Coyle P, Philcox J C, Rofe A M
Division of Clinical Chemistry, Institute of Medical and Veterinary Science, Adelaide SA.
Biol Trace Elem Res. 1993 Jan;36(1):35-49. doi: 10.1007/BF02783778.
The control of metallothionein (MT) synthesis was investigated in freshly prepared rat hepatocytes in experiments of short-term duration. Viability and metabolic function were maintained in incubations of 6-h duration. MT synthesis was measurable in hepatocytes from fed rats at Zn concentrations down to 1 microM. Zn and dexamethasone induced concentration-dependent increases in the synthesis of MT with maximal increases above the 5-h control of 3.2- and 2.5-fold, respectively. Zn induction of MT was first measurable at 2 h and was inhibited by actinomycin C. Although initial (0 h) MT concentrations in hepatocytes from fasted rats were double those from fed rats, after 6-h incubation in the presence of 50 microM Zn, the fasted rat hepatocytes showed only half the MT concentrations of the fed rat hepatocytes. Glucagon and interleukin-6 (IL-6) were less effective inducers and increased MT synthesis by 28 and 17%, respectively. IL-6 (100 U/mL) was found to have an additive effect on MT synthesis above that of Zn alone (1-50 microM) or Zn plus dexamethasone (1 microM). A supernatant from LPS-stimulated macrophages increased MT synthesis by 40%. The basal MT synthesis was not increased by either tumor necrosis factor-alpha (TNF-alpha) or interleukin-1 (IL-1). All incubations were carried out in the presence of RPMI 1640 medium with Hepes (20 mM), bicarbonate (24 mM), and fatty acid-free albumin (FAFA; 0.5% w/v). MT synthesis was also seen using Krebs bicarbonate buffer with glucose (10 mM), Hepes (20 mM), and FAFA (0.5% w/v), and although the level of MT synthesis was less than in RPMI, the increases in concentrations of MT at 5 h were 225, 139, 36 and 20% for Zn, dexamethasone, glucagon, and control, respectively. It is concluded that MT synthesis occurs in freshly prepared hepatocytes and that these cells are responsive to some of the established inducers of MT. This system enables the study of MT synthesis in individual rats in various metabolic and pathological states.
在短期实验中，对新分离的大鼠肝细胞中金属硫蛋白（MT）合成的调控进行了研究。在长达6小时的孵育过程中，细胞活力和代谢功能得以维持。在喂食大鼠的肝细胞中，当锌浓度低至1微摩尔时即可检测到MT的合成。锌和地塞米松可诱导MT合成呈浓度依赖性增加，最大增加量分别比5小时对照组高出3.2倍和2.5倍。锌对MT的诱导作用在2小时时首次可检测到，并被放线菌素C抑制。尽管禁食大鼠肝细胞的初始（0小时）MT浓度是喂食大鼠肝细胞的两倍，但在50微摩尔锌存在下孵育6小时后，禁食大鼠肝细胞的MT浓度仅为喂食大鼠肝细胞的一半。胰高血糖素和白细胞介素-6（IL-6）的诱导作用较弱，分别使MT合成增加28%和17%。发现IL-6（100单位/毫升）对MT合成具有加成作用，其作用超过单独使用锌（1 - 50微摩尔）或锌加地塞米松（1微摩尔）。脂多糖刺激的巨噬细胞上清液使MT合成增加40%。肿瘤坏死因子-α（TNF-α）或白细胞介素-1（IL-1）均未增加基础MT合成。所有孵育均在含有RPMI 1640培养基、Hepes（20毫摩尔）、碳酸氢盐（24毫摩尔）和无脂肪酸白蛋白（FAFA；0.5% w/v）的条件下进行。使用含有葡萄糖（10毫摩尔）、Hepes（20毫摩尔）和FAFA（0.5% w/v）的Krebs碳酸氢盐缓冲液时也观察到了MT合成，尽管MT合成水平低于在RPMI中的水平，但在5小时时，锌、地塞米松、胰高血糖素和对照组的MT浓度增加量分别为225%、139%、36%和20%。结论是，MT合成发生在新分离的肝细胞中，并且这些细胞对一些已确定的MT诱导剂有反应。该系统能够研究处于各种代谢和病理状态的个体大鼠中的MT合成。
