Showing papers by "Kumamoto University published in 1982"

Purification and Characterization of a Ca2+- and Calmodulin-Dependent Protein Kinase from Rat Brain

Abstract: A Ca2+- and calmodulin-dependent protein kinase was purified from rat brain cytosol fraction to apparent homogeneity at approximately 800-fold and with a 5% yield. The purified enzyme had a molecular weight of 640,000 as determined by gel filtration analysis on Sephacryl S-300 and a sedimentation coefficient of 15.3 S by sucrose density gradient centrifugation, and resulted in a single protein band of MW 49,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These results suggest that the native enzyme has a large molecular weight and consists of 11 to 14 identical subunits. The purified enzyme exhibited Km values of 109 and 30 microM for ATP and chicken gizzard myosin light chain, respectively, and Ka values of 12 nM and 1.9 microM for brain calmodulin and Ca2+, respectively. In addition to myosin light chain, myelin basic protein, casein, arginine-rich histone, microtubule protein, and synaptosomal proteins were phosphorylated by the enzyme in a CA2+- and calmodulin-dependent manner. The purified enzyme was phosphorylated without the addition of the catalytic subunits of cyclic AMP-dependent protein kinase. Our findings indicate that there is a multifunctional Ca2+- and calmodulin-dependent protein kinase in the brain and that this enzyme may regulate the reactions of various endogenous proteins.

The relationship between senile plaques and cerebral blood vessels in Alzheimer's disease and senile dementia. Morphological mechanism of senile plaque production.

Abstract: Several kinds of senile plaque found in 6 brains (4 from patients with Alzheimer's disease and 2 from patients with senile dementia) were examined in serial sections by light electron microscopy. The results obtained were as follows. All the senile plaques contained at least some amyloid fibrils, and these seemed to be produced at the basement membranes of capillary endothelial cells and projected into the surrounding parenchyma. Even when the senile plaques themselves appeared to lack amyloid fibrils by light microscopy, at least one degenerable capillary containing amyloid fibrils was demonstrable when serial sections were examined ultrastructurally. The findings described above suggest that the amyloid fibrils which form the cores of the several kinds of senile plaque, seem to be produced at the basement membrane of the endothelial cell. It is speculated that the capillary degeneration with the formation of amyloid fibrils may be primary change in the genesis of senile plaques.

Synthesis of Benzoxazoles, Benzothiazoles and Benzimidazoles and Evaluation of Their Antifungal, Insecticidal and Herbicidal Activities

Abstract: Benzoxazoles, benzothiazoles and benzimidazoles having substituents on the azole and benzene nuclei were synthesized evaluated for antifungal, insecticidal and herbicidal activities. It was found that benzimidazoles tended to exhibit antifungal activity while benzothiazoles tended to show herbicidal activity. Chloro, trifluoromethyl, methoxy and ethoxy groups at the 5 position were potent substituents, and the 2-pyridyl group at the 2 position is a common structural unit. Among several active derivatives, 7-chloro-2-(2-pyridyl) benzimidazole and 2-(2-pyridyl)-5-trifluoromethylbenzothiazole exhibited significant activity against Panonycus citri.

Preparation and evaluation in vitro and in vivo of polylactic acid microspheres containing doxorubicin.

Abstract: DL-Polylactic acid microspheres containing dibucaine were prepared, and release patterns of dibucaine from the microspheres as well as the local anesthetic effects of the drug in the microspheres were examined. The influences of nonsolvents and dibucaine concentration at preparation on the characteristics and dibucaine contents of the microspheres were investigated. Higher pH in nonsolvents and higher dibucaine concentration at preparation resulted in increased dibucaine contents in the microspheres. The release patterns of dibucaine from microspheres varied significantly among microspheres with different dibucaine contents, and the release mechanisms of dibucaine from microspheres were greatly influenced by disintegration of the microspheres. The local anesthetic effects of 0.1 ml aliquots of dibucaine hydrochloride solutions were examined at four concentrations in vivo. Then, the local anesthetic effects of dibucaine in microspheres were compared with those of the above solutions in relation to the release rate of dibucaine from microspheres in vitro. The more the amount of dibucaine, the stronger was the local anesthetic effect. The local anesthetic effect of dibucaine in microspheres lasted much longer (300 h) than that of dibucaine hydrochloride solutions. The profile of local anesthetic effect of dibucaine in microspheres in vivo was different from the release profile in vitro.

Human Erythrocyte Prolidase and Prolidase Deficiency

Abstract: Summary Case. A female patient with prolidase deficiency was examined. Biochemical studies on human prolidase (EC 3.4.13.9) and prolidase deficiency are described. The urine sample from a 32year-old female with prolidase deficiency was examined. Diagnosis was based on clinical features and defects of prolidase in her erythrocytes. She excreted massive amounts of iminopeptides, where three major peptides were identified; aspartyl-proline, glutamyl-proline and glycyl-proline. The prolidase was purified ap proximately 10,000-fold from the normal human erythrocytes through an eight step procedure. The purified enzyme consisted of two identical subunits of which the molecular weight was calculated to be 55.000. The relative cleavage rates of the enzvme for glycyl-L-proline, L-alanyl- prol line, L-leucyl-L-proline, L-prolyl-Lproline, and glycyl-hydroxy-L-proline were 100%, 53%, 27%, 31% and 2%. respectively. he relative substrate specificity of the enzyme offers a reasonable explanation for the presence of a higher level of urinary imidodipeptides in a patient with prolidase deficiency. An attempt at erythrocyte transfusion was performed, aimed at enzyme replacement therapy. After the transfusion (erythrocytes from 800 ml of whole blood), the prolidase activity of the peripheral erythrocyte was elevated to approximately 35% of the normal values and gradually decresed (half-life, 41 days). During this period urinary peptide-bound proline was monitored, but no significant change was observed. Speculation The prolidase activity of transfused erythrocytes is relatively stable; however, intracellular enzyme activity has no effect on net proline loss and skin lesions. Enzyme replacement may be a possible attempt at therapy of the disease, but other forms of replacement should be tried.

The Dopamine Receptor in the Intermediate Lobe of the Rat Pituitary Gland

Abstract: Dopamine is a neurotransmitter in the intermediate lobe (IL) of the rat pituitary gland. The presence of a dopamine receptor can be inferred from the ability of dopamine to diminish the basal or L-isoproterenol-stimulated release of alpha-melanocyte stimulating hormone (alpha-MSH) or to diminish the synthesis of adenosine 3′,5′ monophosphate (cAMP). Based on physiological and biochemical studies the IL dopamine receptor is designated as a D-2 dopamine receptor in the classification schema of Kebabian and Calne (1979) .

Optimisation of discrete-type load-frequency regulators considering generation-rate constraints

Abstract: In the paper a method of designing discrete-type load-frequency regulators of a two-area reheat-type thermal system with generation-rate constraints is presented. The construction of the regulators is based on the conventional tie-line bias control. The regulator parameters are optimised by minimisirig a discrete-type quadratic performance index with a term for presenting a generation-rate constraint. The optimisation of the parameters is achieved by using a Newton-Raphson iterative algorithm. The control effects by the proposed regulators are examined by digital simulations of the system. Furthermore, a suitable means for preventing excessive and unnecessary control action is also proposed considering the constraint.

Design of decentralised load-frequency regulators for interconnected power systems

Abstract: The paper presents a new method of designing decentralised load-frequency regulators for interconnected power systems. Within the framework of this method, the interconnected mutliarea power system is decomposed into several subsystems, each of which is controlled separately by a decentralised regulator. Each subsystem consists of one area and its external equivalent in a simplified form. A decentralised control law for the study area is introduced by using a quadratic performance index. Feedback gains of the decentralised regulator, which minimise the index, are determined by a Newton-Raphson iterative algorithm. The proposed method is applied to an interconnected longitudinal 4-area system, and the effects of the proposed regulator are examined by digital simulations and associated sensitivity analysis of the system. Furthermore, a suitable means for preventing excessive control action is also considered involving a significant system nonlinearity, i.e. some generation rate constraint.

Synthesis and Properties of 4-Diazomethyl-7-methoxycoumarin as a New Fluorescent Labeling Reagent for Alcohols and Carboxylic Acids

Abstract: 4-Diazomethyl-7-methoxycoumarin (DMC) was synthesized as a new fluorescent labeling reagent for hydroxyl and carboxyl compounds for use in high-performance liquid chromatography. Treatment of 4-formyl-7-methoxycoumarin with p-toluenesulfonohydrazide gave the tosylhydrazone in 78% yield, and this was derived to DMC in 78% yield by treatment with 0.2N NaOH. DMC was practically nonfluorescent in solution and possessed excellent storage properties. DMC reacted with alcohols in dichloromethane at room temperature in the presence of HBF4 as a catalyst to give the corresponding fluorescent coumarin ethers and also reacted with carboxylic acids in acetonitrile on heating to give the esters. DMC derivatives of cholestanol and cholesterol were satisfactorily separated by high-performance liquid chromatography. The present studies indicate that DMC is a useful fluorescent reagent for the labeling of alcohols and carboxylic acids.

Further analysis of inhibitory effects of propranolol and local anaesthetics on the calcium current in Helix neurones.

Abstract: 1 The effects of propranolol and local anaesthetics on Ca2+ current (Ica), individually separated from other ionic currents, in Helix neurones were studied under voltage clamp, using a suction pipette technique. 2 Increases in external Ca2+ concentrations overcame the inhibitory action of propranolol on Ica. Double reciprocal plots for peak Ica versus external Ca2+ concentrations in the presence or absence of propranolol did not intersect at the ordinate. 3 Internal application of propranolol (10-4M) inhibited Ica to about 40-60% of the control in a time-dependent manner. 4 Lignocaine and procaine at concentrations of 10-3-10-2 M inhibited Ica without shifting the threshold in the I-V relationships. Internal application of lignocaine (10-3-10-2M) also inhibited Ica: the ratio of depression of the Ica was almost equivalent to that of the agent applied externally. 5 The results provide evidence that propranolol inhibits Ica in the noncompetitive manner with Ca2+ at the cell membrane, and suggest that the agents may occupy the receptor site in the Ca2+-channel somewhere between the outer surface and inner phase of the membrane.

Physiological characteristics of the solitario-parabrachial relay neurons with tongue afferent inputs in rats.

Abstract: Among 180 units in the solitary tract nucleus (NTS) of rats, 34 solitario-parabrachial relay neurons (SP neurons), were identified by antidromic activation from the parabrachial nucleus. The SP neurons were classified into two groups, fast and slow, according to their antidromic latencies. The responsiveness of the SP in comparison with non-SP neurons was studied by electrical stimulation of three tongue nerves: the lingual (L), chorda tympani (CT) and glossopharyngeal (G) nerves. About half the SP neurons produced a single spike with an orthodromic latency of 2–5 ms, while about one third of them discharged more than two spikes. A few neurons gave rise to a long-lasting discharge consisting of five or six spikes. Some SP neurons were excited by stimulation of the tongue afferents with a low stimulus intensity, but other SP neurons produced spikes at only very high voltages. Fast SP cells were not differentiated from slow SP cells, except that latency of orthodromic responses to CT stimulation was significantly shorter in the former than in the latter (P<0.05, Mann-Whitney U-test). Locations of the SP and non-SP neurons, reconstructed histologically, indicate that they do not distribute evenly throughout the mediolateral extent at the rostral pole of the NTS, but clustered in its medial half.

Zinc and copper contents in breast milk of Japanese women

Abstract: Longitudinal studies of zinc and copper contents of breast milk were performed in 65 Japanese mothers during their lactation period until 5 months postpartum. They experienced no problems during pregnancy and at delivery. All women had full term healthy infants. Forty-five were multiparous and 20 primiparous. The highest level of zinc content was found in the colostrum, and subsequently the levels declined (p less than 0.005), as lactation progressed. Copper content was stable during the first month of lactation, and then declined gradually (p less than 0.005). Both mineral levels in breast milk ranged with great variance among the subjects at any stages of lactation. Significantly lower zinc level (p less than 0.01) and higher copper level (p less than 0.005) were found in the serum of lactating women three months postpartum, when compared with non-lactating control women. Factors including lactation history, age of lactating women, serum levels and contents of zinc and copper in the hair did not affect the contents of these trace minerals in the breast milk.

Studies on poisonous metals. IX. Effects of dietary fibers on absorption of cadmium in rats.

Abstract: The effects of dietary fibers on the gastrointestinal absorption of cadmium were studied. The fibers, such as lignin, cellulose, and sodium carboxymethylcellulose (Na CMC), produced a slight decrease in the contents of cadmium in the tissues of rats following a single oral administration of cadmium. In addition, in rats fed continuously with the experimental diets containing cadmium together with fibers, lignin and Na CMC significantly decreased the contents of cadmium in the tissues. These results show that the fibers depressed the intestinal absorption of cadmium. It is suggested that the inhibitory effects of these fibers on the gastrointestinal absorption of cadmium were due to their intrinsic properties, such as ability to bind cadmium and effect on viscosity, and that the differences of inhibitory effects of the fibers on the intestinal absorption of cadmium were related to differences in such physical properties.

Characterization of basal cell epithelioma in the Japanese.

Abstract: Clinicopathological and statistical studies were made on one hundred and fifty-six Japanese cases of basal cell epithelioma. A new formula was presented for the calculation of the growth rate of BCE, as follows, Size of tumor (major axis × minor axis) (mm2) = 1.7363 × T2 + 133.1192 T = Years which had elapsed from the presumed time of onset to the time of the first examination The yearly growth in diameter calculated from the above formula is less than 1.0 mm within ten years after onset, and 1.0 mm to 1.3 mm thereafter. The time needed for BCE to double its size averaged 8.8 years in the present study. In addition to the slow rate of progression, BCE in the Japanese was characterized by its pigmentation; 84% were either partially or totally pigmented, giving the BCE the so-called black and pearly appearance.

Treatment of carcinoma of the gallbladder in Japan

Abstract: Clinical records of 2567 patients (1717 female and 850 male) with primary carcinoma of the gallbladder in Japan, during the past 19 years, were collected by questionnaires sent out to the main 100 surgical institutes in Japan. Eighty-seven per cent of the patients were over fifty years of age. Gallstones were found in 58.8% of 1496 patients. About 50% of the patients with gallstones had cholesterol group stones. Of the 2269 patients who underwent surgical interventions, radical operations were performed in 467 patients (20.6%). Of the 467 patients, a correct preoperative diagnosis was made in only 77 patients (16.3%). Patients in Nevin's Stages I and II, whose lesions had been confined to the muscle layer, showed a good survival rate with only simple cholecystectomy or extended cholecystectomy, while the patients in Stages III, IV and V, whose lesions had spread beyond the muscle layer, showed poor results even with more aggressive surgical approaches. The poor prognosis of the lesions in Stages III, IV and V might be due to inappropriate aggressive procedures. For the lesions involving all the layers of the gallbladder wall, radical resection, such as extended cholecystectomy plus pancreatoduodenectomy, or extended right lobectomy plus pancreatoduodenectomy, might be recommended.

New Fluorescence Probes for Drug-Albumin Intereaction Studies

Abstract: Some new and known coumarincarboxylic acids were synthesized for use as fluorescence probes. The fluorescence properties of these compounds were examined in various solvents and albumin solutions. The fluorescence of 7-anilinocoumarin-4-acetic acid (I) was significantly enhanced in nonpolar solvents and in the presence of human serum albumin (HSA). The binding parameters of I were estimated from the fluorescence enhancement and spectral change of I bound to HSA. The Scatchard plots and the continuous variation plots indicated that only the primary site was capable of enhancing the fluorescence and causing the spectral changes of I. Digitoxin displaced I from its primary site on HSA, but site 1 and site 2 drugs on the basis of Sudlow's classification did not significantly displace the probe. The present data indicate that I may be useful as a third site marker.