| Budget Amount *help |
¥16,000,000 (Direct Cost: ¥15,100,000、Indirect Cost: ¥900,000)
Fiscal Year 2007: ¥3,900,000 (Direct Cost: ¥3,000,000、Indirect Cost: ¥900,000)
Fiscal Year 2006: ¥5,000,000 (Direct Cost: ¥5,000,000)
Fiscal Year 2005: ¥7,100,000 (Direct Cost: ¥7,100,000)
|
|---|
| Research Abstract |
The authors investigated for the host defense mechanisms in bivalve molluscs, especially, phagocytosis and generation of the reactive oxygen intermediates (ROIs) by hemocytes, and determined activities of lysozyme and lectins in the three species of the genera Crassostrea, C. gigas, C. nippona, and C. sikamea. Oyster hemocytes were divided into two morphological types: granulocytes and agranulocytes. The ratios between the granulocytes and agranulocytes varied during the year, however, the number of agranulocytes was always greater than that of granulocytes; approximately 70% of the total cell population was composed of agranulocytes. Granulocytes were more active phagocytes against three species of bacteria tested. Yeast cells were also extensively phagocytized by the granulocytes, but the agranulocytes produced a little phagocytosis of yeast cells. Granulocytes produced large amount of superoxide anion after phorbol stimulation or zymosan phagocytosis, but agranulocytes did not or on
… More
ly slightly produced superoxide anion. A simple method called the phagocytic plaque assay was used to visually present and estimate the phagocytic ability of the hemocytes from the giant Pacific oyster, Crassostrea gigas, using adherent Staphylococcus aureus cells on a plastic dish.
Crassostrea gigas hemocytes might phagocytose yeast cells by recognizing and binding to polysaccharides on the cell wall surface. This study was conducted to determine the differences in phagocytic capacity of C. gigas hemocytes for heat-killed yeast, zymosan, and zymocel, which have similar in size and shape, but which differ in their compositions of polysaccharides on the particle surface. Both agranulocytes and granulocytes exerted strong phagocytic capacity for all tested particles. Especially, zymosan was greatly phagocytosed by some of the granulocytes. To determine whether the phagocytosis of these respective particles depended on their recognition by specific polysaccharide receptors on the hemocyte surface, C. gigas hemocytes were pretreated with soluble a-mannan or laminarin and made to phagocytose the three types of the particles. The percentage of phagocytic cells of laminarin-treated granulocytes decreased significantly for zymosan and zymocel, but not for yeast. These results suggest that C. gigas might have multiple types of hemocytes with phagocytic capacity, some of which depend on β-glucan receptor(s) for phagocytosis.
For this study, we purified a novel isoform of the Manila clam lectin (designated MCL-4) from the plasma of the Manila clam, Ruditapes philippinarum, using affinity chromatography and gel filtration. Native PAGE results showed that the MCL-4 consisted of 70 kDa protein. The MCL-4 was found to be composed of 58-kDa and 43-kDa bands when examined using SDS-PAGE under reducing and non-reducing conditions. The native MCL-4 was revealed as a 147 kDa molecular mass protein by gel filtration. The purified MCL-4 agglutinates calcium-dependently in the erythrocytes of sheep and rabbit, but not in cells of the three species of marine bacteria tested. However, the phagocytic ability of the R. philippinarum hemocytes for the MCL-4-opsonized Vibrio tubiashii cells was significantly greater than that for the BSS-treated bacterial cells. Addition of purified MCL-4 markedly suppressed Alteromonas haloplanktis growth. These results suggest that MCL-4, because of its opsonizing and bacteriostatic properties, might contribute to the host defense mechanisms against invading microorganisms in R. philippinarum. Less
|
