| Budget Amount *help |
¥3,100,000 (Direct Cost: ¥3,100,000)
Fiscal Year 1998: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 1997: ¥2,000,000 (Direct Cost: ¥2,000,000)
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| Research Abstract |
As pressure is the fundamental physical quantity, the knowledge of the role of pressure is useful to understand the whole scheme of cell death. The viability of human immunocytes was investigated after compression at 37ﾟC for 30 minutes using a high pressure apparatus of the piston-cylinder type. In the case of EB3 (EB virus transformed human lymphoblasts), necrotic cells were observed immediately after decompression, and the viability decreased sigmoidally to zero with increasing pressure from 85 to 250 MPa. A 50% cell viability (P_<1/>) was attained at a pressure of 170 MPL Apoptotic cells began to be observed from 8 knirs after decompression (compression at 100 MPa for 30 minutes at 37ﾟC). The apoptosis was confirmed to be pressure-induced by the TUNEL (TdT-mediated dUll' nick end labeling) technique and by optical and electron microscopy. In the case of 50 MPa, the cell counts increased gradually after compression, which means that cell division progressed.
Qualitatively similar res
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ults were obtained with B cell lymphoma (Ramos). P_<1/> in HL-60 (human promyelocytic leukemia cells) and Ramos was lower than that in EB3. In the case of HL-60, apoptotic cells were observed immediately after decompression (compression at 100 MPa for 30 minutes at 37ﾟC). Execution of apoptosis should be different between EB3 and HL-60. Expression of protein related to pressure-induced apoptosis was studied using Western blot analysis. In EB3 cells, compared with uncompressed control, the expression of p53 and Bax increased from 5 lxurs after compression at IOOMPa, whereas that of caspase-3 (CPP32, apoptotic cysteine protease) decreased. Overexpression of p53 may induce caspase-3 processing through Bax activation. The pressure-induced attenuation in a-tubulin expression was observed after compression at lOOMPa, while it was not observed at 50 MPa. HL-60 is p53 deleted cell line. In LIL-60 cells, the expression of phospho p38 MAPK (mitogen-activated protein kinase), Bcl-2 and Bax increased immediatly after decompression at lOOMPa After 2 hours, the expression of an inactive 32-kDa precusor of caspase-3 (CPP32) decreased, and the mature activated p17 form of caspase-3 was detected. These findings suggest that p38 MAPK cascade may be induced by high-pressure, and the participation in the common signaling pathway (caspase cascade) may lead to apoptosis in HL-60 cells.
Our study of human cell biology at high pressure aids the medical application of high pressure in the near future. Less
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