豬心臟纖維母細胞熱休克蛋白質70之合成

本研究之目的在研究豬胚心臟纖維母細胞經加熱處理後熱休克蛋白質合成等特性。由豬胚心臟所分離得到之纖維母細胞經由繼代培養不同代數所得量到的生長曲線顯示，彼此之間的生長趨勢類似。西方免疫吸漬法定量結果則可清楚看到，在不同培養代數之豬胚心臟纖維母細胞desmin及vimentin的量無明顯差異。所得的細胞株經45℃ 60分鐘加熱處理後使其恢復0到12小時，然後以放射性同位素標定法和西方免疫吸漬法來分析細胞熱休克蛋白質合成的質與量。結果清楚指出，熱休克蛋白質70與72可被熱處理所誘導而合成出來，而且在恢復10小時之後可累積至最大量。以上結果指出我們所分離而且經培養數代的豬胚心臟纖維母細胞具有相當的純度以及相當穩定的生長特性及熱休克反應性，是一個研究熱休克蛋白質與其在疾病形成過程中所扮演角色的很好的體外模式。

關鍵字

豬；心臟纖維母細胞；生長曲線；免疫染色；熱休克蛋白質

並列摘要

The purpose of this study was to characterize protein synthesis of cardiac fibroblasts from fetal pigs after heat treatments. There was no significant difference in trend of growth curves between fetal pig cardiac fibroblasts subcultured for various passage numbers. Western blot analysis showed no significant difference in the expression level of desmin and vimentin among the different passages. Further studies of protein synthesis in fetal pig cardiac fibroblast were followed by 45°C for 60 min heat treatment. After treatments, cells were recovered for up to 12 hr and then metabolically labeled with [35S] methionine for 1hr. De novo protein synthesis was monitored by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) and autoradiography. The 70 kD heat shock protein (HSP70) was quantitated by Western blotting using anti-HSP70 antibody. The results showed that the synthesis of HSP70 was heat-inducible and peaked at the period of 4 to 6 hr after recovery. Moreover, HSP70 was accumulated to a maximal level after 10 hr recovery. Thus, the fetal pig cardiac fibroblast cultures we established are of high purity, stability, and able to respond to heat shock treatment. This in vitro model may provide a valuable tool for the study of heat shock response played in cardiac-vascular diseases.

並列關鍵字

Pig ； Cardiac fibroblasts ； Immunofluorescence staining ； Heat shock protein

國際替代計量

豬心臟纖維母細胞熱休克蛋白質70之合成

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