Endothelin-induced Apoptosis of A375 Human Melanoma Cells

作者：Makoto Okazawa, Takuma Shiraki, Haruaki Ninomiya, Shígeo Kobayashi, Tomoh Masaki · 发表于：Journal of Biological Chemistry · 年份：1998 · DOI：10.1074/jbc.273.20.12584 · 被引用次数：107 · 研究领域：Nitric Oxide and Endothelin Effects、Neutrophil, Myeloperoxidase and Oxidative Mechanisms、melanin and skin pigmentation

Endothelin-1 (ET-1) inhibited serum-dependent growth of asynchronized A375 human melanoma cells, and the growth inhibitory effect was markedly enhanced when ET-1 was applied to the cells synchronized at G1/S boundary by double thymidine blocks. Flow cytometric analysis revealed that ET-1 did not inhibit the cell cycle progression after the release of the block but caused a significant increase of the hypodiploid cell population that is characteristic of apoptotic cell death. ET-1-induced apoptosis was confirmed by the appearance of chromatin condensation on nuclear staining and DNA fragmentation on gel electrophoresis. The increase in the hypodiploid cell peak was manifest within 16 h of exposure to 5 nM ET-1. Within the same time range, ET-1 caused actin reorganization and drastic morphological changes of the surviving cells from epithelioid to an elongated bipolar shape. These phenotypical changes were preceded by ET-1-induced increase and nuclear accumulation of the tumor suppressor protein p53. All of these effects of ET-1 were mediated by ETB via a pertussis toxin-sensitive G protein. Flow cytometric analysis with fluorescent dye-labeled ET-1 revealed up-regulation of ETB expressed by the cells in G1/early S phases, and overexpression of the receptor protein by cDNA microinjection conferred the responsiveness (both apoptosis and morphological changes) to ET-1 irrespective of the position of the cell in the cell cycle. These results indicated the presence of ETB-mediated ...