although the divalent metal ion preference is very similar. Conformational change induced by Mg2+ ion titration was monitored by CD. The CD data and the activity-Mg2+ concentration data were analyzed by curve-fitting analysis using equations derived for multiple metal ion binding mechanisms. The data can be explained by a model in which three Mg2+ ions bind to one ribozyme unit.

Base Sequence↗
PubMed1997

Hairpin structure of an RNA 28-mer, which contains a sequence of the enzyme component of a hammerhead ribozyme system: evidence for tandem G:A pairs that are not of side-by-side type.

An RNA 28-mer (Rz28) was obtained as a major product by in vitro transcription with T7 RNA polymerase of a promoter-template DNA, which contains a sequence for the enzyme component, RNA 24-mer (Rz24), of a mutant hammerhead ribozyme system. Sequence analysis and enzymatic probing study showed that Rz28 has 4 extra nucleotides at the 3'-terminus, the sequence of which is complementary to that of the 5'-terminal sequence of Rz24, and forms a stable hairpin structure. NMR studies using a 15N-guanine-labeled derivative suggested that Rz28 contains tandem G:A pairs that are not of the side-by-side type which is found in the crystal structure of hammerhead ribozyme complexes. Comparison of the HMQC spectra of 15N-guanine-labeled Rz28 and Rz24 suggested that Rz24 also contains the same type of tandem G:A pairs.

Adenine↗
PubMed1997

The Spirulina platensis adenylate cyclase gene, cyaC, encodes a novel signal transduction protein.

A cyaC gene encoding an adenylate cyclase of the filamentous cyanobacterium Spirulina platensis was sequenced. The predicted amino acid sequence of the C-terminal region of cyaC is similar to the catalytic domains of adenylate cyclases in other cyanobacteria and eukaryotes. The sequences of other regions are similar to those of proteins consisting of the bacterial two-component signal transduction system: the sensory kinase and the response regulator. The predicted gene product of cyaC contains, from the N-terminal end, a receiver domain of the response regulator protein (R1), a domain similar to the ETR1 of Arabidopsis thaliana, a transmitter domain of the sensory kinase protein, a receiver domain of the response regulator protein (R2), and a catalytic domain of adenylate cyclase. The cyaC gene was expressed as an affinity-tagged protein in Escherichia coli, and the recombinant protein was purified. The purified protein had adenylate cyclase activity which was activated by Mu2+. The results of Western blotting using an anti-CyaC antiserum and the S.platensis cell extract confirmed that cyaC gene is expressed in S. platensis.

Adenylyl Cyclases↗
PubMed1997

Risk factors for neurologic deterioration after revascularization surgery in patients with moyamoya disease.

UNLABELLED: To investigate the risk factors for postoperative neurological deterioration in patients with moyamoya disease, we retrospectively reviewed the perioperative course of 368 cases of revascularization surgery in 216 patients with this disease. Risk factors anecdotally associated with postoperative ischemic events were analyzed by comparing groups with or without a history of such events on the operative day. Ischemic events were noted in 14 cases (3.8%), 4 of which were defined as strokes and the others as transient ischemic attack (TIA). Postoperative neurological deterioration more often developed in patients who suffered from frequent TIAs, had precipitating factors for TIA, and underwent indirect nonanastomotic revascularization. The authors conclude that the incidence of postoperative ischemic events were related more to the severity of moyamoya disease and the type of surgical procedure than to other factors, including anesthetic management. IMPLICATIONS: Although preventing stroke is the major concern for patients with moyamoya disease, risk factors for perioperative cerebral ischemia have not been clarified. We retrospectively analyzed the perioperative course in 368 cases with this disease and found that the severity of the disease and type of surgical procedure were major determinants of postoperative cerebral ischemia.

Adolescent↗
PubMed1997

Role of endogenous somatostatin in postprandial hypersecretion of neurotensin in patients after gastrectomy.

OBJECTIVE: The purpose of this report is to elucidate the mechanism of the hypersecretion of neurotensin (NT) after gastrectomy. SUMMARY BACKGROUND DATA: NT secretion induced by fat ingestion is increased after pancreatoduodenectomy or distal gastrectomy. The hypersecretion of NT in the patients undergoing resection of the upper gastrointestinal tract is suppressed by an exogenous somatostatin (SST) analog. METHODS: We observed simultaneously the secretion of NT and SST in the same patients before and after gastrectomy (n = 7). We also observed the secretion of these hormones induced by intraduodenal (ID) fat infusion in the normal volunteers (n = 6). RESULTS: The response of plasma NT to fat ingestion was significantly increased after gastrectomy compared with that before gastrectomy. The response of plasma SST after gastrectomy was significantly suppressed. The response of plasma NT and SST after ID fat infusion in the normal volunteers was similar to the gastrectomized state. CONCLUSION: Diminution of SST secretion, probably caused by the lack of SST cells in the distal part of the stomach, may play a role in augmenting NT secretion after gastrectomy.

Aged↗
PubMed1997

Ileocolon interposition as a substitute stomach after total or proximal gastrectomy.

OBJECTIVE: The authors evaluated ileocolon interposition as a substitute stomach after total gastrectomy (TG) or proximal gastrectomy (PG). SUMMARY BACKGROUND DATA: Although the jejunum frequently is used for reconstruction to create a substitute stomach after TG or PG, there are few reports on ileocolon interposition. METHODS: The authors performed ileocolon interposition in 47 patients who underwent TG (N = 18) or PG (N = 29) for malignant gastric lesion and evaluated the function of this structure as a substitute stomach using esophagoscopy, manometry, pH-metry, emptying time, oral glucose tolerance test (OGTT), and postoperative body weight changes. RESULTS: No patient reported any reflux symptoms or showed endoscopic findings of reflux esophagitis. These results were well supported by manometry and acid loading pH-metry. Emptying time and OGTT showed good capacity as a reservoir of food, and the postoperative body weight averaged more than 90% of preoperative weight. Clinically, no significant difference between these two groups was recognized during long-term follow-up for up to 12 years after operation. There were no cases of direct operative death, and the 5- and 10-year survival rates were 64.7% and 40.2%, respectively. CONCLUSIONS: Ileocolon interposition after TG or PG has the advantages of preventing postoperative reflux esophagitis and of providing functional replacement of the stomach as a reservoir for ingested food.

Adult↗
PubMed1997

Recovery from neuroendocrinological abnormalities and frontal hypoperfusion after remission in a case with rapid cycling bipolar disorder.

A 51-year-old Japanese woman who had been suffering from a rapid cycling affective disorder (RCAD) for 24 years responded to combined clonazepam and carbamazepine therapy. Before remission, she showed neuroendocrinological and neuroimaging abnormalities such as subclinical hypothyroidism with exaggerated response to thyrotropin releasing hormone (TRH) injection, non-suppression on the dexamethasone suppression test (DST) and hypofrontality in cerebral blood flow. Her symptoms improved remarkably soon after adjunctive clonazepam treatment. After remission, her biological markers gradually returned to normal. First, subclinical hypothyroidism improved 2 months after remission. Next, hypofrontality disappeared 18 months later. Furthermore, non-suppression on the DST normalized 24 months later. The normalization of biological markers with apparent recovery from RCAD suggests a decreased risk of relapse into mood disorder. These findings reiterate the importance of following-up on the biological markers in RCAD for years after remission.

Anticonvulsants↗
PubMed1997

Excessive twitch movements in rapid eye movement sleep with daytime sleepiness.

A man who showed excessive twitch movement, such as fragmentary myoclonus (FM) and periodic movements in sleep (PMS) predominantly during REM sleep, is reported. He complained of excessive daytime sleepiness (EDS). After examination, his twitch movements were shown not to accompany narcolepsy, and his EDS were considered to originate from nocturnal sleep disturbance caused by FM and PMS.

Adult↗
PubMed1997

Working pressure for full balloon inflation during intraaortic balloon pumping.

With changes in heart rate and mean aortic pressure, an intraaortic balloon (IAB) works with incomplete inflation. The effects of initial peak positive pressure and plateau holding positive pressure on IAB volume were investigated and minimum pressures necessary to inflate an IAB to a desired volume were established with a standard IAB catheter (Datascope type, 9.5 Fr, 40 ml). Initial peak pressure greater than 200 mm Hg was adequate to attain the desired volume of the IAB. Plateau positive pressure should be altered in accordance with hemodynamic changes.

Analysis of Variance↗