Bingham M J, Sargeson A M, McArdle H J
Department of Child Health, Ninewells Hospital and Medical School, University of Dundee, United Kingdom.
Am J Physiol. 1997 Jun;272(6 Pt 1):G1400-7. doi: 10.1152/ajpgi.1997.272.6.G1400.
When hepatocytes are incubated with the chelator diamsar, two pools can be identified, which we have termed extractable and nonextractable. On entering the hepatocyte, 67Cu first associates with the extractable pool and, after approximately 2 h, moves to the nonextractable pool. Both pools demonstrate saturation and are filled as a function of Cu concentration and incubation time. Using the Michaelis-Menten equation, we have estimated the size of the pools after incubation with 67Cu for 30 min and 4 h. During this period the extractable pool decreases in size from 200 +/- 27 to 116 +/- 5 pmol/microgram DNA, whereas the nonextractable pool increases from 28 +/- 9 to 77 +/- 11 pmol/microgram DNA. Movement of Cu from the nonextractable pool to the extractable pool is slow and incomplete. Using [3H]diamsar, we demonstrate that uptake of the chelator is not rate limiting and probably does not occur by pinocytosis. Incubation with diamsar does not affect the activity of superoxide dismutase or cytochrome-c oxidase, although it does prevent the incorporation of 67Cu into ceruloplasmin. Incubation with zinc, which induces metallothionein, results in an increase in 67Cu associated with the nonextractable pool, suggesting that 67Cu-metallothionein constitutes at least part of the nonextractable pool.
当肝细胞与螯合剂二氨基丁二酸（diamsar）一起孵育时，可以识别出两个池，我们将其称为可提取池和不可提取池。67Cu进入肝细胞后，首先与可提取池结合，大约2小时后，转移到不可提取池。两个池都表现出饱和现象，并根据铜浓度和孵育时间而被填满。使用米氏方程（Michaelis-Menten equation），我们估计了与67Cu孵育30分钟和4小时后池的大小。在此期间，可提取池的大小从200±27降至116±5 pmol/μg DNA，而不可提取池则从28±9增加到77±11 pmol/μg DNA。铜从不可提取池向可提取池的移动缓慢且不完全。使用[3H]二氨基丁二酸，我们证明螯合剂的摄取不是限速步骤，并且可能不是通过胞饮作用发生的。用二氨基丁二酸孵育不会影响超氧化物歧化酶或细胞色素c氧化酶的活性，尽管它确实会阻止67Cu掺入铜蓝蛋白。用锌孵育可诱导金属硫蛋白，导致与不可提取池相关的67Cu增加，这表明67Cu-金属硫蛋白至少构成了不可提取池的一部分。
