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K Ishioka

Publications and source records attributed to K Ishioka.

At least 19 recordsLinked to original sources
PubMed2000

Canine leptin: cDNA cloning, expression and activity of recombinant protein.

Leptin, the product of the ob gene, is one of the key molecules for the regulation of appetite and whole-body energy balance, and thereby for the pathogenesis of obesity. In an attempt to clarify the roles of leptin in obesity and/or related diseases in companion animals, canine leptin c DNA was cloned by amplifying reverse-transcriptase products of RNA extracted from the adipose tissue of the beagle. A c DNA clone of about 3 kbp contained a 501 bp open reading frame coding a 167-amino acid protein with a 21-amino acid signal peptide. The sequence of a 146-amino acid mature leptin was more than 79 per cent identical to those of other mammals. Northern blot analysis revealed abundant expression of leptin m RNA in adipose tissue, but not in other tissues, in adult beagles. When Chinese hamster ovary cells expressing the rat leptin receptor were stimulated with recombinant canine leptin produced by E. coli, some intracellular signal transduction proteins were phosphorylated, indicating that the recombinant leptin was biologically active. The data reported herein will be helpful for further studies of leptin of the dog in health and disease.

Amino Acid Sequence↗
PubMed2000

Sandwich enzyme-linked immunosorbent assay of canine leptin.

Leptin is the ob gene product secreted from adipocytes in mammals, and thereby its plasma level reflects body fat content. To establish an assay method for leptin in the dog, rabbit anti-canine leptin antibody was obtained using canine recombinant leptin as an antigen. This antibody reacted to canine leptin much stronger than mouse, rat and human leptins. Sandwich enzyme-linked immunosorbent assay (ELISA) using this antibody was developed. The serum leptin levels of 13 healthy dogs were in a range from 1.4 to 5.6 ng/ml with the mean +/- SEM of 3.0 +/- 0.3 ng/ml.

Amino Acid Sequence↗
PubMed1998

Abortive recombination in Escherichia coli ruv mutants blocks chromosome partitioning.

BACKGROUND: All the ruvA, ruvB and ruvC mutants of Escherichia coli are sensitive to treatments that damage DNA, and are mildly defective in homologous recombination. It has been reported that the ruv mutants form nonseptate, multinuclear filaments after low doses of UV irradiation, dependent on the sfiA gene product. In vitro, the RuvAB complex promotes the branch migration of Holliday junctions, and RuvC resolves the junctions endonucleolytically. RESULTS: After a low UV dose (5 J/m2), both delta ruvAB and delta ruvC mutant cells became filamentous, with their chromosomes aggregated in the central region. This corresponded to an increase in nonmigrating DNA on pulsed field gel electrophoresis of the XbaI digested chromosome. Upon further incubation, they produced a large number of anucleoid cells of normal size. A recA mutation, but not a recB mutation, suppressed these phenotypes of the ruv mutants. The ruv polA12(Ts) double mutants were inviable at the nonpermissive temperature and mimicked the morphological phenotypes of the UV irradiated ruv mutants. CONCLUSION: ruvA, B and C mutations block chromosome partitioning in UV irradiated cells because the abortive homologous recombination covalently links chromosomes together. There is a recBCD independent pathway for the recA dependent formation of recombination intermediates. An Ruv-mediated resolution of recombination intermediates is required for the repair of strand breaks produced in UV irradiated cells and in the polA mutant cells.

Adenosine Triphosphatases↗
PubMed1997

ATP-dependent resolution of R-loops at the ColE1 replication origin by Escherichia coli RecG protein, a Holliday junction-specific helicase.

The RecG protein of Escherichia coli is a DNA helicase that promotes branch migration of the Holliday junctions. We found that overproduction of RecG protein drastically decreased copy numbers of ColE1-type plasmids, which require R-loop formation between the template DNA and a primer RNA transcript (RNA II) for the initiation of replication. RecG efficiently inhibited in vitro ColE1 DNA synthesis in a reconstituted system containing RNA polymerase, RNase HI and DNA polymerase I. RecG promoted dissociation of RNA II from the R-loop in a manner that required ATP hydrolysis. These results suggest that overproduced RecG inhibits the initiation of replication by prematurely resolving the R-loops formed at the replication origin region of these plasmids with its unique helicase activity. The possibility that RecG regulates the initiation of a unique mode of DNA replication, oriC-independent constitutive stable DNA replication, by its activity in resolving R-loops is discussed.

Adenosine Triphosphate↗
PubMed1997

Effects of intracoronary fentanyl on left ventricular mechanoenergetics in the excised cross-circulated canine heart.

BACKGROUND: It is still unclear whether fentanyl directly alters left ventricular (LV) contractility and oxygen consumption. This is because of the difficulty in defining and evaluating contractility and energy use independently of ventricular loading conditions and heart rate in beating whole hearts. METHODS: This study was conducted to clarify the mechanoenergetic effects of intracoronary fentanyl in six excised cross-circulated canine hearts. The authors used the framework of the E(max) (a contractility index)-PVA (systolic pressure-volume area, a measure of total mechanical energy)-VO2 (myocardial oxygen consumption per beat) relationship practically independent of ventricular loading conditions. The authors measured LV pressure, volume, coronary flow, and arteriovenous oxygen content difference to calculate E(max), PVA, and VO2. They first obtained the VO2-PVA relationship for varied LV volumes at control E(max). The authors then obtained the VO2-PVA relationship at a constant LV volume, whereas coronary blood fentanyl concentration was increased in steps up to 240 ng/ml. Finally, they obtained the VO2-PVA relationship for varied LV volumes at the final dose of fentanyl. RESULTS: Fentanyl at any concentrations did not significantly change E(max), PVA, and VO2 from the control. The linear end-systolic pressure-volume relations and their slopes were virtually the same between the control and fentanyl volume loading in each heart. Further, either the slope (oxygen cost of PVA) or the VO2 intercept (unloaded VO2) of the linear VO2-PVA relationship remained unchanged by fentanyl. CONCLUSIONS: These results indicate that intracoronary fentanyl produces virtually no effects on LV mechanoenergetics for a wide range of its blood concentration.

Anesthetics, Intravenous↗
PubMed1997

Effects of intracoronary fentanyl on left ventricular mechanoenergetics in the excised cross-circulated canine heart (revised publication).

BACKGROUND: It is still unclear whether fentanyl directly alters left ventricular (LV) contractility and oxygen consumption. This is because of the difficulty in defining and evaluating contractility and energy use independently of ventricular loading conditions and heart rate in beating whole hearts. METHODS: This study was conducted to clarify the mechanoenergetic effects of intracoronary fentanyl in six excised cross-circulated canine hearts. The authors used the framework of the Emax (a contractility index)-PVA (systolic pressure-volume area, a measure of total mechanical energy)-VO2 (myocardial oxygen consumption per beat) relationship practically independent of ventricular loading conditions. The authors measured LV pressure, volume, coronary flow, and arteriovenous oxygen content difference to calculate Emax, PVA, and VO2. They first obtained the VO2-PVA relationship for varied LV volumes at control Emax. The authors then obtained the VO2-PVA relationship at a constant LV volume, whereas coronary blood fentanyl concentration was increased in steps up to 240 ng/ml. Finally, they obtained the VO2-PVA relationship for varied LV volumes at the final dose of fentanyl. RESULTS: Fentanyl at any concentrations did not significantly change Emax, PVA, and VO2 from the control. The linear end-systolic pressure-volume relations and their slopes were virtually the same between the control and fentanyl volume loading in each heart. Further, either the slope (oxygen cost of PVA) or the VO2 intercept (unloaded VO2) of the linear VO2-PVA relationship remained unchanged by fentanyl. CONCLUSIONS: These results indicate that intracoronary fentanyl produces virtually no effects on LV mechanoenergetics for a wide range of its blood concentration.

Anesthetics, Intravenous↗
PubMed1997

Roles of the recG gene product of Escherichia coli in recombination repair: effects of the delta recG mutation on cell division and chromosome partition.

The products of the recG and ruvAB genes of Escherichia coli are both thought to promote branch migration of Holliday recombination intermediates by their junction specific helicase activities in homologous recombination and recombination repair. To investigate the in vivo role of the recG gene, we examined the effects of a recG null mutation on cell division and chromosome partition. After UV irradiation at a low dose (5J/m2), delta recG mutant filamentous cells with unpartitioned chromosomes. A mutation in the sfiA gene, which encodes and SOS-inducible inhibitor of septum formation, partially suppressed filamentation of recG mutant cells, but did not prevent the formation of anucleate cells. The sensitivity of UV light and the cytological phenotypes after UV irradiation of a recA recG double mutant were similar to a recA single mutant, consistent with the role of recG, which is assigned to a later stage in recombinant repair than recA. The recG ruvAB and recG ruvC double mutants were more sensitive to UV, almost as sensitive as the recA mutant and showed more extreme phenotypes concerning filamentation and chromosome nondisjunction, both after UV irradiation and without UV irradiation than either recG or ruv single mutants. The recG polA12 (Ts) mutant, which is temperature sensitive in growth, formed filamentous cells with centrally located chromosome aggregates when grown at nonpermissive temperature similar to the UV irradiated recG mutant. These results support the notion that recG is involved in processing Holliday intermediates in recombination repair in vivo. We suggest that the defect in the processing in the recG mutant results in accumulation of nonpartitioned chromosomes, which are linked by Holliday junctions.

Bacterial Proteins↗
PubMed1996

Molecular analysis of the Pseudomonas aeruginosa genes, ruvA, ruvB and ruvC, involved in processing of homologous recombination intermediates.

In Escherichia coli, the products of the ruvA, ruvB and ruvC genes are all involved in the processing of recombination intermediates (Holliday structures) into recombinant molecules. We cloned a 9.4-kb DNA fragment from Pscudomonas aeruginosa PAO1 in a plasmid by functional complementation of the UV sensitivity of an E. coli strain with ruvABC deleted. In P. aeruginosa, the ruv region seemed to form a non-SOS regulated single operon consisting of orf26-ruvC-ruvA-ruvB, while in this region of E. coli, ruvA and ruvB form an SOS-regulated operon, orf26 and ruvC form a non-SOS operon, and these two operons are split by orf23. The deduced amino acid sequences of P. aeruginosa RuvA, RuvB and RuvC proteins were 55, 72 and 55% identical to those of the corresponding E. coli Ruv proteins. The individual ruv genes of P. aeruginosa complemented the corresponding single ruv mutations of E. coli, suggesting that the P. aeruginosa Ruv proteins can interact functionally with their E. coli Ruv partners in forming heterologous complexes. The sequence alignments of the Ruv proteins were extended by incorporation of data about the putative ruv genes obtained from data banks, and the RuvB sequences were conspicuously more conserved than the RuvA and RuvC sequences.

Amino Acid Sequence↗
PubMed1994

[Effects of urography performed twice at a close interval with contrast media on renal function and morphology in rabbits].

This study was carried out in order to clarify the effects of rapid intravenous urography performed twice at a three-day interval with contrast medium on renal function and morphologic change in normal rabbits as a means to observe progress or to reexamine patients. The ionic high-osmolal amidotrizoate, non-ionic low-osmolal iopamidol, and ionic low-osmolal ioxaglate were used. A second urography was performed and curves of kidney density plotted against time and nephrograms were obtained. Serum BUN and creatinine were measured, and the kidneys were removed for histologic observations. Following amidotrizoate administration, curves of kidney density showed significantly slower clearance than with iopamidol and ioxaglate. Nephrograms at 40 seconds showed that opacification of the renal pelvis and urinary tracts also was poor. Serum creatinine following the second administration of amidotrizoate was significantly higher than the values before and at the first urography, and both serum BUN and creatinine were significantly higher than they were after iopamidol and ioxaglate. Although light microscopic changes were obscure, marked changes were observed by electron microscopy. In conclusion, intravenous urography conducted twice at a close interval with amidotrizoate induces impairment of glomerular function and morphologic change.

Animals↗
PubMed1994

Effects of capsaicin on mechanoenergetics of excised cross-circulated canine left ventricle and coronary artery.

Capsaicin selectively acts on sensory nerve endings in cardiac muscles and coronary arterial smooth muscles. Capsaicin at high doses has cell-nonselective effects including both inhibition of cardiac muscle exciteability and enhancement of vascular smooth muscle tone. We studied whether and how intracoronary infusion of capsaicin affects mechanoenergetics of the excised blood-perfused canine heart and coronary vascular resistance. We found that capsaicin at low concentrations increased Emax (a contracility index) and oxygen consumption (VO2) possibly due to a specific action on capsaicin-sensitive sensory nerves in left ventricular muscles, though in a small number of hearts (3/10). This result coincides with the reported histochemical observations that the distribution of capsaicin-sensitive sensory nerves in the canine left ventricle is not dense. Capsaicin at high doses dose-dependently decreased Emax and proportionally decreased coronary flow. It also lowered the linear VO2-PVA (pressure-volume area; total mechanical energy) relationship without a change in the slope, decreasing unloaded VO2 (VO2 intercept of the VO2-PVA relation). These effects of high-dose capsaicin seem to be direct negative inotropic action on cardiac muscles associated with enhancement of coronary arterial smooth muscle tone, since these effects were not desensitized. No morphological changes of myocardial cells or mitochondria were detected. Therefore, the negative inotropic action is not due to the toxic effect of capsaicin.

Animals↗
PubMed1994

Studies on mutations in male germ cells of transgenic mice following exposure to isopropyl methanesulfonate, ethylnitrosourea or X-ray.

Transgenic mice have recently been used for mutagenesis assays in vivo. The present study was undertaken to clarify whether such assays can detect mutations induced after treatment of male germ cells in mouse with isopropyl methanesulfonate (iPMS), ethylnitrosourea (ENU) or X-ray irradiation. The transgenic mice used for assay are Muta Mouse (MM) strain, which carries 80 copies of the bacterial lacZ gene per cell as targets for mutagenesis. Male MM animals were given a single intraperitoneal injection of 200 mg/kg iPMS, 150 mg/kg ENU or were irradiated with 500 rads of X-rays. Vasa deferential sperm, caudal epididymal sperm and/or whole testes were extracted at various times after treatment with each agent. After the genomic DNA was extracted from each tissue, mutation analysis at the lacZ locus was carried out by the method of Myhr et al. The spontaneous lacZ- mutant frequencies were on the order of 10(-5)-10(-6). The lacZ- mutant frequencies in all treatment groups were increased over the control animals. The iPMS-induced mutant frequency in postmeiotic stages was low. However, ENU induced relatively high mutant frequencies in the spermatogonia. X-rays induced mutant frequencies in the late spermatid and early spermatid stages that were higher than the mutant frequencies in spermatogonia. Mutant frequencies in MM detected after treatment of male germ cells with ENU or X-rays were lower than mutant frequencies detected by the mouse specific-locus test in previous reports. Hence, considering the lower resolution power of the transgenic animal mutagenesis assays using the target lacZ gene compared with the specific locus test, to detect mutations induced in male germ cells, it is not clear whether this assay is a practical alternative to the specific locus test.

Animals↗
PubMed1994

Energetics of the negative and positive inotropism of pentobarbitone sodium in the canine left ventricle.

OBJECTIVE: Pentobarbitone (sodium) is an anaesthetic widely used in animal experiments. It is known to be a cardiovascular depressant and a coronary dilator, but its effects on myocardial energetics in relation to its negative and positive (due to Gregg's phenomenon) inotropism have not been studied. The aim of this study was therefore to determine whether and how pentobarbitone affects cardiac mechanoenergetics compared with other negative inotropic agents for which data are already available. METHODS: The effects of graded doses of intracoronary pentobarbitone on mechanoenergetics were studied in the excised cross circulated left ventricles of 12 dogs. The framework of the Emax (a contractility index)--VO2 (myocardial oxygen consumption)--PVA (systolic pressure-volume area, a measure of total mechanical energy) relationships was fully utilised. RESULTS: Pentobarbitone increased Emax at low doses in five of the 12 hearts. In two of these five hearts, a marked coronary dilatation was found. Pentobarbitone decreased Emax dose dependently at high doses in all the hearts and lowered the VO2 intercept but not the slope (oxygen cost of PVA) of the VO2-PVA relation. There was no difference in oxygen cost of Emax between pentobarbitone and CaCl2, although they have opposite inotropism. These findings suggest that pentobarbitone depresses myocardial mechanoenergetics via suppression of total calcium handling in the excitation-contraction-relaxation coupling. CONCLUSIONS: Pentobarbitone at low doses partly acts as a positive inotropic agent, but at high doses it acts as a negative inotropic agent like beta blockers and calcium antagonists on cardiac mechanoenergetics in canine blood perfused hearts.

Animals↗
PubMed1994

Estimation of total carbon dioxide contents in canine coronary arterial and venous whole blood samples.

We examined whether total carbon dioxide content (CCO2) estimation equations for human whole blood in the literature are applicable to canine coronary arterial and venous blood samples. PCO2 of the tested blood samples covered 19-52 mmHg; PO2 19-398 mmHg. Three CCO2 estimation equations developed by Kelman, Godfrey, and Douglas et al yielded CCO2 values over a range of 26.7-54.9 vol%, where plasma CCO2 values calculated by the Henderson-Hasselbalch equation ranged over 27.7-62.2 vol%. The estimated blood CCO2 values were almost identical among the three equations. They also closely correlated with plasma CCO2 values measured after hemolysis with saponin. We conclude that the human whole blood CCO2 estimation equations are applicable to canine coronary arterial and venous blood samples. These CCO2 equations may be used for canine cardiac metabolic studies.

Animals↗
PubMed1994

A high-resolution line sensor-based photostereometric system for measuring jaw movements in 6 degrees of freedom.

Photostereometry has widely been applied to the measurement of mandibular movements in 6 degrees of freedom. In order to improve the accuracy of this measurement, we developed a system utilizing small LEDs mounted on the jaws in redundant numbers and a 5000 pixel linear charge-coupled device (CCD) as a photo-sensor. A total of eight LEDs are mounted on the jaws, in two sets of four, by means of connecting facebows, each weighing approximately 55 g. The position of the LEDs are detected in three-dimensions by two sets of three CCD cameras, located bilaterally. The position and orientation of the mandible are estimated from the positions of all LEDs measured in the sense of least-squares, thereby effectively reducing the measurement errors. The static overall accuracy at all tooth and condylar points was considered to lie within 0.19 and 0.34 mm, respectively, from various accuracy verification tests.

Biomedical Engineering↗