Characterization of cell death induced by ethacrynic acid in a human colon cancer cell line DLD-1 and suppression by N-acetyl-L-cysteine
Characterization of cell death induced by ethacrynic acid in a human colon cancer cell line DLD-1 and suppression by N-acetyl-L-cysteine
复制标题
DOI：
10.1111/j.1349-7006.2003.tb01371.x
复制
发表时间：
2003-10-01
期刊：
影响因子：
5.7
通讯作者：
Tsuchida, S
中科院分区：
文献类型：
--
作者：
Aizawa, S;Ookawa, K;Tsuchida, S
由于依他尼酸（EA），一种SH改性剂以及谷胱甘肽S-转移酶（GST）抑制剂，已被建议在一些细胞系中诱导凋亡，其对人结肠癌细胞系DLD-1的影响进行了研究。EA在20-40 μ M时促进细胞增殖，而在60-100 μ M时引起细胞死亡。Caspase抑制剂不能阻断细胞死亡，也没有检测到DNA梯状条带的形成。然而，聚（ADP-核糖）聚合酶被切割成82-kDa的片段，不同于特异性用于腺苷酸分解的85-kDa片段。该82-kDa片段不被抗半胱天冬酶3切割的PARP片段的抗体识别。N-乙酰-L-半胱氨酸（NAC）完全抑制EA诱导的细胞死亡，而3（2）-叔丁基-4-羟基苯甲醚或吡咯烷二硫代氨基甲酸铵盐则不能。谷氨酸（GSH）水平在EA处理的细胞中呈剂量依赖性增加，并且这种增加几乎不受NAC的影响。25-75 μ M EA处理的细胞中丝裂原活化蛋白激酶（MAPK）激酶（MEK）1、细胞外信号调节激酶（ERK）1和GST P1-1表达增加，而c-Jun N-末端激酶（JNK）1和p38 MAPK在100 μ M EA处理的细胞中显著降低。NAC抑制EA诱导的这些MAPK和GST P1-1的改变。p38 MAPK抑制剂SB 203580和FR 167653剂量依赖性地增强EA诱导的细胞死亡。MEK抑制剂U 0126不影响EA诱导的细胞死亡。这些研究表明，EA诱导的细胞死亡伴随着一种新的PARP片段化，但没有DNA片段化。p38 MAPK在EA诱导的细胞死亡中起抑制作用。
Since ethacrynic acid (EA), an SH modifier as well as glutathione S-transferase (GST) inhibitor, has been suggested to induce apoptosis in some cell lines, its effects on a human colon cancer cell line DLD-1 were examined. EA enhanced cell proliferation at 20-40 muM, while it caused cell death at 60-100 muM. Caspase inhibitors did not block cell death and DNA ladder formation was not detected. Poly(ADP-ribose) polymerase, however, was cleaved into an 82-kDa fragment, different from an 85-kDa fragment that is specific for apoptosisis. The 82-kDa fragment was not recognized by antibody against PARP fragment cleaved by caspase 3. N-Acetyl-L-cysteine (NAC) completely inhibited EA-induced cell death, but 3(2)-t-butyl-4-hydroxyanisole or pyrrolidinedithiocarbamate ammonium salt did not. Glutathione (GSH) levels were dose-dependently increased in cells treated with EA and this increase was hardly affected by NAC addition. Mitogen-activated protein kinase (MAPK) kinase (MEK) 1, extracellular signal-regulated kinase (ERK) 1 and GST P1-1 were increased in cells treated with 25-75 muM EA, while c-Jun N-terminal kinase (JNK) 1 and p38 MAPK were markedly decreased by 100 muM EA. NAC repressed EA-induced alterations in these MAPKs and GST P1-1. p38 MAPK inhibitors, SB203580 and FR167653, dose-dependently enhanced EA-induced cell death. An MEK inhibitor, U0126, did not affect EA-induced cell death. These studies revealed that EA induced cell death concomitantly with a novel PARP fragmentation, but without DNA fragmentation. p38 MAPK was suggested to play an inhibitory role in EA-induced cell death.