The heterocyclic amine, 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole induces apoptosis in cocultures of rat parenchymal and nonparenchymal liver cells.
The heterocyclic amine, 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole induces apoptosis in cocultures of rat parenchymal and nonparenchymal liver cells.
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杂环胺 3-氨基-1,4-二甲基-5H-吡啶并[4,3-b]吲哚可诱导大鼠实质和非实质肝细胞共培养物的细胞凋亡。
DOI：
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发表时间：
2001
影响因子：
3.8
通讯作者：
T. Hashimoto
中科院分区：
文献类型：
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作者：
H. Ashida;K. Kihara;Y. Nonaka;I. Fukuda;B. Shiotani;T. Hashimoto
在本研究中，我们研究了 3-氨基-1,4-二甲基-5H-吡啶并[4,3-b]吲哚 (Trp-P-1) 在实质和非实质肝细胞共培养物中的细胞凋亡机制，因为肝脏由多种细胞类型组成，并且它们对化学物质协同反应。研究发现，共培养物比每种细胞类型单独培养物更容易受到 Trp-P-1 导致的细胞死亡的影响。在共培养物中，Trp-P-1 诱导 DNA 片段化，同时激活 18-kDa 核酸内切酶。 Trp-P-1 (30 microM) 导致线粒体中 Bid 蛋白水平快速增加，并在处理后 15 分钟使细胞色素 c 从线粒体渗漏到细胞质中。另一方面，处理后 2 小时，随着 p53 蛋白水平和 NF-kappa B 的 DNA 结合活性增加，在线粒体组分中检测到 Bax 蛋白增加和 Bcl-2 蛋白减少。Caspase-8 在 30 分钟内被激活，随后使用相应的肽底物测量下游 caspase 的激活。通过蛋白质印迹分析，caspase-3、聚（ADP-核糖）聚合酶和蛋白激酶 C-delta 的裂解也证实了 caspase 的激活。 caspase-8 的肽抑制剂减少了 DNA 梯的形成和下游 caspase 的激活，但 caspase-9 抑制剂和作为 NF-kappa B 抑制剂的吡咯烷二硫代氨基甲酸盐仅表现出部分抑制，表明 caspase-8 是级联中的顶端 caspase。这些结果得出这样的结论：Trp-P-1 主要驱动涉及 Bid 的 caspase-8 介导的途径，并伴有涉及 Bax、Bcl-2 和 caspase-9 的 p53/NF-kappa B 介导的旁路途径的延迟。
In this study, we investigated the mechanism of apoptosis by 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) in cocultures of parenchymal and nonparenchymal liver cells, since the liver consists of various cell types and they cooperatively respond to chemicals. It was found that cocultures were more susceptible to cell death by Trp-P-1 than culture of each cell type alone. In cocultures, Trp-P-1 induced DNA fragmentation accompanied by the activation of 18-kDa endonuclease. Trp-P-1 (30 microM) caused a rapid increase in Bid protein level in mitochondria and the leakage of cytochrome c from mitochondria into the cytosol 15 min after treatment. On the other hand, an increase in Bax protein and a decrease in Bcl-2 protein were detected in the mitochondrial fraction 2 h after treatment following the increases in p53 protein level and DNA binding activity of NF-kappa B. Caspase-8 was activated within 30 min followed by the activation of downstream caspases as measured using the corresponding peptide substrates. The activation of caspases was also confirmed by cleavage of caspase-3, poly(ADP-ribose)polymerase, and protein kinase C-delta as analyzed by Western blotting. A peptide inhibitor of caspase-8 diminished DNA ladder formation and the activation of downstream caspases, but a caspase-9 inhibitor and pyrrolidinedithiocarbamate as an inhibitor of NF-kappa B showed only partial inhibition, suggesting that caspase-8 is the apical caspase in the cascade. These results led to the conclusion that Trp-P-1 mainly drives the caspase-8-mediated pathway that involves Bid, accompanied by a delay in the p53/NF-kappa B-mediated side pathway that involves Bax, Bcl-2, and caspase-9.