Institute of Nutrition and Food Technology (INTA), University of Chile, El Líbano 5524, Macul 5540, Casilla 13811, Santiago 11, Chile.
Biol Trace Elem Res. 2011 Sep;142(3):407-14. doi: 10.1007/s12011-010-8819-6. Epub 2010 Aug 25.
Acute and chronic cellular responses to changes in copper availability are not clear when these changes are mild to moderate, as what often occur in human daily life. The aims of the study were to develop an in vitro copper challenge in peripheral mononuclear cells (PMNCs) obtained from healthy individuals with different preconditioning copper treatments, and measure copper and iron content, and MT2A and TfR mRNA abundance after the copper challenge. (1) Screening using clinical and biochemical indicators defined healthy participants, who received 8 mg Cu/day (copper sulfate) or placebo for 2 months. (2) Mononuclear cells were obtained on days 0, 2 (acute changes), and 60 (chronic changes). (3) Cells were challenged with a 1, 5, and 20 μM Cu-histidine for 20 h, at T0, T2, and T60. Cells from both supplemented and placebo individuals showed a clear trend to increase copper content when there was more copper in the media. Increases were greater in the supplemented group, larger with 20 μM Cu (p < 0.02, one-way ANOVA), and mostly not significant when incubated with 5 μM Cu. By two-way ANOVA, differences were significant by treatment and by time (both p < 0.001). Differences between T0/T2 and T0/T60 were also significant (both p < 0.001). Changes of iron content were significant by treatment and time (two-way ANOVA); mRNA relative abundance of MT2A changed significantly and paralleled those of copper concentration, but TfR transcripts did not change. An in vitro challenge of PMNC showed specific changes of cellular copper and MT2A, while changes of iron content and TfR mRNA abundance were not consistent. PMNCs appear as good candidates to assess changes of cellular copper availability. That results differed after acute (T2) and chronic (T60) supplementation suggests that acute and chronic changes are handled differently by these cells.
当铜的变化轻微到中度时，如在人类日常生活中经常发生的那样，急性和慢性细胞对铜可用性变化的反应尚不清楚。本研究的目的是在来自接受不同预处理铜处理的健康个体的外周血单核细胞 (PMNC) 中建立体外铜挑战，并在铜挑战后测量铜和铁含量以及 MT2A 和 TfR mRNA 丰度。(1) 使用临床和生化指标筛选定义健康参与者，他们每天接受 8 mg Cu/day（硫酸铜）或安慰剂治疗 2 个月。(2) 在第 0 天、第 2 天（急性变化）和第 60 天（慢性变化）获得单核细胞。(3) 在 T0、T2 和 T60 时，用 1、5 和 20 μM Cu-组氨酸对细胞进行 20 h 的挑战。补充和安慰剂个体的细胞在培养基中含有更多铜时，铜含量明显呈上升趋势。补充组的增加幅度更大，20 μM Cu 时增加幅度更大（p < 0.02，单向方差分析），而用 5 μM Cu 孵育时则大多不显著。通过双向方差分析，处理和时间均有显著差异（均 p < 0.001）。T0/T2 和 T0/T60 之间的差异也具有统计学意义（均 p < 0.001）。铁含量的变化通过处理和时间的双向方差分析有统计学意义；MT2A 的 mRNA 相对丰度发生显著变化，并与铜浓度平行，但 TfR 转录本没有变化。PMNC 的体外挑战显示出细胞铜和 MT2A 的特定变化，而铁含量和 TfR mRNA 丰度的变化并不一致。PMNC 似乎是评估细胞铜可用性变化的良好候选物。急性（T2）和慢性（T60）补充后的结果不同表明，这些细胞对急性和慢性变化的处理方式不同。
