The pyrrolizidine alkaloid senecionine induces CYP-dependent destruction of sinusoidal endothelial cells and cholestasis in mice
The pyrrolizidine alkaloid senecionine induces CYP-dependent destruction of sinusoidal endothelial cells and cholestasis in mice
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DOI：
10.1007/s00204-019-02582-8
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发表时间：
2019-10-12
影响因子：
6.1
通讯作者：
Reif, Raymond
中科院分区：
文献类型：
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作者：
Hessel-Pras, Stefanie;Braeuning, Albert;Reif, Raymond
吡咯里西啶生物碱（Pyrrolizidine alkaloids，PA）是一种广泛存在的植物毒素，可引起人类和其他哺乳动物严重的肝损伤，其机制尚不完全清楚。因此，我们研究了PA肝毒性在体内的发展，在小鼠中使用急性毒性剂量的PA senecietin，结合活体双光子显微镜，组织学，临床化学，并在体外实验与原代小鼠肝细胞和肝窦内皮细胞（LSEC）。我们观察到中央周围LSEC坏死以及升高的正弦标记蛋白在血清中的千里光碱治疗的小鼠和增加的正弦血小板聚集在受损的组织区域。体外实验表明，对新鲜分离的LSECs没有细胞毒性，高达500 μ M senecietin。然而，通过与原代小鼠肝细胞预孵育进行的seneciblide代谢活化增加了对培养的LSEC的细胞毒性，EC 50约为22 μ M。细胞色素P450（P450）-依赖性的seneciprimidine生物激活证实在PA还原酶缺陷小鼠中，没有观察到PA诱导的肝毒性。因此，seneciprin的毒性代谢产物由肝CYP产生，并可能部分从肝细胞释放，导致肝小叶中心周围区域的LSEC破坏。通过活体双光子成像对肝胆汁盐转运的分析显示，荧光胆汁盐类似物从肝窦进入肝细胞的摄取延迟，消除延迟。这伴随着肝胆盐转运蛋白如Abcb 11或Abcc 1的转录失调。总之，尽管毒性代谢物在邻近的中央周围肝细胞中以CYP依赖性方式形成，但千里光素破坏LSEC。
Pyrrolizidine alkaloids (PAs) are widely occurring phytotoxins which can induce severe liver damage in humans and other mammalian species by mechanisms that are not fully understood. Therefore, we investigated the development of PA hepatotoxicity in vivo, using an acutely toxic dose of the PA senecionine in mice, in combination with intravital two-photon microscopy, histology, clinical chemistry, and in vitro experiments with primary mouse hepatocytes and liver sinusoidal endothelial cells (LSECs). We observed pericentral LSEC necrosis together with elevated sinusoidal marker proteins in the serum of senecionine-treated mice and increased sinusoidal platelet aggregation in the damaged tissue regions. In vitro experiments showed no cytotoxicity to freshly isolated LSECs up to 500 mu M senecionine. However, metabolic activation of senecionine by preincubation with primary mouse hepatocytes increased the cytotoxicity to cultivated LSECs with an EC50 of approximately 22 mu M. The cytochrome P450 (CYP)-dependency of senecionine bioactivation was confirmed in CYP reductase-deficient mice where no PA-induced hepatotoxicity was observed. Therefore, toxic metabolites of senecionine are generated by hepatic CYPs, and may be partially released from hepatocytes leading to destruction of LSECs in the pericentral region of the liver lobules. Analysis of hepatic bile salt transport by intravital two-photon imaging revealed a delayed uptake of a fluorescent bile salt analogue from the hepatic sinusoids into hepatocytes and delayed elimination. This was accompanied by transcriptional deregulation of hepatic bile salt transporters like Abcb11 or Abcc1. In conclusion, senecionine destroys LSECs although the toxic metabolite is formed in a CYP-dependent manner in the adjacent pericentral hepatocytes.