Electron spin resonance studies on the lipoxygenase reaction by spin trapping and spin labelling methods.

The rate of oxygenation and that of trapping linoleic acid free radicals in the lipoxygenase [EC 1.13.11.12] reaction were measured in the presence of linoleic acid, oxygen, and nitrosobenzene at various concentrations, with a Clark oxygen electrode and ESR spectroscopy. The results were interpreted under the assumption that the free radical of linoleic acid, an intermediate of the lipoxygenase reaction, reacts competitively with oxygen or nitrosobenzene. The oxidation of the iron in the active site of lipoxygenase caused by the spin label reagent, 2-(10-carboxydecyl)-2-hexyl-4,4-dimethyl-3-oxazolidinyloxyl, was also observed by ESR- and fluorescence-spectroscopy.     

Fluroescence properties of chlorophyll a and b monomolecular films at the air-water interface.

The fluorescence properties of chlorophyll a and b monomolecular films at the air-water interface were measured by a high sensitivity fluorophotometer using the photon-counting method. The fluorescence intensity of chlorophyll molecules in monomolecular films in the absence of any diluents did not decrease simply with the mean distance of chlorphyll molecules. Over the range of the mean distances from 27 to 21 A, three fluorescence components (peaks at 685, 695 and 715 nm) of chlorophyll a were observed. In the case of chlorophyll beta, two fluorescence components (peaks at 667 and 685 nm) were observed over the range of the mean distances from 34 to 24 A. When the mean distance was 18 A, the short wavelength component of chlorophyll beta disappeared, and only the long avelength component was observed.     

Acetylcholine receptor-controlled ion flux in electroplax membrane vesicles: A minimal mechanism based on rate measurements in the millisecond to minute time region

The dependence of acetylcholine receptor-controlled transmembrane ion flux on carbamylcholine concentration was measured in the msec time region, using membrane vesicles and a quench flow technique. 4 Measurements were made: (1) transmembrane ion influx, (2) rate of inactivation of the receptor by carbamylcholine, (3) rate of recovery, and (4) ion influx mediated by “inactivated” receptor. The minimal model, based on the measurements, accounts for the time dependence of receptor-controlled ion flux over a 200-fold carbamylcholine concentration range. The maximum flux rate of 84 sec^?1 indicates that we have succeeded in measuring the receptor-controlled processes which give rise to electrical signals in cells.     

Establishment of Alternative Culture Method for Spermatogonial Stem Cells Using Knockout Serum Replacement

Since spermatogonial stem cells (SSCs) are capable of both self-renewal and differentiation to daughter cells for subsequent spermatogenesis, the development of an efficient in vitro culture system is essential for studies related to spermatogenesis. Although the currently available system is serum-free and contains only chemically-defined components, it highly relies upon bovine serum albumin (BSA), a component with batch-to-batch quality variations similar to those of fetal bovine serum. Thus, we searched for an alternative BSA-free culture system that preserved the properties of SSCs. In this study, we utilized Knockout Serum Replacement (KSR) in the SSC culture medium, as a substitute for BSA. The results demonstrated that KSR supported the continuous growth of SSCs in vitro and the SSC activity in vivo without BSA, in a feeder-cell combination with mouse embryonic fibroblasts. The addition of BSA to KSR further facilitated cell cycle progression, whereas a transplantation assay revealed that the addition of BSA did not affect the number of SSCs in vivo. The combination of KSR with BSA also allowed the elimination of GFRA1 and FGF2, and the reduction of the GDNF concentration from 20 ng/ml to 5 ng/ml, while maintaining the growth rate and the expression of SSC markers. Furthermore, KSR was also useful with SSCs from non-DBA/2 strains, such as C57BL/6 and ICR. These results suggested that KSR is an effective substitute for BSA for long-term in vitro cultures of SSCs. Therefore, this method is practical for various studies related to SSCs, including spermatogenesis and germ stem cell biology.     

Molecular phylogeny of the yellow peach moth,Conogethes punctiferalis (Lepidoptera: Crambidae): distribution of two genetic lineages across Japan

Mating disruption using synthetic sex pheromones is an effective pest-control method to prevent the evolution of insecticide resistance. Although the sex pheromone of the yellow peach moth Conogethes punctiferalis (Guenée) (Lepidoptera, Crambidae) has already been identified and artificially synthesized, mating disruption in the field has been less successful than expected. Such ineffectiveness may be caused by genetic variation in the attractiveness of the sex pheromone across moth populations. To evaluate this possibility, we evaluated the genetic population structure of this species across 10 populations in Japan. We found that two clades were present at the mtDNA COI region, each clade was sympatrically distributed, and no genetic structure was detected among the populations. In addition, the frequencies of the two clades did not differ between the samples in pheromone traps using two types of blend and those hand collected from infested fruit. Our mating experiment revealed random mating between the two clades. Our results strongly suggest that the genetic variation in the attractiveness of the sex pheromone is minimal and symmetrical gene flow likely occurs between the clades in the field.

     

The gender differences in health effects of environmental cadmium exposure and potential mechanisms

On a viewpoint of gender differences in Cd body burden and its health effects, we reviewed the population-based research including our own which conducted in Japan, Thailand, Australia, Poland, Belgium and Sweden to assess health effects of human exposure to environmental cadmium and their potential mechanisms. As a result, six risk factors in Cd health effects in women have been identified; (1) more serious type of renal tubular dysfunction, (2) difference in calcium metabolism and its regulatory hormones, (3) kidney sensitivity; difference in P450 phenotype, (4) pregnancy, (5) body iron store status, and (6) genetic factors. Further studies of Cd toxicity targeted to women would now appear necessary.