Eom Jae-Won, Lee Jong-Min, Koh Jae-Young, Kim Yang-Hee
Department of Molecular Biology, Sejong University, Seoul, 143-747, South Korea.
Department of Integrative Bioscience and Biotechnology, Sejong University, Seoul, 143-747, South Korea.
Mol Brain. 2016 Feb 9;9:14. doi: 10.1186/s13041-016-0194-6.
We reported that zinc neurotoxicity, a key mechanism of ischemic neuronal death, was mediated by poly ADP-ribose polymerase (PARP) over-activation following NAD(+)/ATP depletion in cortical cultures. Because AMP-activated protein kinase (AMPK) can be activated by ATP depletion, and AMPK plays a key role in excitotoxicity and ischemic neuronal death, we examined whether AMPK could be involved in zinc neurotoxicity in mouse cortical neuronal cultures.
Compound C, an AMPK inhibitor, significantly attenuated zinc-induced neuronal death. Activation of AMPK was detected beginning 2 h after a 10-min exposure of mouse cortical neurons to 300 μM zinc, although a significant change in AMP level was not detected until 4 h after zinc treatment. Thus, AMPK activation might not have been induced by an increase in intracellular AMP in zinc neurotoxicity. Furthermore, we observed that liver kinase B1 (LKB1) but not Ca(2+)/calmodulin-dependent protein kinase kinase β (CaMKKβ), was involved in AMPK activation. Although STO-609, a chemical inhibitor of CaMKKβ, significantly attenuated zinc neurotoxicity, zinc-induced AMPK activation was not affected, which suggested that CaMKKβ was not involved in AMPK activation. Knockdown of LKB1 by siRNA significantly reduced zinc neurotoxicity, as well as zinc-induced AMPK activation, which indicated a possible role for LKB1 as an upstream kinase for AMPK activation. In addition, mRNA and protein levels of Bim, a pro-apoptotic Bcl-2 family member, were noticeably increased by zinc in an AMPK-dependent manner. Finally, caspase-3 activation in zinc-induced neuronal death was mediated by LKB1 and AMPK activation.
The results suggested that AMPK mediated zinc-induced neuronal death via up-regulation of Bim and activation of caspase-3. Rapid activation of AMPK was detected after exposure of cortical neuronal cultures to zinc, which was induced by LKB1 activation but not increased intracellular AMP levels or CaMKKβ activation. Hence, blockade of AMPK in the brain may protect against zinc neurotoxicity, which is likely to occur after acute brain injury.
我们曾报道，锌神经毒性是缺血性神经元死亡的关键机制，在皮质培养物中，烟酰胺腺嘌呤二核苷酸（NAD(+)）/三磷酸腺苷（ATP）耗竭后，其由聚腺苷酸二磷酸核糖聚合酶（PARP）过度激活介导。由于AMP激活的蛋白激酶（AMPK）可被ATP耗竭激活，且AMPK在兴奋性毒性和缺血性神经元死亡中起关键作用，因此我们研究了AMPK是否参与小鼠皮质神经元培养物中的锌神经毒性。
AMPK抑制剂Compound C显著减轻了锌诱导的神经元死亡。在小鼠皮质神经元暴露于300μM锌10分钟后2小时开始检测到AMPK激活，尽管直到锌处理后4小时才检测到AMP水平有显著变化。因此，在锌神经毒性中，AMPK激活可能不是由细胞内AMP增加诱导的。此外，我们观察到肝脏激酶B1（LKB1）而非钙/钙调蛋白依赖性蛋白激酶激酶β（CaMKKβ）参与了AMPK激活。尽管CaMKKβ的化学抑制剂STO-609显著减轻了锌神经毒性，但锌诱导的AMPK激活未受影响，这表明CaMKKβ不参与AMPK激活。用小干扰RNA（siRNA）敲低LKB1显著降低了锌神经毒性以及锌诱导的AMPK激活，这表明LKB1可能作为AMPK激活的上游激酶发挥作用。此外，促凋亡Bcl-2家族成员Bim的mRNA和蛋白水平以AMPK依赖的方式被锌显著上调。最后，锌诱导的神经元死亡中的半胱天冬酶-3激活由LKB1和AMPK激活介导。
结果表明，AMPK通过上调Bim和激活半胱天冬酶-3介导锌诱导的神经元死亡。在皮质神经元培养物暴露于锌后检测到AMPK快速激活，这是由LKB1激活诱导的，而非细胞内AMP水平增加或CaMKKβ激活所致。因此，在脑中阻断AMPK可能预防锌神经毒性，而锌神经毒性可能在急性脑损伤后发生。
Zotero 插件
