Saberi Behnam, Shinohara Mie, Ybanez Maria D, Hanawa Naoko, Gaarde William A, Kaplowitz Neil, Han Derick
Research Center for Liver Diseases, Keck School of Medicine, University of Southern California, Los Angeles, CA 90089-9121, USA.
Am J Physiol Cell Physiol. 2008 Jul;295(1):C50-63. doi: 10.1152/ajpcell.90654.2007. Epub 2008 May 7.
Recent studies have suggested that, in certain cases, necrosis, like apoptosis, may be programmed, involving the activation and inhibition of many signaling pathways. In this study, we examined whether necrosis induced by H(2)O(2) is regulated by signaling pathways in primary hepatocytes. A detailed time course revealed that H(2)O(2) treated to hepatocytes is consumed within minutes, but hepatocytes undergo necrosis several hours later. Thus, H(2)O(2) treatment induces a "lag phase" where signaling changes occur, including PKC activation, Akt (PKB) downregulation, activation of JNK, and downregulation of AMP-activated kinase (AMPK). Investigation of various inhibitors demonstrated that PKC inhibitors were effective in reducing necrosis caused by H(2)O(2) (80%). PKC inhibitor treatment decreased PKC activity but, surprisingly, also upregulated Akt and AMPK, suggesting that various PKC isoforms negatively regulate Akt and AMPK. Akt did not appear to play a significant role in H(2)O(2)-induced necrosis, since PKC inhibitor treatment protected hepatocytes from H(2)O(2) even when Akt was inhibited. On the other hand, compound C, a selective AMPK inhibitor, abrogated the protective effect of PKC inhibitors against necrosis induced by H(2)O(2). Furthermore, AMPK activators protected against H(2)O(2)-induced necrosis, suggesting that much of the protective effect of PKC inhibition was mediated through the upregulation of AMPK. Work with PKC inhibitors suggested that atypical PKC downregulates AMPK in response to H(2)O(2). Knockdown of PKC-alpha using antisense oligonucleotides also slightly protected (22%) against H(2)O(2). Taken together, our data demonstrate that the modulation of signaling pathways involving PKC and AMPK can alter H(2)O(2)-induced necrosis, suggesting that a signaling "program" is important in mediating H(2)O(2)-induced necrosis in primary hepatocytes.
最近的研究表明，在某些情况下，坏死与凋亡一样，可能是程序性的，涉及许多信号通路的激活和抑制。在本研究中，我们检测了过氧化氢（H₂O₂）诱导的坏死是否受原代肝细胞信号通路的调控。详细的时间进程显示，给予肝细胞的H₂O₂在数分钟内被消耗，但肝细胞数小时后才发生坏死。因此，H₂O₂处理诱导了一个信号变化发生的“延迟期”，包括蛋白激酶C（PKC）激活、蛋白激酶B（Akt，又称PKB）下调、c-Jun氨基末端激酶（JNK）激活以及腺苷酸活化蛋白激酶（AMPK）下调。对各种抑制剂的研究表明，PKC抑制剂能有效减少H₂O₂所致的坏死（约80%）。PKC抑制剂处理降低了PKC活性，但令人惊讶的是，同时也上调了Akt和AMPK，提示不同的PKC亚型对Akt和AMPK起负调控作用。Akt似乎在H₂O₂诱导的坏死中不起重要作用，因为即使Akt被抑制，PKC抑制剂处理仍能保护肝细胞免受H₂O₂损伤。另一方面，选择性AMPK抑制剂化合物C消除了PKC抑制剂对H₂O₂诱导坏死的保护作用。此外，AMPK激活剂能保护细胞免受H₂O₂诱导的坏死，提示PKC抑制的大部分保护作用是通过上调AMPK介导的。使用PKC抑制剂的研究提示，非典型PKC在H₂O₂作用下下调AMPK。使用反义寡核苷酸敲低PKC-α也能对H₂O₂提供轻微的保护（约22%）。综上所述，我们的数据表明，涉及PKC和AMPK的信号通路调节可改变H₂O₂诱导的坏死，提示信号“程序”在介导原代肝细胞H₂O₂诱导的坏死中起重要作用。
