The diurnal variations in content of arginine-vasopressin in the supraoptic nucleus, the paraventricular nucleus and the suprachiasmatic nucleus of rats were determined using radioimmunoassay. In the supraoptic nucleus and the paraventricular nucleus the arginine-vasopressin level was relatively constant during the light phase (the inactive phase). When it became dark, the level of arginine-vasopressin lowered during the early and middle dark phase and then increased to the highest level during the late dark phase. In the suprachiasmatic nucleus the level was stable during the light phase, while in the early and the late dark phase it was significantly higher than that in the middle dark phase.

Animals↗

PubMedDate not supplied

Beating-heart endoscopic coronary artery surgery.

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Aged↗

PubMedDate not supplied

Biosynthesis of polyhydroxyalkanoates (PHA) by recombinant Ralstonia eutropha and effects of PHA synthase activity on in vivo PHA biosynthesis.

Recombinant strains of Ralstonia eutropha PHB 4, which harbored Aeromonas caviae polyhydroxyalkanoates (PHA) biosynthesis genes under the control of a promoter for R. eutropha phb operon, were examined for PHA production from various alkanoic acids. The recombinants produced poly(3-hydroxybutyrate-co-3-hydroxyhexanoate) [P(3HB-co-3HHx)] from hexanoate and octanoate, and poly(3-hydroxybutyrate-co-3-hydroxyvalerate-co-3-hydroxypentano ate) [P(3HB-co-3HV-co-3HHp)] from pentanoate and nonanoate. One of the recombinant strains, R. eutropha PHB 4/pJRDBB39d3 harboring ORF1 and PHA synthase gene of A. caviae (phaC(Ac)) accumulated copolyesters with much more 3HHx or 3HHp fraction than the other recombinant strains. To investigate the relationship between PHA synthase activity and in vivo PHA biosynthesis in R. eutropha, the PHB- 4 strains harboring pJRDBB39d13 or pJRDEE32d13 were used, in which the heterologous expression of phaC(Ac) was controlled by promoters for R. eutropha phb operon and A. caviae pha operon, respectively. The PHA contents and PHA accumulation rates were similar between the two recombinant strains in spite of the quite different levels of PHA synthase activity, indicating that the polymerization step is not the rate-determining one in PHA biosynthesis by R. eutropha. The molecular weights of poly(3-hydroxybutyrate) produced by the recombinant strains were also independent of the levels of PHA synthase activity. It has been suggested that a chain-transfer agent is generated in R. eutopha cells to regulate the chain length of polymers.

Acyltransferases↗

PubMedDate not supplied

Improved production of poly(4-hydroxybutyrate) by Comamonas acidovorans and its freeze-fracture morphology.

Production of poly(4-hydroxybutyrate) [P(4HB)] by Comamonas acidovorans JCM10181 was studied by introducing additional copies of its PHA synthase gene and the beta-ketothiolase gene. A multi-copy-number broad-host-range plasmid vector, pJRD215, was modified to contain the strong hybrid trc promoter in order to express these genes in the wild-type C. acidovorans. Increased copy-number of genes resulted in significant increase in the activities of corresponding enzymes, which could further be increased by inducing with isopropyl-beta-D-thiogalactopyranoside (IPTG), indicating that the expression is under the transcriptional control of the trc promoter. P(4HB) biosynthesis in the recombinant C. acidovorans increased 2-fold to constitute more than 60 wt% of the dry cell weight. No significant decrease in the number-average molecular weights of P(4HB) in the recombinant strain was observed when compared with that of the wild-type. Freeze-fracture electron microscopy of intracellular P(4HB) granules revealed almost similar fracture morphology to the well-known mushroom-type deformation shown by polyhydroxyalkanoates with medium-chain-length monomers.

Acetyl-CoA C-Acyltransferase↗

PubMedDate not supplied

Properties and biodegradability of ultra-high-molecular-weight poly[(R)-hydroxybutyrate] produced by a recombinant Escherichia coli.

Ultra-high-molecular-weight poly[(R)-3-hydroxybutyrate] (P(3HB)) (Mw = 3-11 x 10(6)) was produced from glucose by a recombinant Escherichia coli XL1-Blue (pSYL105) harboring Ralstonia eutropha H16 polyhydroxyalkanoate (PHA) biosynthesis genes. Morphology of ultra-high-molecular-weight P(3HB) granules in the recombinant cells was studied by transmission electron microscopy. The recombinant E. coli contained several P(3HB) granules within a cell. Freeze-fracture morphology of ultra-high-molecular-weight P(3HB) granules showed the needle-type as that of P(3HB) granules in R. eutropha. Both the P(3HB) granules in wet cells and wet native granules isolated from the recombinant cells proved to be amorphous on the X-ray diffraction patterns. Mechanical properties of ultra-high-molecular-weight P(3HB) films were markedly improved by stretching over 400%, resulting from high crystallinity and highly oriented crystal regions. Biodegradability of the films of ultra-high-molecular-weight P(3HB) was tested with an extracellular polyhydroxybutyrate depolymerase from Alcaligenes faecalis T1. The rate of enzymatic erosion of P(3HB) films was not dependent of the molecular weight but was dependent of the crystallinity. In addition, it is demonstrated that all ultra-high-molecular-weight P(3HB) films were completely degraded at 25 degrees C in a natural river freshwater within 3 weeks.

Acyltransferases↗

PubMedDate not supplied

Structure and enzymatic degradation of poly(3-hydroxybutyrate) copolymer single crystals with an extracellular PHB depolymerase from Alcaligenes faecalis T1.

Lamellar single crystals of four random copolymers of (R)-3-hydroxybutyrate with different hydroxyalkanoates: poly(3-hydroxybutyrate-co-8 mol%-3-hydroxyvalerate) (P(3HB-co-8%-3HV)), poly(3-hydroxybutyrate-co-10 mol%-4-hydroxybutyrate) (P(3HB-co-10%-4HB)), poly(3-hydroxybutyrate-co-8 mol%-3-hydroxyhexanoate) (P(3HB-co-8%-3HH)) and poly(3-hydroxybutyrate-co-10 mol%-6-hydroxyhexanoate) (P(3HB-co-10%-6HH)), were grown from dilute solutions of chloroform and ethanol. All single crystals have lath-shaped morphology and the second monomer units seem to be excluded from the P(3HB) crystal, on the basis of the electron diffraction diagrams. The enzymatic degradation of P(3HB-co-8%-3HH) and P(3HB-co-10%-6HH) single crystals was investigated with an extracellular PHB depolymerase from Alcaligenes faecalis T1. Adsorption of an extracellular PHB depolymerase, examined using an immuno-gold labelling technique, demonstrated a homogeneous distribution of enzyme molecules with a low concentration on the crystal surfaces. Enzymatic degradation of single crystals progressed from the edges and ends of crystals to yield narrow cracks along their long axes and the small crystal fragments. Lamellar thicknesses of single crystals and molecular weights of copolymer chains remained unchanged during the enzymatic hydrolysis. The above results support the hypothesis that the hydrophobic adsorption of the enzyme contributes to increase the mobility of molecular chains of single crystals and generate the disordered chain-packing regions. The active-site of PHB depolymerase takes place preferentially at the disordered chain-packing regions of crystal edges and ends with endo-exo enzymatic hydrolysis behaviour, termed processive degradation.

Adsorption↗

PubMedDate not supplied

Crystallization behavior and thermal properties of melt-crystallized poly[(R)-3-hydroxybutyric acid-co-6-hydroxyhexanoic acid] films.

Melt-crystallized films of poly[(R)-3-hydroxybutyric acid-co-10mol% 6-hydroxyhexanoic acid] (P[(R)-3HB-co-6HH]) were prepared by isothermal crystallization at various temperatures for 3 days, and subsequently stored at room temperature after the films formed well-developed and volume-filled spherulites. The lamellar morphologies and properties of melt-crystallized films were characterized by means of wide-angle X-ray diffraction, small-angle X-ray scattering, differential scanning calorimetry, transmission electron microscopy, and atomic force microscopy. The melting endotherm of P[(R)-3HB-co-6HH] films was composed of a broad peak starting around room temperature and of a sharper peak starting above the isothermal crystallization temperature. The stacking of flat-on lamellae with lamellar periodicity of 8-10 nm was detected on the surface of P[(R)-3HB-co6HH] films after the primary crystallization at 110 degrees C. On storage at room temperature above the Tg (-5 degrees C) of copolyester, thin crystals of 1-4 nm thickness appeared on the surface of P[(R)-3HB-co-6HH] films crystallized at 110 degrees C. These results suggest that long sequences of (R)-3HB units in a random copolyester form relatively thick P[(R)-3HB] crystalline lamellae during the primary crystallization process at a given crystallization temperature, while shorter sequences of (R)-3HB units, which are incapable of crystallizing at a given crystallization temperature, form relatively thin crystalline lamellae during the subsequent crystallization process at room temperature.

Calorimetry, Differential Scanning↗

PubMedDate not supplied

Structural effects on enzymatic degradabilities for poly[(R)-3-hydroxybutyric acid] and its copolymers.

Poly[(R)-3-hydroxybutyric acid] and its copolymers were prepared by biosynthetic and chemosynthetic methods. The films of polyesters were prepared by both the solution-cast and melt-crystallized techniques. The enzymatic degradation of polyester films was carried out at 37 degrees C in an aqueous solution (pH 7.4) of PHB depolymerase from Alcaligenes faecalis. The rate of enzymatic erosion on the solution-cast films increased markedly with an increase in the fraction of second monomer units up to 10-20 mol% to reach a maximum value followed by a decrease in the erosion rate. Analysis of the water-soluble products liberated during the enzymatic degradation of polyester films showed the formation of a mixture of monomers and oligomers of (R)-3HB and hydroxyalkanoic acids units, suggesting that the active site of PHB depolymerase recognizes at least three monomeric units as substrate for the hydrolysis of ester bonds in a polymer chain. The rate of enzymatic erosion of melt-crystallized polyester films decreased with an increase in crystallinity. PHB depolymerase predominantly hydrolyzed the polymer chains in the amorphous phase and subsequently eroded crystalline phase. In addition, the enzymatic degradation of crystalline phase by PHB depolymerase progressed from the edges of crystalline lamellar stacks. The enzymatic erosion rate of crystalline phase in polyester films decreased with an increase in the lamellar thickness.

Alcaligenes↗

PubMedDate not supplied

Lamellar thickening and cocrystallization of poly(hydroxyalkanoate)s on annealing.

Lamellar thickening behavior of microbial polyesters, poly(3-hydroxybutyrate) [P(3HB)], poly(3-hydroxybutyrate-co-3-hydroxyvalerate) [P(3HB-co-3HV)] and poly(3-hydroxybutyrate-co-4-hydroxybutyrate) [P(3HB-co-4HB)] annealed at various temperatures was investigated to make sure of the occurrence of cocrystallization of both components. All the copolymers showed steep increases in melting points accompanied by partial melting as the annealing temperature increased up to just below the melting points. In contrast, long periods of P(3HB-co-7mol% 3HV) increased to twice, similar to those of P(3HB), with increasing annealing temperature up to just below the melting point, while long periods of P(3HB-co-7mol% 4HB) and P(3HB-co-92mol% 3HV) only increased up to one and a half times. Lattice indices of unit cell of the former crystal were increased slightly, while those of the latter crystal remained unchanged. These results imply that the P(3HB) crystal can occlude the 3HV component to some extent, but hardly includes the 4HB component, and P(3HV) crystal also excludes the 3HB component.

Calorimetry, Differential Scanning↗

PubMedDate not supplied

Progressive intraventricular drop of early diastolic flow velocity reflects impaired active ventricular diastolic function in hypertensive heart disease: comparative study between early diastolic and atrial contraction phases.

Left ventricular (LV) diastolic function in the early diastolic phase includes both active and passive processes, but in the atrial contraction phase it includes only passive processes. To elucidate the relation between the intraventricular dispersion of the flow velocity in diastole and LV diastolic process, 31 normal volunteers and 12 patients with hypertensive heart disease were studied. In these subjects the flow velocity pattern at the mitral tip was recorded simultaneously with regional flow velocity patterns 1, 2, or 3 cm from the mitral tip toward the apex, respectively, with multigate pulsed Doppler echocardiography from the apical long-axis view with the guidance of Doppler color-flow imaging. Although the ratio of regional peak flow velocity/mitral peak flow velocity in the atrial contraction phase decreased from the mitral tip to the apex to the same degree in the normal volunteers and patients with hypertensive heart disease, there was a significant difference in the intraventricular dispersion of the early diastolic flow velocity between the two groups. These results suggest that the progressive intraventricular drop of the flow velocity in the early diastolic phase in patients with hypertensive heart disease may reflect the impairment of active rather than passive LV diastolic function.

Adult↗

PubMedDate not supplied

Decreased and abnormal left ventricular filling in acute heart failure: role of pericardial constraint and its mechanism.

Pericardial constraining force is minimal in normal hearts; however, it is considered to be prominent in moderate to severe heart failure. Thus, effects of the pericardium on pulsed Doppler transmitral flow velocity pattern were examined in 17 dogs with acute left ventricular dysfunction. Left ventricular dysfunction with left ventricular end-diastolic pressure > or = 15 mm Hg was produced by injection of microspheres into the left coronary artery. Transmitral flow velocity pattern, left atrial and left ventricular diameters, and high-fidelity left atrial and left ventricular pressures were recorded before and after pericardiectomy. In five of the 17 dogs, mitral regurgitation with giant "v" wave of left atrial pressure occurred with reductions of left ventricular systolic pressure and peak rate of the left ventricular pressure fall (dP/dt) after pericardiectomy. In the other 12 dogs, peak early and late diastolic filling velocities increased with a decrease in left ventricular minimal pressure and increases in left arterial and left ventricular diameters and left atrial and left ventricular compliance after pericardiectomy. In these 12 dogs, left atrial to left ventricular crossover pressure, left ventricular end-diastolic pressure, and references for left ventricular relaxation did not change after pericardiectomy. Thus the release from pericardial constraining force in severe heart failure may increase chamber compliance of the left ventricle and left atrium and, in turn, increase peak early and late diastolic filling velocities through an increment in forward transmitral pressure gradient. Increased pericardial constraining force is a possible cause limiting left ventricular filling and hence cardiac output in heart failure.

Acute Disease↗

PubMedDate not supplied

Peak early diastolic filling velocity may decrease with preload augmentation: effect of concomitant increase in the rate of left atrial pressure drop in early diastole.

Doppler-determined transmitral flow velocity pattern has been shown to depend on transmitral pressure gradient, and left atrial (LA) pressure has been considered to be important in determining transmitral pressure gradient in early diastole and peak early diastolic filling velocity (E). In recent studies in human beings, however, it was proved that E did not necessarily change with LA pressure. This may be because concomitant changes in other factors masked the effect of LA pressure. To investigate the relation between transmitral flow velocity pattern and hemodynamic parameters during preload intervention over the wide range of LA pressure, pulsed Doppler transmitral flow velocity pattern and high-fidelity LA and left ventricular (LV) pressures were simultaneously recorded during rapid volume loading to the LA. Data at three stages, at control, at moderate volume loading (the median LA-to-LV crossover pressure during the volume loading), and at advanced volume loading (the maximal crossover pressure during the volume loading), were compared with one another in 11 dogs. A mean value of E increased with the crossover pressure up to moderate volume loading but did not further increase at advanced volume loading. In the data pooled from all experimental stages in all dogs, the changes in E did not correlate with those in the crossover pressure, but correlated weakly with those in the difference between the crossover pressure and LV minimum pressure (r = 0.45, p < 0.05). E decreased at advanced volume loading in three of 11 dogs with a steep LA pressure drop in early diastole although the changes in the difference between the crossover pressure and LV minimum pressure in the three dogs were similar to those in the other eight dogs. The changes in a rate of LA pressure drop in early diastole associated with advanced volume loading inversely correlated with those in E (r = -0.79, p < 0.01). Thus, E may decrease with an extreme increase in LA pressure; this change may be due to an associated increase in a rate of LA pressure drop in early diastole. This finding suggests that at high LA pressure the increased rate of LA pressure drop in early diastole appears to decrease LV filling and hence to reduce stroke volume.

Animals↗

PubMedDate not supplied

Dobutamine stress ultrasonic myocardial tissue characterization in patients with dilated cardiomyopathy.

Although acoustic properties of the myocardium are different between patients with cardiomyopathy and normal subjects, the frequency of the abnormal properties in patients with cardiomyopathy is unknown. We assessed the frequency of abnormal acoustic properties of the myocardium detectable with integrated backscatter in patients with cardiomyopathy and attempted more sensitive ultrasonic tissue characterization by combining dobutamine stress testing in patients with cardiomyopathy with apparently normal acoustic properties of the myocardium at rest. The magnitude of cyclic variation of integrated backscatter and calibrated myocardial integrated backscatter at end diastole were measured in 36 normal subjects and 40 patients with dilated cardiomyopathy. Either one of the integrated backscatter parameters was abnormal in 30 of 40 patients with cardiomyopathy. Dobutamine stress ultrasonic tissue characterization was performed in 10 patients with cardiomyopathy with normal values of both integrated backscatter parameters and 10 normal subjects. Calibrated myocardial integrated backscatter did not change during dobutamine infusion in any subject. The magnitude of cyclic variation in integrated backscatter increased in normal subjects but did not change in patients with cardiomyopathy despite a comparative associated increase in the systolic wall thickening during dobutamine infusion. Abnormal acoustic properties are detectable at rest with myocardial integrated backscatter about in three quarters of patients with cardiomyopathy. A combination of dobutamine stress testing would provide more sensitive ultrasonic myocardial tissue characterization and may make it possible to detect subtle changes in the acoustic properties of the myocardium in patients with dilated cardiomyopathy. Therefore dobutamine stress ultrasonic tissue characterization may detect mild dilated cardiomyopathy.

Adult↗

PubMedDate not supplied

Effect of nicardipine versus enalapril on plasma endothelin-1 in hypertensive patients with type 2 diabetes mellitus.

We compared the effects of dihydropyridine type Ca channel blocker slow-release nicardipine and angiotensin converting enzyme inhibitor enalapril on plasma endothelin-1 (ET-1) levels in hypertensive type 2 diabetic patients (n=20). Nicardipine or enalapril was administered for 6 months by a crossover design. Nicardipine and enalapril comparably lowered blood pressure. Enalapril significantly reduced urinary albumin excretion in microalbuminuric patients, whereas nicardipine did not. Urinary beta2-microglobulin excretion was significantly increased during nicardipine treatment. However, both drugs significantly reduced plasma ET-1 as compared with pretreatment levels, close to that in healthy control (2.9 +/- 0.3 pg/ml in control, 4.8 +/- 0.3 pg/ml before treatment, 3.2 +/- 0.3 pg/ml during nicardipine vs before treatment p<0.05, 2.9 +/- 0.4 pg/ml during enalapril vs before treatment p<0.01). The decrease in plasma ET-1 was significantly correlated with the increase in natriuresis in normoalbuminuric patients treated with enalapril ( r= -0.82, p<0.01) but not in those treated with nicardipine. Although nicardipine and enalapril had different renal effects, both drugs equally suppressed plasma ET-1 levels in hypertensive patients with type 2 diabetes.

Albuminuria↗