Characterization and epitope mapping of monoclonal antibodies to the nucleocapsid protein of severe acute respiratory syndrome coronavirus

The sudden emergence of severe acute respiratory syndrome (SARS) at the end of 2002 resulted in 774 reported deaths from more than 8000 cases worldwide. As no effective vaccines or antiviral agents are available, the most effective measure to prevent the expansion of a SARS epidemic is the rapid diagnosis and isolation of SARS patients. To establish specific diagnostic methods, we generated nine clones of monoclonal antibodies to nucleocapsid protein (NP) of SARS-coronavirus (SARS-CoV). On immunofluorescent antibody assay and Western blotting analysis, none of the monoclonal antibodies showed cross-reactivity to authentic and recombinant NPs of human coronavirus (HCoV) 229E strain. To determine the region on the NP molecule where the monoclonal antibodies bind, we generated four truncated recombinant NPs and analyzed the reactivity between monoclonal antibodies and truncated NPs. Two monoclonal antibodies reacted with a truncated NP covering from amino acid residues 111 to 230, and seven reacted with another truncated NP covering from amino acid residues 221 to 340. Epitope mapping analysis indicated that monoclonal antibody SN5-25 recognized the amino acid sequence Q(245)TVTKK(250) On SARS-NP. Within the epitope, Q245, T246, V247, K249, and K250 appeared to form an essential motif for monoclonal antibody SN5-25 to bind. The information about binding sites and epitopes of monoclonal antibodies may be useful for the development of new diagnostic methods for SARS and for analyzing the function of N protein of SARS-CoV.     

Follicle-stimulating hormone (FSH) stimulates the expression of Pin1, a peptidyl-prolyl isomerase, in the bovine granulosa cells

A peptidyl-prolyl isomerase, Pin 1, has been shown to play a role in the regulation of cell cycle progression, both in vitro and in vivo. However, the involvement of Pin 1 during follicular development is not well understood. The aim of this study was first to investigate the expression of Pin 1 mRNA in the granulosa and theca cells of the follicle at different developmental stages of follicles in the bovine ovary, and second, to examine the effects of follicle-stimulating hormone (FSH) and estradiol (E2) on the expression of Pin 1 in the cultured bovine granulosa cells. Follicles were classified into four groups based on the diameter (dominant follicles >8.5mm in diameter, subordinate follicles <8.5mm in diameter) and the relative levels of E2 and progesterone (P4) (E2:P4>1, estrogen active; E2:P4<1, estrogen inactive): i.e. preovulatory dominant follicles (POFs); E2 active dominant follicles (EADs); E2 inactive dominant follicles (EIDs); small follicles (SFs). The expression of the Pin 1 gene was significantly increased in the granulosa cells of EADs as compared with those of other follicles, whereas its expression in theca cells did not differ among follicles at different developmental stages. The concentration of 5 ng/ml FSH alone and the combination of 1 ng/ml E2 and 5 ng/ml FSH stimulated the expression of the Pin 1 gene in bovine granulosa cells. Our data provide the first evidence that Pin 1 expression in the granulosa cells but not the theca cells changes during follicular development, and that FSH stimulate the expression of the Pin 1 gene. These results suggest that Pin 1 regulates the timing of cell proliferation and may act as an intracellular signal responder in the granulosa cells during bovine follicle development.     

Abstracts of Nippon Dojo-Hiryogaku Zasshi (Japanese Journal of Soil Science and Plant Nutrition)

The electrokinetic behavior of colloidal particles in three waterlogged soils at 38°C was investigated with reference to the stability changes of soil colloidal suspensions under reductive conditions. The dispersed clay particles of the three soils exhibited a negative zeta (ζ) potential. The absolute value of the ζ-potential, |-ζ|, of these soils in the earlier period of waterlogging decreased, which caused the flocculation of clay particles. The concentrations of divalent cations, i.e., Fe²⁺ and Ca²⁺ in the soil solutions were estimated to be higher than their critical flocculation concentrations (CFCs) on the basis of the observed CFCs of Fe²⁺ and Ca²⁺ for the clay suspension of halloysite as a reference. With the progression of the reduction process, clay particles of one soil still exhibited a low |- ζ| and flocculated. The concentrations of Fe²⁺ and Ca²⁺ in the soil solutions were estimated to be higher than their CFCs, respectively. The clay particles of two sandy soils, however, showed an increase in |- ζ| due to the increase in pH and dispersed. The concentrations of Fe²⁺ and Ca²⁺ in the soil solutions were estimated to be lower than their CFCs, respectively. The stability changes of the soil colloidal suspensions by these divalent cations under sequential soil-reduction can be explained by the alteration of the Stern potential (- ψ _s ), which determines the repulsion energy related to the potential energy of interaction between two particles. The apparent decrease in the Ca²⁺ concentration of the soil solutions in the later period of waterlogging was explained largely by the re-adsorption of water-soluble Ca²⁺ on the exchange sites of soil clays with the decrease in the Fe²⁺ concentration in the soil solution.     

Comparative study of anti-oncogenic microRNA-145 in canine and human malignant melanoma

MicroRNA-145 (miRNA-145; miR-145) is aberrantly expressed in most of human cancers and plays a significant role in carcinogenesis and cancer progression. In the current study, we focused on how miR-145 plays a role in canine and human malignant melanomas. MiR-145 was significantly downregulated in canine malignant melanoma tissues and canine melanoma cell lines, as well as human melanoma cell lines tested. The ectopic expression of miR-145 showed a significant growth inhibition in both canine and human melanoma cells tested, and the effect was achieved partly by suppressing c-MYC in canine melanoma LMeC and in human melanoma A2058 and Mewo cells. At the same time, a suppressive tendency on cell migration in canine melanoma KMeC cells and significant suppression of cell migration in human melanoma A2058 cells by suppressing FASCIN1 were also found. These findings suggest that miR-145 acts as a tumor suppressor in both canine and human malignant melanomas.     

Dried-bonito aroma components enhance salivary hemodynamic responses to broth tastes detected by near-infrared spectroscopy

To elucidate the effects of aroma from dried bonito (katsuo-bushi) on broth tastes caused by the central integration of flavor, optical imaging of salivary hemodynamic responses was conducted using near-infrared spectroscopy (NIRS). A reconstituted dried bonito flavored broth produced a significantly larger hemodynamic response than the odorless broth taste solutions for 5 of the 10 panelists, who felt that the combination of the aroma with the tastes was congruent. In the remaining 5 panelists who felt the combination incongruent, the flavored broth did not cause the enhancement of response. Moreover, when the odor-active smoky parts were removed from the flavoring, the reconstituted flavoring did not enhance the response in the former five panelists. These results indicate that NIRS offers a sensitive method to detect the effect of specific congruent aroma components from dried-bonito broth on the taste-related salivary hemodynamic responses, dependent on the perceptual experience of the combination of aromas and tastes.     

Alternaria host-selective toxins: determinant factors of plant disease

Seven diseases caused by pathotypes of Alternaria alternata, which produced host-selective toxins (HSTs), a diverse group of low-molecular-weight secondary metabolites, are known, and each HST has an essential role as a determinant of pathogenicity in all interactions between the plant host and A. alternata. Although these HST-producing pathotypes are morphologically indistinguishable, each has a distinct host range and can be distinguished by its specificity on the respective host plant, hence their designation as pathotypes of A. alternata. In 1933, Tanaka made the first discovery of a HST; fungus-free culture filtrates of A. kikuchiana (now called A. alternata Japanese pear pathotype) were toxic to susceptible cultivar Nijisseiki, but not to resistant cultivars. Over the 80 years since then, the structure of HST molecules, target sites and mode of actions of HSTs, and the molecular genetics of HST production regulating by supernumerary chromosomes encoding HST gene clusters have been studied extensively. We focus this review on studies of low-molecular-weight HSTs produced by A. alternata and give an overview of various types of HST studies.     

Expressions of lipid oxidation markers, N(ε)-hexanoyl lysine and acrolein in cisplatin-induced nephrotoxicity in rats

The purpose of this study was to evaluate whether N(ε)-hexanoyl lysine (N(ε)-HEL) and acrolein reflect the severity of cisplatin-induced nephrotoxicity. Immunoexpression of N(ε)-HEL and acrolein in kidneys and their urinary concentration were examined up to day 4 post-cisplatin injection in rats. Cisplatin-induced tubular injury was observed histopathologically on days 2-4 after injection and became more severe time-dependently. On days 2-4, N(ε)-HEL and acrolein were immunostained in the cytoplasm of damaged tubular cells. Their immunostaining intensity and urinary levels increased as tubular injury became more severe. These results suggest that expressions of N(ε)-HEL and acrolein were associated with the pathogenesis of cisplatin-induced nephrotoxicity.     

Larval survival,development and growth in the horsehair crab,Erimacrus isenbeckii,cultured under different photoperiod conditions

Larval sinking causes larval mass mortality during seed production in the horsehair crab, Erimacrus isenbeckii. Under normal light conditions, horsehair crab larvae generally show negative phototactic behaviour and sink in their rearing seawater. It has been proposed that culturing larvae in the dark may prevent larval sinking. Herein, we examined the effect of photoperiods on horsehair crab larval survival and development to facilitate the development of larval rearing techniques that prevent sinking. Newly hatched larvae were reared with Artemia to the first crab stage in 2‐L beakers under five photoperiods: 0L:24D, 6L:18D, 12L:12D, 18L:6D and 24L:0D. Larvae survived and molted to the first crab stage under all tested photoperiod conditions. The survival rate improved with increasing light period, whereas the developmental period for each larval stage decreased with increasing light period. Longer light periods increased the carapace length at the first crab stage. Our results suggest that larvae could be cultured to the first crab stage in large‐scale tanks under constant darkness. However, significantly improved larval performance under longer photoperiodic conditions indicates a need for developing alternative culture techniques to control larval behaviour in the seed production tank.     

Amelioration Effects of Pedogenetic Acidification Process on Physical Properties of Soils in Japanese Non-Volcanic Areas with Special Reference to Charge Characteristics

The charge characteristics and physical properties of acid soils were compared with those of reference clays to elucidate the effect of the pedogenetic acidification process on soil physicochemical properties. The soil physics such as clay dispersibility and aggregate stability of the acid soils are considered to be ameliorated by acidification process, which is attributable to the increase in amorphous Al hydroxides and interlayering of 2:1 clays. This means that the pedogenetic acidification process plays an important role in maintaining a desirable soil structure in Japanese non-volcanic areas from the aspect of environmental conservation. The involved mechanism was discussed with special reference to the modification of charge characteristics of clays and binding agents between soil particles.     

Spawning grounds and maturation status in adult flathead flounder Hippoglossoides dubius off Niigata Prefecture, Sea of Japan

Investigations were conducted on the flathead flouder Hippoglossoides dubius to analyze the environmental conditions around the spawning grounds and the maturational status of spawners in the Sea of Japan off Niigata. In both sexes, larger fish matured earlier than smaller individuals, and participated in breeding from the early part of the spawning season. Males were always the predominant sex found within the spawning ground, likely because males spent longer periods in the spawning grounds than females. This finding further suggests that males matured earlier and maintained sexual activity for longer periods than females within the spawning grounds. Submarine topography and water temperature were concluded to be important factors affecting the formation of the spawning grounds. The spawning ground was located within a restricted area where bathymetric lines curve towards a shallow area. Adult fish usually occur in the Japan Sea Proper Water (<1 °C), but spawners gathered just below the surface mixing layer where water temperature was 5–10°C. Hence, it was thought that the flounder spawn at the upper tolerable water temperature limit (i.e. the shallowest accessible depth) to release their offspring into the surface layer.