Effects of arildone on the immunogenicity of formalin-inactivated polioviruses

Concentrated and purified Sabin and virulent strains of poliovirus types 1, 2 and 3 were inactivated with formalin at 37 C. By addition of 5.4 microM arildone, an antiviral agent, to the virus suspension, the stability of D antigen increased in both Sabin and virulent strains of all types, especially in virulent type 1 Mahoney strain. The drug had neither any inhibitory nor enhancing effect on the formalin inactivation. When antibody response was compared in guinea pigs, Sabin strains inactivated in the absence of arildone were less immunogenic against homotypic virulent strains than inactivated vaccine prepared from virulent strains. On the other hand, Sabin strains inactivated in the presence of arildone were equally immunogenic. These results indicate that it is possible to prepare from Sabin strains a potent and safe inactivated vaccine having an immunogenicity comparable to that prepared from virulent strains.     

Properties of glutamate dehydrogenase purified from Bacteroides fragilis

The dual pyridine nucleotide-specific glutamate dehydrogenase [EC 1.4.1.3] was purified 37-fold from Bacteroides fragilis by ammonium sulfate fractionation, DEAE-Sephadex A-25 chromatography twice, and gel filtration on Sephacryl S-300. The enzyme had a molecular weight of approximately 300,000, and polymeric forms (molecular weights of 590,000 and 920,000) were observed in small amounts on polyacrylamide gel disc electrophoresis. The molecular weight of the subunit was 48,000. The isoelectric point of the enzyme was pH 5.1. This glutamate dehydrogenase utilized NAD(P)H and NAD(P)+ as coenzymes and showed maximal activities at pH 8.0 and 7.4 for the amination with NADPH and with NADH, respectively, and at pH 9.5 and 9.0 for the deamination with NADP+ and NAD+, respectively. The amination activity with NADPH was about 5-fold higher than that with NADH. The Lineweaver-Burk plot for ammonia showed two straight lines in the NADPH-dependent reactions. The values of Km for substrates were: 1.7 and 5.1 mM for ammonium chloride, 0.14 mM for 2-oxoglutarate, 0.013 mM for NADPH, 2.4 mM for L-glutamate, and 0.019 mM for NADP+ in NADP-linked reactions, and 4.9 mM for ammonium chloride, 7.1 mM for 2-oxoglutarate, 0.2 mM for NADH, 7.3 mM for L-glutamate, and 3.0 mM for NAD+ in NAD-linked reactions. 2-Oxoglutarate and L-glutamate caused substrate inhibition in the NADPH- and NADP+-dependent reactions, respectively, to some extent. NAD+- and NADH-dependent activities were inhibited by 50% by 0.1 M NaCl. Adenine nucleotides and dicarboxylic acids did not show remarkable effects on the enzyme activities.     

Changes in the Content of Phytochrome I and II Apoproteins in Embryonic Axes of Pea Seeds during Imbibition

The contents of phytochrome I and II in crude extracts fromembryonic axes of Pisum sativum cv. Alaska seeds were immunochemicallydetermined using purified pea phytochrome I and II as standards.We have produced and used three different types of mouse monoclonalanti-pea phytochrome antibodies (mAP) such as one reacting preferentiallywith phytochrome I, one with phytochrome II, and one with bothI and II. Phytochrome II was separated from I in the samplesusing immobilized column chromatography with mAPl. The amountsof two phytochrome species were quantitatively measured withwestern blotting and ELISA. Ca. 0.2 µg /axis of phytochromeI and ca. 0.05 µg /axis of phytochrome II were detectedby ELISA after imbibition for 12 h in the dark, though smallamounts of both were detected in dry axes. Ca. 0.05 µg/axis each of phytochrome I and II were detected by ELISA afterimbibition for 12 h in the light, and the results were confirmedby western blotting. This study showed that phytochrome II isnot green-tissue-specific, being also found in dark-imbibedembryonic axes, and that although light significantly lowersthe content of phytochrome I in the axis, it does not significantlyaffect that of phytochrome II. (Received June 10, 1987; Accepted August 27, 1987)     

Accurate Evaluation of 1,2-Diacylglycerol in Gerbil Forebrain Using HPLC and In Situ Freezing Technique

Gerbil forebrains were frozen in situ to inactivate the tissues, and 1,2-diacylglycerols were first measured quantitatively by HPLC. Although 1,2-diacylglycerols were completely recovered from the HPLC column, the control amount of 1,2-diacylglycerol in gerbil forebrain was only 79.6 nmol/g wet weight, which is about one-fourth of that previously reported for gerbil brain inactivated by liquid N2 after decapitation instead of in situ freezing. The fatty acid composition of 1,2-diacylglycerols in gerbil forebrain was first reported and the control 1,2-diacylglycerols were richer in palmitic acid than in stearic acid or arachidonic acid, which is rather different from the data previously reported for mouse or rat brain obtained by decapitation and analyzed by traditional TLC methods. The amount of 1,2-diacylglycerol increased by 82.9% in gerbil forebrain during 5 min of ischemia induced by bilateral carotid ligation. Arachidonic acid and stearic acid were abundant in the 1,2-diacylglycerols produced by 5 min of ischemia. Thus we were able to obtain accurate values of the amount and the fatty acid composition of 1,2-diacylglycerols in gerbil forebrains using HPLC and in situ freezing technique.     

Possible involvement of calmodulin and the cytoskeleton in electrofusion of plant protoplasts

Calmodulin antagonists, trifluoperazine, chlorpromazine, calmidazolium, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7), strongly inhibited the electrofusion of barley (Hordeum vulgare L. cv Moor) protoplasts with a marked increase of broken fusion products, after 60 minutes of incubation. W-5, a dechlorinated analog of W-7, was found less effective for the inhibition than W-7. Ethyleneglycol-bis(β- aminoethylether)-N,N′-tetraacetic acid a Ca²⁺ chelator, La³⁺, a surface Ca²⁺ antagonist, and verapamil, a Ca²⁺ channel blocker, also inhibited electrofusion. Dielectrophoresis was inhibited by La³⁺. A microtubule inhibitor, vinblastine, inhibited electrofusion strongly while colchicine, slightly. A microfilament inhibitor, cytochalasin B, promoted fused cells to become spherical while phalloidin did not affect electrofusion.     

Promotion of flower formation by atrazine and diuron in seedlings of Asparagus

Flower formation in 25-d-old seedling of Asparagus officinalis L was increased by soaking the seeds for 12 d in solutions of atrazine (2-chloro-4-ethylamino-6-isopropylamino-1,3,5-triazine) or diuron (3-(3,4-dichlorophenyl)-1,1-dimethylurea), two herbicides known to affect electron flow of photosystem II, from approx. 4% in controls to 37% in treatments with 400 M of atrazine or about 100 M diuron. Both herbicides inhibited plant growth, but treatment of seeds with other growth inhibitors (absicic acid, (2-chloroethyl)trimethylammonium chloride, polyethylene glycol) did not affect flower formation although inhibiting seed germination and plant growth. The increase of flower formation by the two herbicides permits identification of the sex of Asparagus plants at an early developmental stage.     

Genotypic variability for callus formation and plant regeneration in rice (Oryza sativa L.)

Summary Sixty rice varieties (Oryza sativa L.), belonging to three subspecies, japonica, indica and javanica (some japonicaXindica hybrids were included), were compared for their capacity for callus growth and plant regeneration. Tissue cultures initiated from mature seeds on Murashige and Skoog's (1962) medium with 2 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D) were transferred to a medium containing 0.02 mg/l 2,4-D and 10 mg/l kinetin, from which plantlets were regenerated. Large variabilities in callus growth and plant regeneration potentials were revealed among the varieties tested. Most japonica varieties formed a callus that weighed more than 100 mg per seed 30 days after inoculation, and showed a relatively high regenerative potential, whereas indica varieties, japonica-indica hybrids and javanica varieties showed poor callus growth and plant regeneration, although considerable varietal variation was observed in each subspecies. The callus growth potential was not correlated with the plant regeneration potential. Histological observations revealed that the epithelium cells of the scutellum mainly proliferated to form a callus, from which shoot and root primordia were differentiated independently from each other. The shoot primordia developed into plantlets when roots were formed adventitiously. In a few cases, shoots and roots were bilaterally initiated from a single primordium.     

Genetic polymorphism of coagulation factor XIII B subunit in the Japanese population: description of three new rare alleles

Polyacrylamide gel isoelectric focusing (PAGIEF) of neuraminidase-treated EDTA plasma samples followed by electroblotting with enzyme immunoassay was performed to further investigate coagulation factor XIII B subunit (FXIII B) polymorphism. In 435 Japanese subjects PAGIEF patterns of FXIII B were classified into five common and three rare allotypes. This suggested that the FXIII B*2 allele existed in the Japanese population in the same manner as in Caucasians. Three new rare allotypes were considered to be controlled by three rare alleles which were designated FXIII B*13, FXIII B*14, and FXIII B*15, respectively. The gene frequencies calculated from 435 Japanese subjects were FXIII B*1 = 0.2977, FXIII B*2 = 0.0184, FXIII B*3 = 0.6805, FXIII B*13 = 0.0011, FXIII B*14 = 0.0011, and FXIII B*15 = 0.0011, respectively.     

Effect of intracerebroventricular administration of rat calcitonin gene-related peptide (CGRP), human calcitonin and [Asu1,7]-eel calcitonin on gastric acid secretion in rats

The effect of intracerebroventricular injection of rat calcitonin gene-related peptide (CGRP), human calcitonin (CT) and [Asu1,7]-eel CT on the volume and acidity of gastric juice was examined in the pylorus-ligated male rats. These 3 peptides were effective in suppressing both the volume and acidity of secreted gastric juice. Their potency on a molar basis, however, was markedly different; [Asu1,7]-eel CT was most potent, followed by human CT and finally by rat CGRP. These finding suggest that CGRP could not substitute for [Asu1,7]-eel or human CT in exerting the suppressive effect of gastric acid secretion.     

Receptor dynamics and tyrosine aminotransferase induction during the course of chronic treatment of rats with glucocorticoid

The changes in the cytosol glucocorticoid receptor (GR) content during a long-term administration of a glucocorticoid were studied to examine the mechanism of the development of steroid hormone resistance. Dexamethasone (Dex) (0.2 microgram/ml and 2.0 micrograms/ml) was given to adrenalectomized rats, and the GR content was determined using the exchange assay 1, 10, 20 and 50 days after the start of administration. The activity of tyrosine aminotransferase (TAT) in the cytosol was also assayed as a measure of the biological responsiveness of these animals to the administered glucocorticoid. The dissociation constant (Kd) was elevated and the Bmaxs of the GR in the cytosol were decreased by the lower concentration of Dex. The Bmaxs decreased to 30% of the untreated controls within 24 h and this lower level was maintained as long as the hormone treatment continued. On the other hand, the cytosol obtained from animals treated with 2.0 micrograms/ml of Dex for 20-24 days did not show any measurable amount of binding to 3H-Dex. The activity of TAT was elevated 24 h after the administration of Dex but decreased gradually and steadily with time during the experimental period. To examine the biological potency of remaining GR in the liver cytosol, 2.0 micrograms/ml Dex was again administered after a long-term treatment. This treatment eliminated the remaining GR completely and induced TAT at almost the same rate as observed in the untreated control animals.(ABSTRACT TRUNCATED AT 250 WORDS)