Nonsteady motion in unloaded contractions of single frog cardiac cells.

We studied the mode of shortening of enzymatically isolated single frog cardiac cells with a high-speed videosystem to see whether or not shortening is smooth. The segmental shortening of the cell in response to electrical stimulation exhibited a clear pause following the initial shortening over a distance of approximately 11 nm/half-sarcomere. Several preparations showed a second pause following the initial one. Nonsteady motion with a pause lasted usually a few tens of milliseconds. The duration of nonsteady motion was shorter in cells with large velocities of steady shortening following the pause than those with smaller velocities.     

Sequential actions of glucose oxidase and peroxidase in molecular films assembled by layer-by-layer alternate adsorption

Molecular films of protein/polyion layers were assembled by means of alternate adsorption through electrostatic interaction. Glucose oxidase (GOD) and peroxidase (POD) were assembled in combination with sodium poly(styrenesulfonate) (PSS) and poly(ethyleneimine) (PEI), respectively. Enzyme activities of those films on specific substrates (glucose and H(2)O(2)) were examined by coloring reaction of dye DA67. A multienzyme film containing GOD layer and POD layer was prepared by alternate adsorption of POD/PSS followed by PEI/GOD. Sequential redox reaction of glucose/H(2)O(2)/DA67 was demonstrated successfully with this supramolecular system. (c) 1996 John Wiley & Sons, Inc.     

Environmental control for the large-scale production of plants through in vitro techniques

Leafy or chlorophyllous explants of a number of plant species currently micropropagated have been found to have high photosynthetic ability. Their growth and development have been promoted on sugar-free medium rather than on sugar-containing medium, provided that the environmental factors, such as CO₂ concentration, light intensity and relative humidity, are controlled for promoting photosynthesis and transpiration of explants/shoots/plantlets in vitro. Thus, environmental control is essential for promoting photosynthetic growth and development of in vitro plantlets. Several types of sugar-free (photoautotrophic) culture systems for large-scale micropropagation of plants have been developed. Advantages of sugar-free over conventional (heterotrophic or photomixotrophic) micropropagation systems are as follows: growth and development of plantlets in vitro are faster and more uniform, plantlets in vitro have less physiological and morphological disorders, biological contamination in vitro is less, plantlets have a higher percentage of survival during acclimatization ex vitro, and larger culture vessels could be used because of less biological contamination. Hence, production costs could be reduced and plant quality could be improved significantly with photoautotrophic micropropagation. Methods for the measurement and control of in vitro environments and the beneficial effects of environmental control on photosynthetic growth, development, and morphogenesis in large-scale production of micropropagated plantlets are presented. This revised version was published online in June 2006 with corrections to the Cover Date.     

Clinical effect of recombinant human granulocyte colony-stimulating factor (rhG-CSF) on various types of neutropenia including cyclic neutropenia

Recombinant human granulocyte colony-stimulating factor (rhG-CSF) was investigated for its clinical efficacy in the treatment of various types of neutropenia (3 cases with idiopathic neutropenia of suspected drug induction, 5 cases with idiopathic neutropenia of other origin, and 2 cases with cyclic neutropenia). Treatment with glycosylated rhG-CSF produced in the Chinese Hamster Ovary cells at dose levels of 2–5g/kg/day caused rapid increases of neutrophil counts associated with an improvement of the infection. In cyclic neutropenia patients, marked reduction in the duration of the neutropenic period was observed with rhG-CSF administration started before the period. Intercurrent stomatitis, which occurred in 1 patient, was markedly milder as compared to a previous episode which occurred during an untreated neutropenic period.The treatment of rhG-CSF was well tolerated and no adverse events were observed, nor was there any detectable anti-rhG-CSF antibody in any patients studied; hence the clinical use of rhG-CSF is considered to be safe.These results suggest beneficial effects of rhG-CSF on the recovery of neutrophil counts in cyclic and other types of idiopathic neutropenias, as well as for the treatment of neutropenia-associated infection.     

Rapid detection of a minute amount of lipopolysaccharide from Salmonella in large volumes by poly(ethyleneimine)-cloth enzyme immunoassay

Utilizing the rapidity in capturing Salmonella lipopolysaccharide (LPS) and the macroporous nature of poly(ethyleneimine) immobilized polyester cloth (PEI-cloth), a PEI-cloth disk set in a Swinnex cartridge joined at both ends with 10 ml plastic syringes was repeatedly passed through with an LPS sample in large volumes. The captured LPS on the disk were detected at up to 1 pg, which is equivalent to the amount produced of 16 cells, by cloth enzyme immunoassay.     

Cypris larvae (Cirripedia: Balanomorpha) display auto-fluorescence in nearly species-specific patterns

The identification of balanomorph larvae plays an important role in ecological study and for protection against biofouling. However, it is difficult to identify species of cyprids (settling larvae) of balanomorph barnacles, as they show remarkably similar morphology. Some authors have suggested distinguishing cyprids of different species by carapace length, pigmentation, and fine carapace detail. However, such criteria are only applicable to a narrow range of balanomorph species. Recently, we were have serendipitously found species-specific distribution of fluorescent substances in cultured cyprids obtained from adult balanomorph barnacles, collected near the coast of Japan. Fluorescent patterns (FPs) of cyprids from 11 species were classified into five major groups. Cyprids specimens collected from the field were estimated, based on the FPs and other morphological characteristics (pigmentation and carapace length), after which their species were identified using the following two criteria: the morphology of adults derived from the field cyprids that adhered to a culture dish, and cyprid 12S rRNA gene sequence. The results of species estimation by FPs largely corresponded to the correct species identification. Other FP groups were found in the field cyprids. This study of FPs should be helpful for identification of cyprid species.     

Removal of the side-chain glucose groups from schizophyllan improves the thermal stability of the polycytidylic acid complexes under the physiological conditions

Thermal stabilization of the complex between polycytidylic acid [poly(C)] and the modified schizophyllan (SPG) whose hydrophilic side-chain glucose groups are selectively removed utilizing mild Smith-degradation has been investigated. With the decrease in the side-chain glucose groups of schizophyllan, the complex with poly(C) can be considerably stabilized compared with unmodified SPG; for example, the T(m) value after the removal of the side-chain glucose groups from 33.3 (unmodified) to 1.0 is enhanced by 14 degrees C. In addition, the thermal stabilization effect is even operative under the physiological conditions ([NaCl] = 0.15 mol dm(-3)). This effect is exerted owing to the construction of the hydrophobic atmosphere around the complex. Although schizophyllan lost the side-chain glucose groups, it still kept the protection effect of the bound poly(C) chain against RNaseA-mediated hydrolysis as observed for unmodified schizophyllan. The assessment of the cytotoxicity for A375:human malignant melanoma, and HL60:human promyelocytic leukemia revealed that the modified schizophyllan scarcely increases the cytotoxicity. These results indicate that the present modification for schizophyllan is of great significance in a viewpoint to develop the practical gene carriers operative even under the physiological conditions.     

Lipid-binding form is a key conformation to induce a programmed cell death initiated in tobacco BY-2 cells by a proteinaceous elicitor of cryptogein

Cryptogein, a proteinaceous elicitor secreted by Phytophthora cryptogea , induces a remarkable hypersensitive cell death in tobacco cells. Two cryptogein mutants were analysed to characterize the induction mechanism of cell death; one was a newly synthesized mutant N93A whose 93rd Asn residue was changed to Ala, the other was K13V whose Lys at position 13 was replaced with Val. The effect of these mutations was evaluated in terms of extracellular alkalization, production of active oxygen species (AOS) and progression to death. The mutation N93A resulted in a reduction in activity to 71.0, 74.6 and 24.5% for original rates of extracellular alkalization, AOS production and cell death progression, respectively. In the case of the K13V mutation, these rates changed to 114, 3.38 and 7.40%, respectively. The lipid-binding activities of the mutants were analysed using fluorogenic lipid of dehydroergosterol. The results for N93A and K13V were 38.3 and 3.40% compared with the wild type, respectively. These findings indicate that the lipid-binding form was the only conformation to induce the production of AOS and programmed cell death in plants.