Department of Pathology, Metabolic Diseases Institute, University of Cincinnati, Cincinnati, Ohio 45237, USA.
Mol Pharmacol. 2012 Nov;82(5):1001-7. doi: 10.1124/mol.112.079863. Epub 2012 Aug 23.
c-Jun NH(2)-terminal kinase (JNK) activation plays a major role in acetaminophen (APAP)-induced hepatotoxicity. However, the exact mechanism of APAP-induced JNK activation is incompletely understood. It has been established that apoptosis signal-regulating kinase 1 (ASK1) regulates the late phase of APAP-induced JNK activation, but the mitogen-activated protein kinase kinase kinase that mediates the initial phase of APAP-induced JNK activation has not been identified. Oxidative stress produced during APAP metabolism causes JNK activation, which promotes mitochondrial dysfunction and results in the amplification of oxidative stress. Therefore, inhibition of the initial phase of JNK activation may be key to protection against APAP-induced liver injury. The goal of this study was to determine whether mixed-lineage kinase 3 (MLK3) mediates the initial, ASK1-independent phase of APAP-induced JNK activation and thus promotes drug-induced hepatotoxicity. We found that MLK3 was activated by oxidative stress and was required for JNK activation in response to oxidative stress. Loss of MLK3 attenuated APAP-induced JNK activation and hepatocyte death in vitro, independent of receptor-interacting protein 1. Moreover, JNK and glycogen synthase kinase 3β activation was significantly attenuated, and Mcl-1 degradation was inhibited in APAP-treated MLK3-knockout mice. Furthermore, we showed that loss of MLK3 increased expression of glutamate cysteine ligase, accelerated hepatic GSH recovery, and decreased production of reactive oxygen species after APAP treatment. MLK3-deficient mice were significantly protected from APAP-induced liver injury, compared with wild-type mice. Together, these studies establish a novel role for MLK3 in APAP-induced JNK activation and hepatotoxicity, and they suggest MLK3 as a possible target in the treatment of APAP-induced liver injury.
c-Jun NH(2)-末端激酶（JNK）的激活在乙酰氨基酚（APAP）诱导的肝毒性中起着重要作用。然而，APAP 诱导 JNK 激活的确切机制尚不完全清楚。已经证实凋亡信号调节激酶 1（ASK1）调节 APAP 诱导的 JNK 激活的晚期阶段，但介导 APAP 诱导的 JNK 激活初始阶段的丝裂原激活蛋白激酶激酶激酶尚未确定。APAP 代谢过程中产生的氧化应激导致 JNK 激活，从而促进线粒体功能障碍，并导致氧化应激的放大。因此，抑制 JNK 激活的初始阶段可能是防止 APAP 诱导的肝损伤的关键。本研究的目的是确定混合谱系激酶 3（MLK3）是否介导 APAP 诱导的 JNK 激活的初始、ASK1 非依赖性阶段，从而促进药物诱导的肝毒性。我们发现 MLK3 被氧化应激激活，并且是 JNK 对氧化应激反应的激活所必需的。MLK3 的缺失减弱了体外 APAP 诱导的 JNK 激活和肝细胞死亡，与受体相互作用蛋白 1 无关。此外，APAP 处理的 MLK3 敲除小鼠中 JNK 和糖原合成酶激酶 3β 的激活明显减弱，Mcl-1 降解受到抑制。此外，我们表明 MLK3 的缺失增加了谷氨酸半胱氨酸连接酶的表达，加速了肝 GSH 的恢复，并减少了 APAP 处理后的活性氧的产生。与野生型小鼠相比，MLK3 缺陷型小鼠对 APAP 诱导的肝损伤有明显的保护作用。总之，这些研究确立了 MLK3 在 APAP 诱导的 JNK 激活和肝毒性中的新作用，并表明 MLK3 可能是治疗 APAP 诱导的肝损伤的一个潜在靶点。
