Hydrogen peroxide induces intracellular calcium overload by activation of a non-selective cation channel in an insulin-secreting cell line
Hydrogen peroxide induces intracellular calcium overload by activation of a non-selective cation channel in an insulin-secreting cell line
复制标题
DOI：
10.1074/jbc.274.2.833
复制
发表时间：
1999-01-08
影响因子：
4.8
通讯作者：
Ashford, MLJ
中科院分区：
文献类型：
--
作者：
Herson, PS;Lee, K;Ashford, MLJ
用Fura-2荧光法研究了H2 O2对胰岛素分泌细胞系CRI-G1 [Ca（2+）]（i）的影响。H（2）O（2）（1-10 mM）引起游离[Ca（2+）]（i）的双相增加，在3分钟内观察到初始升高，在暴露30分钟后观察到第二次更大的升高。细胞外钙的去除阻断了[Ca（2+）]（i）的晚期而非初始升高，Thapsigargin不影响对H（2）O（2）的反应，但激活了电容性钙内流，10 μ M La（3+）消除了这种作用，同时记录细胞膜电位和[Ca（2+）]（i），表明[Ca（2+）]（i）对H2 O2的双相反应，并显示[Ca（2+）]（i）的晚期增加（i）与细胞膜电位崩溃时间上一致。将Ca（i）（2+）缓冲至低纳摩尔水平可防止[Ca（2+）]（i）和H（2）O（2）诱导的去极化增加的两个阶段。H（2）O（2）诱导的[Ca（2+）]（i）的晚期升高可被细胞外应用100 μ M La（3+）所阻止。La（3+）（100 μ M）抑制H（2）O（2）诱导的阳离子电流和NAD激活的阳离子（NS（NAD））通道活性。H（2）O（2）增加了完整CRI-G1细胞中的NAD/NADH比率，与细胞[NAD]增加一致。这些数据表明，H（2）O（2）增加[NAD]，其与增加的[Ca（2+）]（i）结合，激活NS（NAD）通道，引起不受调节的Ca（2+）进入和随后的细胞死亡。
Fura-2 fluorescence was used to investigate the effects of H(2)O(2) on [Ca(2+)](i) in the insulin-secreting cell line CRI-G1. H(2)O(2) (1-10 mM) caused a biphasic increase in free [Ca(2+)](i), an initial rise observed within 3 min and a second, much larger rise following a 30-min exposure. Extracellular calcium removal blocked the late, but not the initial, rise in [Ca(2+)](i), Thapsigargin did not affect either response to H(2)O(2), but activated capacitive calcium entry, an action abolished by 10 mu M La(3+), Simultaneous recordings of membrane potential and [Ca(2+)](i) demonstrated the same biphasic [Ca(2+)](i) response to H(2)O(2) and showed that the late increase in [Ca(2+)](i) coincided temporally with cell membrane potential collapse. Buffering Ca(i)(2+) to low nanomolar levels prevented both phases of increased [Ca(2+)](i) and the H(2)O(2)-induced depolarization. The H(2)O(2)-induced late rise in [Ca(2+)](i) was prevented by extracellular application of 100 mu M La(3+). La(3+) (100 mu M) inhibited the H(2)O(2)-induced cation current and NAD-activated cation (NS(NAD)) channel activity in these cells. H(2)O(2) increased the NAD/NADH ratio in intact CRI-G1 cells, consistent with increased cellular [NAD], These data suggest that H(2)O(2) increases [NAD], which, coupled with increased [Ca(2+)](i), activates NS(NAD), channels, causing unregulated Ca(2+) entry and consequent cell death.