Determining time of death: temperature-dependent postmortem changes in calcineurin A, MARCKS, CaMKII, and protein phosphatase 2A in mouse
Determining time of death: temperature-dependent postmortem changes in calcineurin A, MARCKS, CaMKII, and protein phosphatase 2A in mouse
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DOI：
10.1007/s00414-009-0343-x
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发表时间：
2009-07-01
影响因子：
2.1
通讯作者：
O'Day, Danton H.
中科院分区：
文献类型：
--
作者：
Poloz, Yekaterina O.;O'Day, Danton H.
虽然死后间隔（PMI）的确定在任何死亡调查中都是关键且基本的步骤，但用于PMI估算的合适生化方法的发展仍处于起步阶段。本研究关注小鼠中钙调神经磷酸酶A（CnA）、钙调蛋白依赖性激酶II（CaMKII）、豆蔻酰化富丙氨酸C激酶底物（MARCKs）和蛋白磷酸酶2A（PP2A）的温度依赖性死后降解。结果表明，MARCKs、CaMKII以及肺组织的使用似乎不值得进一步研究用于确定人类的PMI。在骨骼肌中，CnA在死后间隔的前48小时内经历了快速的温度依赖性裂解（60→约57 kDa）。在21℃时，这种转变在24小时内完成。相比之下，PP2A在最初的24小时内增加，之后在21℃时降解，但在5℃和10℃时可稳定长达96小时。添加蛋白酶抑制剂和MDL - 28170可抑制CnA的60→约57 kDa死后转变，这表明钙蛋白酶途径介导了这种分解。蛋白酶体抑制（MG - 132）和钙调蛋白拮抗（氯米达唑）也抑制了这种转变，这表明其他蛋白质降解途径也在起作用。相反，所有的蛋白酶抑制剂和氯米达唑（但不是乙二醇四乙酸）都导致PP2A水平升高。本文根据开发一种可用于现场的人类死后间隔确定的生化检测方法以及理解死亡时启动的蛋白质降解途径对数据进行了讨论。
While the determination of postmortem interval (PMI) is a crucial and fundamental step in any death investigation, the development of appropriate biochemical methods for PMI estimation is still in its infancy. This study focused on the temperature-dependent postmortem degradation of calcineurin A (CnA), calmodulin-dependent kinase II (CaMKII), myristoylated alanine-rich C-kinase substrate (MARCKs), and protein phosphatase 2A (PP2A) in mice. The results show that MARCKS, CaMKII, and the use of lung tissue do not appear to warrant further study for the determination of PMI in humans. In skeletal muscle, CnA underwent a rapid temperature-dependent cleavage (60 -> aEuro parts per thousand 57 kDa) over the first 48 h of postmortem interval. At 21A degrees C, this transformation was completed within 24 h. In contrast, PP2A increased within the first 24 h after which it degraded at 21A degrees C but remained stable for up to 96 h at 5A degrees C and 10A degrees C. The 60 -> aEuro parts per thousand 57 kDa postmortem conversion of CnA was inhibited by addition of protease inhibitors and MDL-28170 indicating a calpain pathway mediates this breakdown. Proteasome inhibition (MG-132) and calmodulin antagonism (calmidazolium) also inhibited this conversion suggesting that other protein degradation pathways also are in play. In contrast, all of the protease inhibitors and calmidazolium but not ethylene glycol tetraacetic acid led to increased levels of PP2A. The data are discussed in terms of developing a useable field-based biochemical assay for postmortem interval determination in humans and understanding the protein degradation pathways that are initiated upon death.