| Budget Amount *help |
¥6,500,000 (Direct Cost: ¥6,500,000)
Fiscal Year 1988: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1987: ¥6,000,000 (Direct Cost: ¥6,000,000)
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| Research Abstract |
G-actin was purified from the acetone powder prepared using the rat intestinal smooth muscle and, thereafter, it was treated with performic acid. Mice were sensitized with the denaturated actin. Thereafter, the spleen cells of the sensitized mice were isolated and fused with myeloma cells by means of an electric hybridizer. Antibody-secreting elone was established and anti-rat actin actin monoclonal IgG was obtained. Tubulin was isolated from the rat brain and anti-tubulin monoclonal IgG was prepared similarly. Guinea-pigs were sensitized with ovalbumin according to conventional techniques. When sensitized guinea-pig lung mast cells were isolated and exposed to antigen, granules were pushed out on to the cell surface. Subsequently, microfilaments were projected with extruded granules. Using anti-actin antibody or phalloidine stainings, it was revealed that these filaments were consisted mainly of actin. The conformational changes in microfilaments took place in accordance with an incre
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ase of intracellular Ca^<2+> concentrations. When intracellular Ca levels became lower close to the control level, the extruded filaments became shorter and the granules were reincorporated into the cytoplasma. By means of electron microscopy, it was revealed that a complicated, dense network of filaments exist in the mast cells connecting plasma membrane and granules, and the granules are interconnected by these filaments. Using anti-actin and anti-tubulin antibodies, it was clearly shown that the complicated networks are mostly consisted of microfilaments and microtubules. It was assumed that these cytoskeletons play a variety of important roles not only in expelling the granules out of the cell but also in altering the cell shape. Furthermore, intracellular distribution of the microtubules was studied using rat basophilic leukemia cell line. Consequently, it became evident that the microtubules are densely located in the periphery of the cell, indicating that these microtubules are important in exerting the morphological changes of the cell as well as in adhering to the tissues. Less
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