Paramyxovirus reverse genetics is coming of age.
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Animals↗
Biomedical subjects
A Kato
Publications and source records attributed to A Kato.
Molecular characterization of the PhoP-PhoQ two-component system in Escherichia coli K-12: identification of extracellular Mg2+-responsive promoters.
We identified Mg2+-responsive promoters of the phoPQ, mgtA, and mgrB genes of Escherichia coli K-12 by S1 nuclease analysis. Expression of these genes was induced by magnesium limitation and depended on PhoP and PhoQ. The transcription start sites were also determined, which allowed us to find a (T/G)GTTTA direct repeat in their corresponding promoter regions.
Adenosine Triphosphatases↗
Sendai virus gene start signals are not equivalent in reinitiation capacity: moderation at the fusion protein gene.
In paramyxovirus transcription, viral RNA polymerase synthesizes each monocistronic mRNA by recognizing the gene start (S) and end (E) signals flanking each gene. These signal sequences are well conserved in the virus family; nevertheless, they do exhibit some variations even within a virus species. In Sendai virus (SeV) Z strain, the E signals are identical for all six genes but there are four (N, P/M/HN, F, and L) different S signals with one or two nucleotide variations. The significance of these variations for in vitro and in vivo replication has been unknown. We addressed this issue by SeV reverse genetics. The luciferase gene was placed between the N and P gene so that recombinant SeVs expressed luciferase under the control of each of the four different S signals. The S signal for the F gene was found to drive a lower level of transcription than that of the other three, which exhibited comparable reinitiation capacities. The polar attenuation of SeV transcription thus appeared to be not linear but biphasic. Then, a mutant SeV whose F gene S signal was replaced with that used for the P, M, and HN genes was created, and its replication capability was examined. The mutant produced a larger amount of F protein and downstream gene-encoded proteins and replicated faster than wild-type SeV in cultured cells and in embryonated eggs. Compared with the wild type, the mutant virus also replicated faster in mice and was more virulent, requiring a dose 20 times lower to kill 50% of mice. On the other hand, the unique F start sequence as well as the other start sequences are perfectly conserved in all SeV isolates sequenced to date, including highly virulent fresh isolates as well as egg-adapted strains, with a virulence several magnitudes lower than that of the fresh isolates. This moderation of transcription at the F gene may therefore be relevant to viral fitness in nature.
Animals↗
Decreased plasma tryptophan associated with deep white matter lesions in elderly subjects.
The aim was to identify potentially treatable risk factors for cerebral white matter lesions often found on MRI in elderly persons. findings were assessed on 1.0 T MRI of 178 subjects living in the community and aged 60 years or older. Participants underwent standardised evaluations including standard questionnaires, a physical and neurological examination, cognitive function tests, electrocardiogram, a complete blood chemistry panel, and plasma amino acid measurements. Brain MRI infarcts, deep white matter lesions (DWMLs), and periventricular hyperintensities were found in 26%, 43%, and 29% of the 178 participants, respectively. Subjects with DWMLs were significantly older and had a higher frequency of hypertension, higher systolic blood pressure, and more brain infarcts, but lower plasma concentrations of tryptophan. In the multivariate model, greater age and lower plasma tryptophan concentrations were independently associated with DWMLs. Tryptophan concentrations were inversely related to DWML grading, whereas hypertension and brain infarction were more common in subjects with higher extents of DWMLs. The present study suggests that greater age and lower plasma tryptophan concentrations were important in producing DWMLs in elderly subjects.
Aged↗
A comparative study of Embelia schiperi and Embelia keniensis on blood glucose and triglycerides in normal and epinephrine-induced hyperglycemic mice.
Intraperitoneal administration of the methanol extract of Embelia schiperi (ES) to normal mice caused a significant decrease in blood glucose (p < 0.01) and a significant increase in triglycerides 4 hours after administration at 100 mg/kg (p < 0.01). The toluene fraction of Embelia keniensis methanol extract (TS) showed hypoglycemic and lipid lowering activity 7 hours after intraperitoneal administration at 100 mg/kg. In addition, TS (100 mg/kg) administration significantly decreased blood glucose in epinephrine-induced hyperglycemic mice (p < 0.01). Moreover, ES tended to increase while TS tended to decrease the blood triglycerides in epinephrine-induced hyperglycemic mice. On the other hand, no changes in blood cholesterol were observed after the administration of ES or TS in normal and epinephrine-induced hyperglycemic mice. We found that two species from Embelia, ES and TS, have different activities on blood glucose and triglycerides in normal and epinephrine-induced hyperglycemic mice.
Animals↗
Interleukin-10 inhibits pulmonary NF-kappaB activation and lung injury induced by hepatic ischemia-reperfusion.
Hepatic ischemia and reperfusion cause local and remote organ injury. This injury culminates from an integrated cascade of proinflammatory cytokines, chemokines, and adhesion molecules, many of which are regulated by the transcription factor nuclear factor-kappaB (NF-kappaB). The anti-inflammatory cytokine interleukin-10 (IL-10) has been shown to have inhibitory effects on NF-kappaB. The objective of the current study was to determine whether IL-10 could suppress pulmonary NF-kappaB activation and ensuing lung injury induced by hepatic ischemia-reperfusion. C57BL/6 mice underwent partial hepatic ischemia with or without intravenous administration of IL-10. Hepatic ischemia-reperfusion resulted in pulmonary NF-kappaB activation, increased mRNA expression of tumor necrosis factor-alpha (TNF-alpha), and macrophage inflammatory protein-2 (MIP-2), as well as increased pulmonary neutrophil accumulation and lung edema. Administration of IL-10 suppressed lung NF-kappaB activation, reduced TNF-alpha and MIP-2 mRNA expression, and decreased pulmonary neutrophil recruitment and lung injury. The data suggest that IL-10 protects against hepatic ischemia and reperfusion-induced lung injury by inhibiting lung NF-kappaB activation and the resulting pulmonary production of proinflammatory mediators.
Animals↗
Urea transport processes are induced in rat IMCD subsegments when urine concentrating ability is reduced.
Infusing urea into low-protein-fed mammals increases urine concentration within 5-10 min. To determine which urea transporter may be responsible, we measured urea transport in perfused IMCD3 segments [inner medullary collecting duct (IMCD) segments from the deepest third of the IMCD] from low-protein-fed rats. Basal facilitated urea permeability increased 78%, whereas active urea secretion was completely inhibited. This suggests that upregulation of facilitated urea transport may mediate the rapid increase in urine concentration. Next, expression of active urea transporter(s) in perfused IMCDs was determined in rats with other causes of reduced urine concentrating ability. In untreated and water diuretic rats, IMCD1 segments showed no active urea transport, nor did IMCD2 segments from untreated or hypercalcemic rats. In IMCD1 segments from hypercalcemic rats, active urea reabsorption was induced. The induced active urea reabsorption was completely inhibited by replacing perfusate Na+ with N-methyl-D-glucamine (NMDG+). Active urea secretion was completely inhibited in IMCD3 segments from hypercalcemic rats. In contrast, water diuresis stimulated active urea secretion in IMCD2 segments. The induced active urea secretion was inhibited by phloretin, ouabain, triamterene, or replacing perfusate Na+ with NMDG+. In conclusion, the response of active urea transporters to reductions in urine concentrating ability follows two paradigms: one occurs with hypercalcemia or a low-protein diet, and the second occurs only in water diuresis.
Animals↗
Dose-dependent intestinal and hepatic first-pass metabolism of midazolam, a cytochrome P450 3A substrate with differently modulated enzyme activity in rats.
The dose-dependent first-pass metabolism of midazolam, a cytochrome P450 (CYP) 3A substrate, was separately estimated in the intestine and liver after administration into a jejunal loop of rats with differently modulated enzyme activity. Modulation of CYP3A enzyme activity of Sprague-Dawley rats was performed by pretreating the rats with inducers such as dexamethasone or by co-administering ketoconazole (an inhibitor) with midazolam. Bioavailabilities of midazolam administered into the jejunal loop at a dose of 10 micromol were 12% in untreated (control) rats, and 2% in dexamethasone-pretreated rats. Co-administered ketoconazole (2 micromol) significantly increased the bioavailability to 53% and 7%, respectively, in these rats. The intestinal first-pass metabolism of midazolam administered into the jejunal loop at a dose of 50 nmol in untreated and dexamethasone-pretreated rats, estimated by the mesenteric blood-collecting method in-situ, was 25% and 49% of absorbed amount, respectively. The intestinal first-pass metabolism of midazolam was reduced when ketoconazole (0.5 micromol) was co-administered or when the dose of midazolam was increased to 0.5 micrommol in these rats. Assuming that the contribution of intestinal first-pass metabolism could be negligible when midazolam was administered at a much higher dose of 10 micromol, the estimated hepatic first-pass metabolism of midazolam at a dose of 10 micromol in untreated rats, dexamethasone-pretreated rats, untreated rats given ketoconazole, and dexamethasone-pretreated rats given ketoconazole was, respectively, 86, 97, 46, and 92% of the amounts absorbed. In conclusion, the dose-dependent intestinal first-pass metabolism and the hepatic first-pass metabolism of midazolam in rats with differently modulated CYP3A activities was quantitatively estimated by in-vivo and in-situ absorption studies.
Animals↗
Molecular analysis of a tryptophan-2-monooxygenase gene (IaaM) of Agrobacterium vitis.
Tryptophan-2-monooxygenase genes occur in a number of bacteria and encode the conversion of tryptophan to the plant hormone precursor indole-3-acetamide. The role of these genes in the plant-bacteria interaction is often unclear. However, their function as a virulence determinant is established for Pseudomonas savastanoi and Agrobacterium tumefaciens. Some members of the Agrobacteria, such as Agrobacterium vitis have a limited host range. We have characterized the tryptophan-2-monooxygenase (iaaM) gene of A. vitis strain AG162 and show it is different from other A. vitis strains and related to iaaM of A. rhizogenes. The sequence of AG162 iaaM was deposited in the Genbank database under the accession number AF142716.
Amino Acid Sequence↗
Air drying method using nitrous oxide for chromosome counting in maize.
Nitrous oxide (N2O), colchicine, trifluralin, amiprophos-methyl, 8-hydroxyquinoline, and temperature pretreatments (cold and cold-hot-cold) were compared for chromosome counting in maize (Zea mays L.). Pretreated root tips were prepared by enzymatic maceration and air drying, and the number of countable figures and mitotic indexes were recorded. N2O treatment at 10 atm for 3 hr produced the largest number of countable chromosome figures (266.5 per preparation) and an average of 44.2 nonoverlapped chromosome figures per preparation. Treatment with 0.04% 8-hydroxyquinoline for 3 hr exhibited a moderate number of countable chromosome figures (53.9 per preparation). The effects of colchicine, trifluralin, amiprophos-methyl and temperature pretreatments were limited.
Air↗
Little benefit from mild hypothermia therapy for severely head injured patients with low intracranial pressure.
OBJECT: This study was performed to determine whether mild hypothermia therapy is essential for the treatment of severely head injured patients in whom intracranial pressure (ICP) can be maintained below 20 mm Hg by using conventional therapies. METHODS: Sixteen consecutive severely head injured patients fulfilled the following criteria: the patient's ICP was maintained below 20 mm Hg by using fluid restriction, hyperventilation, and high-dose barbiturate therapy; and the patient had a Glasgow Coma Scale score of 8 or less on admission. After conventional therapies had been applied, the patients were divided randomly into two groups: the mild hypothermia group (HT group; eight patients) and the normothermia group (NT group; eight patients). The HT group received mild hypothermia (intracranial temperature 34 degrees C) therapy for 48 hours followed by rewarming at 1 degree C per day for 3 days, whereas the NT group received normothermia (intracranial temperature 37 degrees C) therapy for 5 days. Specimens of cerebrospinal fluid (CSF) taken from an intraventricular catheter every 24 hours were analyzed for the presence of excitatory amino acids ([EAAs] glutamate, aspartate, and glycine) and cytokines (tumor necrosis factor-alpha, interleukin [IL]-1beta, IL-6, IL-8, and IL-10). The two groups did not differ significantly in patient age, neurological status, or level of ICP. There were no significant differences in daily changes in CSF concentrations of EAAs and cytokines between the two groups. The incidence of pneumonia was slightly higher in the HT group compared with the NT group (p = 0.059). The incidence of diabetes insipidus associated with hypernatremia was significantly higher in the HT group compared with that in the NT group (p < 0.01). The two groups did not differ with respect to their clinical outcomes. CONCLUSIONS: The authors recommend normothermia therapy for the treatment of severely head injured patients in whom ICP can be maintained at lower than 20 mm Hg by using conventional therapies, because mild hypothermia therapy does not convey any advantage over normothermia therapy in such patients.
Adolescent↗
Application of a rigid endoscope to the microsurgical management of 54 cerebral aneurysms: results in 48 patients.
OBJECT: To enhance visual confirmation of regional anatomy, endoscopy was introduced during microsurgery for cerebral aneurysms. The risks and benefits are analyzed in the present study. METHODS: The endoscopic technique was used during microsurgery for 54 aneurysms in 48 patients. Forty-three aneurysms were located in the anterior circulation and 11 were in the posterior circulation. Thirty-eight aneurysms (70.4%) had not ruptured. All ruptured aneurysms in the present series produced Hunt and Hess Grade I or II subarachnoid hemorrhage. After initial exposure achieved with the aid of a microscope, the rigid endoscope was introduced to confirm the regional anatomy, including the aneurysm neck and adjacent structures. The necks of 43 aneurysms were clipped using microscopic control or simultaneous microscopic/endoscopic control. After clipping, the positions of the clip and nearby structures were inspected using the endoscope. Use of the neuroendoscope provided useful information that further clarified the regional anatomy in 44 cases (81.5%) either before or after neck clipping. In nine cases (16.7%), these details were available only with the use of the endoscope. In five cases (9.3%), the surgeons reapplied the clip on the basis of endoscopic information obtained after the initial clipping. There were two cases in which surgical complications were possibly related to the endoscopic procedures (one patient with asymptomatic cerebral contusion and another with transient oculomotor palsy). CONCLUSIONS: It is the authors' impression that the use of the endoscope in the microsurgical management of cerebral aneurysms enhanced the safety and reliability of the surgery. However, there is a prerequisite for the surgeon to be familiar with this instrumentation and fully prepared for the risks and inconveniences of endoscopic procedures.
Adult↗
A case of cicatricial pemphigoid producing severe dysuria due to labial adhesion.
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Aged↗
A Phase I and pharmacokinetic study of TNP-470 administered weekly to patients with advanced cancer.
A Phase I study of angiogenesis inhibitor TNP-470 was conducted in patients with advanced cancer. TNP-470 (25-235 mg/m2) was administered i.v. over 4 h once a week to patients who had solid tumors refractory to the best available treatment or with a high risk of recurrence and who had normal renal, hepatic, and hematological function and no evidence of coagulopathy. The aims of the study were to determine the maximum tolerated dose, dose-limiting toxicities (DLTs), and the pharmacokinetics of TNP-470 given on a once-weekly schedule. Thirty-six patients, ages 23-75 (median, 54 years), with an Eastern Cooperative Oncology Group performance status of 0-2 were treated. The number of patients at each dose level (mg/m2) were 6 (25), 3 (50), 3 (75), 3 (100), 3 (133), 12 (177), and 6 (235). The principal toxicities of TNP-470 were dizziness, lightheadedness, vertigo, ataxia, decrease in concentration and short-term memory, confusion, anxiety, and depression, which occurred at doses of 133, 177, and 235 mg/m2. Two patients treated at 235 mg/m2 experienced DLT in the form of grade III cerebellar neurotoxicity after 6 weeks of treatment. Overall, these neurological symptoms were dose-related, had an insidious onset, progressively worsened with treatment, and resolved completely within 2 weeks of stopping the drug. One patient with malignant melanoma had stabilization of the previously growing disease for 27 weeks while on the treatment. Two patients, one with adenocarcinoma of the colon and the other with a soft tissue sarcoma, had no clinically detectable disease but were at high risk for recurrence at the initiation of treatment and received 13 months and > 3 years of treatment, respectively, with no evidence of disease recurrence. The remaining patients had progression of their disease after 1-6 months of treatment. The mean plasma half-life (t(1/2)) of TNP-470 and its principal metabolite, AGM-1883, were extremely short (harmonic mean, t(1/2) of 2 and 6 min, respectively) with practically no drug detectable in the plasma by 60 min after the end of the infusion. MII, an inactive metabolite, had a considerably longer t(1/2) of approximately 2.6 h. Mean peak TNP-470 concentrations were > or = 400 ng/ml at doses > or = 177 mg/m2. On the basis of this study, the maximum tolerated dose of TNP-470 administered on a weekly schedule was 177 mg/m2 given i.v over 4 h. The principal DLT was neurotoxicity, which appeared to be dose-related and was completely reversible. On the basis of the short plasma t(1/2) of TNP-470, exploration of a prolonged i.v. infusion schedule is warranted.
Adult↗
Antitumor effect of vesnarinone on human hepatocellular carcinoma cell lines.
Vesnarinone, a quinolinone derivative, induces apoptosis and differentiation in some tumor cell lines. We examined the antitumor effect of vesnarinone on three human hepatocellular carcinoma (HCC) cell lines. Vesnarinone suppressed the proliferation of all three HCC cell lines, but did not induce apoptosis and differentiation. Vesnarinone has also been reported to inhibit both nucleoside and nucleobase transport. Concomitant administration of vesnarinone and a de novo nucleotide synthesis inhibitor potentiated the growth-inhibitory effect of vesnarinone on HCC cells. Vesnarinone may be useful as a new biochemical modulator of anticancer agents.
Antineoplastic Agents↗
In vitro assembly of an archaeal D-L-N RNA polymerase subunit complex reveals a eukaryote-like structural arrangement.
Archaeal RNA polymerases (RNAPs) resemble the eukaryotic nuclear RNAPs in complexity, and many of their subunits display a high degree of sequence similarity to their eukaryotic counterparts. Here we describe specific protein-protein contacts present between individual recombinant RNAP subunits from the archaeon Methanococcus jannaschii. Subunits D and L interact specifically with each other in two-hybrid assays. D also interacts under the same conditions with the RPB11 and AC19 subunits from the yeast Saccharomyces cerevisiae, suggesting that essential elements of the binding surface between these proteins have been conserved across the archaeal/eukaryotic evolutionary domain boundary. Interactions between L and RPB3 or AC40 were, however, not detectable. Recombinant D and L subunits associate under in vitro conditions and copurify with each other during size-exclusion chromatography. Addition of an another recombinant subunit (N) to the D-L complex results in the formation of a triple complex. This D-L-N complex resembles the RPB3-RPB11-RPB10 or AC40-AC19-RPB10 complexes in eukaryotic RNAPIIand RNAPI/RNAPIII, respectively. Our data provide evidence for a close similarity in the quaternary arrangement of a subset of archaeal and eukaryotic RNA polymerase subunits and the conservation of the protein-protein contacts formed between them.
Archaea↗
Enhanced secretion of hydrophobic peptide fused lysozyme by the introduction of N-glycosylation signal and the disruption of calnexin gene in Saccharomyces cerevisiae.
The insertion of a hydrophobic pentapeptide (Phe-Phe-Val-Ala-Pro) into the C-terminus in hen egg white lysozyme by genetic modification resulted in an unstable structure which caused little secretion in a yeast expression system, although this modification is useful to enhance bactericidal action to gram-negative bacteria [Ibrahim et al. (1994) J. Biol. Chem. 269, 5059-5063]. To enhance the secretion of the unstable hydrophobic pentapeptide fused lysozymes (H5-Lz), we attempted to introduce the signal sequence (Asn-X-Ser/Thr) of N-linked glycosylation into lysozyme and to suppress the quality control of the unstable mutant in the yeast expression system. The polymannosyl hydrophobic fused lysozyme (H5/G49N-Lz) having the N-glycosylation signal sequence was expressed in the medium at 3.4 times that of unglycosylated lysozyme. Further, the secretion of the unstable mutant lysozyme was done in the Saccharomyces cerevisiae disrupted calnexin gene to avoid the degradation of the unstable mutant by the quality control. Although disruption of the calnexin gene did not lead to gross effects on the levels of growth of S. cerevisiae (W303-1b), the secretion amount of H5/G49N-Lz in calnexin disrupted S. cerevisiae was 2.5 times larger than that in wild type S. cerevisiae. These results suggest that the secretion of unstable glycosylated lysozyme (H5/G49N) was suppressed by the quality control function of calnexin and that the disruption of calnexin is effective to increase the secretion of unstable glycosylated protein.
Animals↗