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K Uchida

Publications and source records attributed to K Uchida.

At least 1,027 records · Page 57Linked to original sources
PubMed1991

Molecular cloning and characterization of a ras p21-like GTP-binding protein (24KG) from rat liver.

We have isolated cDNA clones from a rat liver cDNA library that encode a ras p21-like small GTP-binding protein (24KG) which was purified from the microsomes-Golgi complex fraction of the rat liver. The cloning was accomplished using polymerase chain reaction amplified with a set of oligonucleotide primers which were designed from the partial amino acid sequences for 24KG. The cDNA contained an open reading frame encoding a 216 amino acid protein with a calculated Mr weight of 24,397. This Mr weight was similar to that of the purified 24KG estimated by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The sequence analysis of 24KG revealed that a 24KG cDNA is the rat counterpart of a rab11 cDNA cloned from a Madin-Darby canine kidney cell cDNA library. The 1.0-kilobase 24KG mRNA corresponding to the isolated cDNA was also detected in various rat tissues, such as brain, testis, spleen, and heart.

Amino Acid Sequence↗
PubMed1991

Novel muteins of human tumor necrosis factor alpha.

For chemical synthesis of a gene coding for human tumor necrosis factor alpha (TNF-alpha), DNA sequence predicted by the amino acid sequence of human TNF molecule was prepared. Codons were chosen according to the codon usage in Escherichia coli (E. coli). The 490 bp gene was assembled by enzymic ligation of 42 oligonucleotides and was cloned into a vector (pKK223-3) for high expression of active TNF-alpha in E. coli. With use of site-directed mutagenesis on this DNA, five different muteins of TNF-alpha were synthesized. TNF-M1 and TNF-M4 have deletions of His-73 and Gln-102, respectively. These deletions didn't cause loss of the cytotoxic activity against L929 cells. TNF-M5, which has a substitution of Asp-10 to Arg, had the similar cytotoxic activity to that of TNF-alpha. The cytotoxic spectra against several tumor cells were not changed by this substitution. TNF-M3 has an amino acid substitution of Glu-116 to His which occupies this position in human TNF-beta. This substitution didn't change the cytotoxicity. In addition, evidence was presented that the change of the carboxyl terminal residue doesn't always influence the cytotoxic activity of TNF-alpha. Many different muteins were also isolated by random mutagenesis with hydroxylamine-HCl. One of the muteins, which carries a mutation of His-15 to Tyr, lost the cytotoxic activity almost completely.

Amino Acid Sequence↗
PubMed1991

Release of glucose-mediated catabolite repression due to a defect in the membrane fraction of phosphoenolpyruvate: mannose phosphotransferase system in Pediococcus halophilus.

A spontaneous mutant 9R-4 resistant to 2-deoxyglucose (2DG) was derived from a wild-type strain Pediococcus halophilus I-13. Phosphoenolpyruvate (PEP)-dependent glucose-6-phosphate formation by the permeabilized 9R-4 cells was less than 5% of that observed with the parent I-13. In vitro complementation of PEP-dependent 2DG-6-phosphate formation was assayed with combination of the cytoplasmic and membrane fractions prepared from the I-13 and the mutants (9R-4, and X-160 isolated from nature), which were defective in PEP:mannose phosphotransferase system (man:PTS). The defects in man:PTS of both the strain 9R-4 and X-160 were restricted to the membrane fraction (e.g. EIIman), not to the cytoplasmic one. Kinetic studies on the glucose transport with intact cells and iodoacetate-treated cells also supported the presence of two distinct transport systems in this bacterium as follows: (i) The wild-type I-13 possessed a high-affinity man:PTS (Km = 11 microM) and a low-affinity proton motive force driven glucose permease (GP) (Km = 170 microM). (ii) Both 9R-4 and X-160 had only the low-affinity system (Km = 181 microM for 9R-4, 278 microM for X-160). In conclusion, a 2DG-induced selective defect in the membrane component (EIIman) of the man:PTS could partially release glucose-mediated catabolite repression but not fructose-mediated catabolite repression in soy pediococci.

Biological Transport↗
PubMed1991

Effect of flomoxef on blood coagulation and alcohol metabolism.

The effect of flomoxef, a newly developed oxacephem antibiotic with an N-hydroxyethyltetrazolethiol (HTT) side chain, on blood coagulation and alcohol metabolism was compared with that of a series of cephalosporin antibiotics with N-methyltetrazolethiol (NMTT), thiadiazolethiol (TDT) or methylthiadiazolethiol (MTDT) side chains in position 3' of the cephalosporin nucleus known to cause hypoprothrombinemia and bleeding in patients who are malnourished, debilitated and/or of high age. A disulfiram-like effect caused by inhibition of aldehyde dehydrogenase was observed for NMTT-containing antibiotics. Studies were carried out on healthy volunteers and on rats. Eight-day treatment with 2 g flomoxef i.v. once or twice daily in five and six healthy male volunteers, respectively, did not cause any significant changes in prothrombin time (PT), coagulation factors II, VII, IX or X, in hepaplastin values or fibrinogen levels, activated partial thromboplastin time (APTT), platelet counts, bleeding time, or collagen- and ADP-induced platelet aggregation. Inhibition of vitamin K epoxide reductase was observed in rats treated with flomoxef, yet to a much lesser extent than observed for cephalosporins with NMTT, TDT or MTDT side chains. This defect was quickly normalized by vitamin K injection. There were no differences between oxacephem (1-O) and cephem (1-S) compounds with respect to effects on blood clotting and platelet aggregation. Flomoxef and its side chain HTT showed no influence on alcohol metbolism.

Animals↗
PubMed1991

Reductive cleavage and regeneration of the disulfide bonds in Streptomyces subtilisin inhibitor (SSI) as studied by the carbonyl 13C NMR resonances of cysteinyl residues.

Four enhanced carbonyl carbon resonances were observed when Streptomyces subtilisin inhibitor (SSI) was labeled by incorporating specifically labeled [1-13C]Cys. The 13C signals were assigned by the 15N, 13C double-labeling method along with site-specific mutagenesis. Changes in the spectrum of the labeled protein ([C]SSI) were induced by reducing the disulfide bonds with various amounts of dithiothreitol (DTT). The results indicate that, in the absence of denaturant, the Cys71-Cys101 disulfide bond of each SSI subunit can be reduced selectively. This disulfide bond, which is in the vicinity of the reactive site scissile bond Met73-Val74, is more accessible to solvent than the other disulfide bond, Cys35-Cys50, which is embedded in the interior of SSI. This half-reduced SSI had 65% of the inhibitory activity of native SSI and maintained a conformation similar to that of the fully oxidized SSI. Reoxidation of the half reduced-folded SSI by air regenerates fully active SSI which is indistinguishable with intact SSI by NMR. In the presence of 3 M guanidine hydrochloride (GuHCl), however, both disulfide bonds of each SSI subunit were readily reduced by DTT. The fully reduced-unfolded SSI spontaneously refolded into a native-like structure (fully reduced-folded state), as evidenced by the Cys carbonyl carbon chemical shifts, upon removing GuHCl and DTT from the reaction mixture. The time course of disulfide bond regeneration from this state by air oxidation was monitored by following the NMR spectral changes and the results indicated that the disulfide bond between Cys71 and Cys101 regenerates at a much faster rate than that between Cys35 and Cys50.

Bacterial Proteins↗
PubMed1991

Differential effects of ursodeoxycholic acid and ursocholic acid on the formation of biliary cholesterol crystals in mice.

The preventive effect of 3 alpha, 7 beta, 12 alpha-trihydroxy-5 beta-cholanoic acid (ursocholic acid) and ursodeoxycholic acid on the formation of biliary cholesterol crystals was studied in mice. Cholesterol crystals developed with 80% incidence after feeding for five weeks a lithogenic diet containing 0.5% cholesterol and 0.25% sodium cholate. When 0.25% ursocholic acid or ursodeoxycholic acid was added to the lithogenic diet, the incidence as well as the grade (severity) of the gallstones were reduced. Plasma and liver cholesterol levels were decreased by ursodeoxycholic acid but not by ursocholic acid. Gallbladder cholesterol and phospholipid levels were decreased by both bile acids. The biliary bile acid level was decreased by ursocholic acid but not by ursodeoxycholic acid. After feeding ursocholic acid, its level in the bile was about 25% and the levels of cholic acid and beta-muricholic acid decreased. Fecal sterol excretion was not changed by ursocholic acid, but was increased by ursodeoxycholic acid. After feeding ursocholic acid, fecal excretion of deoxycholic acid, cholic acid, and ursocholic acid increased. No differences were found between mice, with or without gallstones, in plasma and liver cholesterol levels, biliary phospholipid and bile acid levels, fecal sterol and bile acid levels, and biliary and fecal bile acid composition. The results suggest that the lower incidence of crystal formation after treatment with ursocholic acid is probably by a different mechanism than with ursodeoxycholic acid. In the mouse model, ursodeoxycholic acid exerts its effect at least partially, by decreasing cholesterol absorption. Ursocholic acid is well absorbed and excreted into bile and transformed into deoxycholic acid by the intestinal microflora in mice.

Animals↗
PubMed1991

Effects of water-immersion-induced stress and intraperitoneal administration of brain-gut peptides upon immunoreactive thyrotropin-releasing hormone and prostaglandin E2 concentrations in the rat stomach.

The effects of water-immersion-induced stress and intraperitoneal (i.p.) administration of selected neuropeptides on the levels of thyrotropin-releasing hormone (TRH) and prostaglandin E2 (PGE2) were studied in the rat stomach. Water-immersion caused a significant decrease immunoreactive-TRH (ir-TRH) concentrations in the stomach, and a significant increase in ir-TRH concentrations in the gastric juice. The concentrations of PGE2 were significantly increased at 0.5-4 hrs, and significantly decreased at 6-8 hrs after water-immersion. In the experiment of i.p. administration of selected neuropeptides, the level of ir-TRH in the stomach was significantly decreased after VIP injection, whereas it was significantly increased after beta-endorphin injection. The concentration of PGE2 was significantly decreased in the stomach after i.p. administration of TRH and VIP. However, it did not change after beta-endorphin injection. These results indicate that some neuropeptides may participate in regulating the endogenous level of PGE2 and that these interrelations between neuropeptides and PGE2 may be important as ulcerogenic factors in stress ulcers induced by water-immersion in the rat.

Animals↗
PubMed1991

Clinical study of a new fecal occult blood test using a combination assay of hemoglobin and transferrin.

The applicability of a new immunological fecal occult blood test in which hemoglobin (Hb) and transferrin (Tf) are simultaneously assayed was evaluated. The mean absorbance and standard deviation (510/630 nm) obtained by this test was 0.840 +/- 0.805 in 51 fecal samples from patients with colon cancer, 0.248 +/- 0.305 in 95 samples from patients with colon polyps, and 0.104 +/- 0.053 in 110 samples from control patients; these values differed significantly (P less than 0.005). Hb and Tf concentrations were separately determined in the same fecal samples, and qualitative evaluation was performed with a cutoff value of 5.1 micrograms/g feces for Hb and 0.4 micrograms/g feces for Tf. Hb or Tf was positive in 41 of the 51 samples in the colon cancer group, 33 of the 95 in the colon polyp group, and 3 of the 110 in the control group. Qualitative analysis of the values obtained by the combination assay of Hb and Tf with a cutoff value of 0.200 revealed positive rates of 41/51 in the colon cancer group, 33/95 in the colon polyp group, and 4/110 in the control group. These results suggest the usefulness of a combination assay of Hb and Tf as a fecal occult blood test.

Aged↗
PubMed1991

Ventilation-perfusion scintigram in diabetics.

We carried out ventilation and perfusion scintigraphies and pulmonary function tests in 20 diabetics under 50 years of age. 99mTc-MAA perfusion scintigrams showed evidence of minimal nonuniformity (MNU) in four cases (20%) and nonsegmental defect (NSD) in eight cases (40%). There was a ventilation defect in the single-breath image in one case (5%) and a delayed washout in three cases (15%) upon 133Xe ventilation scintigram. In the NSD group, the mean diffusing capacity value was abnormally low and the mean duration of the diabetes was long compared with other groups. The frequency of perfusion defects was higher than that of ventilation abnormalities; moreover, abnormal findings on ventilation scintigrams were very mild compared with those of perfusion defects. Perfusion defects correlated significantly with a decrease in diffusing capacity. These findings suggest that the disturbance in pulmonary arterial perfusion caused a decrease in diffusing capacity in diabetics.

Adult↗
PubMed1991

ras gene alterations in invasive and non-invasive rat bladder carcinomas induced by N-methyl-N-nitrosourea.

We have established a reliable method to induce invasive and non-invasive carcinomas in the heterotopically transplanted urinary bladder of rats by repeated injection of N-methyl-N-nitrosourea (MNU), and examined the alterations of the ras oncogenes and ras oncogene product (p21) in the induced tumours. The incidence of muscle-invasive carcinomas was proportional to the total dose of MNU. When 5, 6 or 12 doses of MNU were used, muscle invasive carcinomas developed in 22, 58 or 45% of animals, respectively, after a mean observation period, respectively, of 54 +/- 9, 45 +/- 13 and 38 +/- 3 weeks. Whereas activated H-ras gene was detected in only one non-invasive carcinoma by DNA transfection assay, seven of 18 non-invasive and invasive carcinomas showed activated ras p21 when examined by immunoblot analysis. Amplification or rearrangement of myc or epidermal growth factor (EGF) receptor gene was not observed. The results indicate that alterations of ras gene may be involved in the development of rat bladder carcinomas but not of invasiveness.

Animals↗
PubMed1991

Total and split renal function assessed by ultrasound Doppler techniques.

We evaluated total and split renal functions from the pattern for renal arterial blood flow detected by ultrasound Doppler in healthy subjects and patients with varying degrees of renal function and disorders other than renovascular hypertension or severe aortic valvular disease. A renal-time pulsed ultrasonic echo-Doppler device at 2.5 MHz was used with a translumbar approach. The ratio of peak diastolic (D) to systolic (S) velocity correlated well with both p-aminohippurate clearance and creatinine clearance. Acceleration time was correlated with the clearance of neither compound. To evaluate the clinical usefulness of ultrasound Doppler in the assessment of split renal function, we compared the D/S ratio with the renal function obtained by radionuclide methods for individuals. The split renal glomerular filtration rate, calculated by a method which makes use of the early renal uptake of 99mTc-diethylenetriam-inepentaacetic acid, correlated well with the D/S ratio. These results indicate that the ultrasonic measurement of renal arterial blood flow by the pulsed Doppler method should be useful for assessment of total and split renal functions.

Adolescent↗
PubMed1991

Evaluation of the invasive potential of superficial bladder cancer by adenosine triphosphate measurement.

In order to predict the malignant potential of superficial bladder cancer, in 121 patients with this malignancy adenine nucleotide levels (adenosine triphosphate, adenosine diphosphate, and adenosine monophosphate) and energy charge in bladder cancer cells were determined. The nucleotides were analyzed according to the method of Bücher, and the energy charge was calculated by applying the formula of Atkinson. It was clearly pointed out that superficial bladder tumors, which became invasive and/or metastatic, had higher levels of adenylate compounds than those which became not. Our results show that a high correlation exists between adenine nucleotide content and tumor progression.

Adenosine Diphosphate↗
PubMed1991

Angiotensin II receptor and postreceptor events in adrenal glomerulosa cells from streptozotocin-induced diabetic rats with hypoaldosteronism.

Streptozotocin-induced chronic diabetic rats develop hyporeninemic hypoaldosteronism. The hypoaldosteronism is associated with selective unresponsiveness of aldosterone to angiotensin II (AII) and an atrophy of the zona glomerulosa. To assess the nature of the adrenal unresponsiveness to AII, we examined the [125I]monoiodoAII binding and the responses of pregnenolone formation and aldosterone production to AII using adrenal glomerulosa cells from diabetic rats 6 weeks after an injection of streptozotocin. Comparisons were made using the cells from control rats treated with vehicle. Diabetic rats had low levels of plasma renin activity, plasma 18-hydroxycorticosterone, and plasma aldosterone, and normal levels of plasma corticosterone and plasma potassium. The zona glomerulosa width was narrower in diabetic than in control rats. Scatchard analysis of the AII binding data demonstrated that the number and affinity of the receptors were similar in the cells from control and diabetic rats. When corrected to an uniform number of cells per group, baseline levels of pregnenolone formation and aldosterone production were similar in the cells from control and diabetic rats. However, cells from diabetic rats had a less sensitive and lower response of both pregnenolone formation and aldosterone production to AII. In contrast, the effect of ACTH on pregnenolone formation and aldosterone production was similar in the cells from control and diabetic rats. These results indicate that the main defect responsible for the hypoaldosteronism may be located on some step(s) mediating between AII receptors and conversion of cholesterol to pregnenolone, presumably on the calcium messenger system, with a disturbance downstream from AII binding.

Aldosterone↗
PubMed1991

Lack of enhanced responsiveness of plasma 18-hydroxycorticosterone and aldosterone to adrenocorticotropin as well as to angiotensin-II during moderate sodium depletion in type II diabetic subjects with normoreninemia.

Responses of plasma aldosterone (PA) and its precursor steroids to alpha ACTH-(1-24) (85.2 nmol, iv) injection and graded angiotensin-II (AII) infusions (3.90 and 7.80 pmol/kg.min for 30 min at each dose) on both 170 and 100 mmol sodium intakes were assessed in 25 type II diabetic subjects with normoreninemia and 11 age-matched normal subjects. The diabetic subjects and the normal subjects did not differ in mean blood pressure, serum electrolytes, and creatinine clearance, except for an increase in fasting plasma glucose (P less than 0.001) in the diabetic subjects. A 100-mmol sodium intake for 4 days produced an increase in basal plasma renin activity (P less than 0.01), plasma 18-hydroxycorticosterone (18-OHB; P less than 0.05), and PA (P less than 0.05), and a decrease in urinary sodium excretion (P less than 0.001) in the two groups. These parameters were similar in the two groups. ACTH injection produced significant increases in plasma cortisol, plasma corticosterone, plasma 18-OHB, and PA (P less than 0.005 or P less than 0.001), and graded AII infusions produced significant increases in plasma 18-OHB and PA (P less than 0.05 or P less than 0.01) during both 170- and 100-mmol sodium intakes in the two groups. In the diabetic subjects, however, the responses of plasma 18-OHB and PA to both ACTH injection and graded AII infusions on a 100-mmol, but not on a 170-mmol, sodium intake were subnormal (P less than 0.05 or P less than 0.01) and were similar to those on a 170-mmol sodium intake. These results indicate a lack of enhanced responsiveness of plasma 18-OHB and PA to both ACTH and AII during moderate sodium restriction in nonazotemic type II diabetic subjects with normoreninemia. It appears that these subjects have selective unresponsiveness of adrenal zona glomerulosa to sodium depletion per se, but not to ACTH or AII.

18-Hydroxycorticosterone↗
PubMed1991

[Effect of (+)-S-145 calcium salt dihydrate, an orally active antagonist of the thromboxane A2/prostaglandin endoperoxide receptor, on platelet aggregation].

The effect of (+)-S-145, (1R, 2S, 3S, 4S)-(5Z)-7-(3-phenylsulfonylaminobicyclo [2.2.1] hept-2-yl) heptenoic acid on human and guinea pig platelet aggregation was examined. (+)-S-145 sodium salt inhibited human platelet aggregation induced by arachidonic acid (AA), 9,11-methanoepoxy-PGH2 (U 46619), collagen, ADP or epinephrine with the IC50 being 0.047-0.146 microM in an in vitro system. When (+)-S-145 calcium salt dihydrate was administered orally to guinea pigs, it inhibited AA-, U-46619- or collagen-induced platelet aggregation dose-dependently with the minimum effective dose being 0.03 mg/kg, and the effective duration being maximally 3 hr. The inhibiting potency and effective duration of (+)-S-145 calcium salt dihydrate after multiple administrations, once a day (0.5 mg/kg) for 7 days, were almost the same as those after a single administration. Although (+)-S-145 sodium salt showed a partial agonist effect (shape change) on platelets in vitro, the effect diminished after pretreatment of the platelets with a lower dose of this compound. These data suggest that (+)-S-145 calcium salt dihydrate is an orally effective potent platelet aggregation inhibitor.

Adenosine Diphosphate↗
PubMed1991

Effect of a high dose of cimetidine on biliary, intestinal and fecal bile acids in rats.

Effect of an intraperitoneal injection of cimetidine at a daily dose of 160 mg/kg for two weeks on biliary lipid secretion, fecal excretion of bile acids and intestinal bile acids was studied in rats. Bile flow, biliary lipid secretion and fecal excretion of bile acids remained unchanged, while the pool size of bile acids decreased by cimetidine treatment. Chenodeoxycholic acid increased with a concomitant decrease of beta-muricholic acid in the biliary bile acid composition of cimetidine-treated rats.

Animals↗
PubMed1991

Pathological studies on cerebral amyloid angiopathy, senile plaques and amyloid deposition in visceral organs in aged dogs.

The relationship between cerebral lesions such as amyloid angiopathy or senile plaques and amyloid deposition in the visceral organs were studied in 90 autopsy cases of dogs, 0 to 19-year-old. Cerebral amyloid angiopathy was detected in 28 aged dogs (mean age: 13.7-year-old) and was found mostly in or around the wall of cerebral meningeal arterioles and capillaries of the neocortex. That condition was often accompanied by cerebral hemorrhage in dogs more than 9 years of age. Senile plaques were detected in the neocortex of the brain of 12 dogs (mean age: 13.2-year-old) and classified into 3 subtypes, i.e., "diffuse plaque", "primitive plaque" and "classical plaque". Among those 3 subtypes of senile plaques, amyloid containing plaques were small in number. In the visceral organs of dogs with cerebral amyloid angiopathy, amyloid deposition was found in the vascular walls or connective tissues of small intestines at a high frequency and sometimes in the vascular walls of the heart, lung, liver and thyroid gland as well as in atrioventricular valves. Amyloid in both cerebral and visceral organs was congophilic and showed green birefringence under poralized light even after potassium permanganate oxidation.

Aging↗