Bresgen Nikolaus, Ohlenschläger Ingo, Fiedler Brigitte, Wacht Nathalie, Zach Susanne, Dunkelmann Bettina, Arosio Paolo, Kuffner Elisabeth, Lottspeich Friedrich, Eckl Peter M
Department of Cell Biology, University of Salzburg, Salzburg, Austria.
J Cell Physiol. 2007 Jul;212(1):157-64. doi: 10.1002/jcp.21009.
Previously we have demonstrated an apoptosis inducing activity for a rat hepatocyte conditioned medium (CM) presumably mediated by acidic isoferritins. Here, we present support for this assumption since isoferritins purified from different rat hepatocyte CM significantly enhanced the frequency of apoptotic cells in primary rat hepatocytes, an effect completely inhibited by a neutralizing anti-H-ferritin antibody. The apoptosis induction appears to be related to a 43 kDa ferritin subunit contained in the isoferritins released from primary hepatocytes, presumably representing a ferritin heavy/light chain heterodimer. In addition, these isoferritins immunologically crossreact with antibodies raised against placental isoferritin p43-PLF (which also contains a 43 kDa ferritin subunit) and melanoma-derived H-chain ferritin, representing ferritin isoforms which reveal immunomodulatory properties. Furthermore, p53 and FasL are upregulated upon isoferritin treatment in a time dependent mode, and apoptosis induction can be suppressed by neutralizing anti-FasL antibodies. Proapoptotic Bid is upregulated too and translocated into mitochondria in primary hepatocytes exposed to the isoferritins purified from the CM. Finally, epidermal growth factor (EGF) and dexamethasone (DEX), which counteract proapoptotic mitochondrial signalling, almost completely abolished the proapoptotic effect of the hepatocyte derived isoferritins. In conclusion, our findings demonstrate that acidic isoferritins with homology to immunomodulatory ferritin isoforms (p43-PLF, melanoma-derived-H-chain ferritin) are released from hepatocytes in vitro, and are able to stimulate upregulation of p53 and mediate apoptosis involving Fas (CD95) signalling as well as addressing the intrinsic mitochondrial proapoptotic pathway.
此前我们已证明大鼠肝细胞条件培养基（CM）具有凋亡诱导活性，推测是由酸性异铁蛋白介导的。在此，我们为这一假设提供支持，因为从不同大鼠肝细胞CM中纯化的异铁蛋白显著提高了原代大鼠肝细胞中凋亡细胞的频率，而这种效应被一种中和性抗H-铁蛋白抗体完全抑制。凋亡诱导似乎与原代肝细胞释放的异铁蛋白中所含的一种43 kDa铁蛋白亚基有关，推测它代表一种铁蛋白重链/轻链异二聚体。此外，这些异铁蛋白与针对胎盘异铁蛋白p43-PLF（其也含有一个43 kDa铁蛋白亚基）和黑色素瘤来源的H链铁蛋白产生的抗体发生免疫交叉反应，这些铁蛋白异构体具有免疫调节特性。此外，异铁蛋白处理后，p53和FasL会以时间依赖性方式上调，并且中和性抗FasL抗体可抑制凋亡诱导。促凋亡蛋白Bid也会上调，并在暴露于从CM中纯化的异铁蛋白的原代肝细胞中转移至线粒体。最后，对抗促凋亡线粒体信号的表皮生长因子（EGF）和地塞米松（DEX）几乎完全消除了肝细胞来源的异铁蛋白的促凋亡作用。总之，我们的研究结果表明，与具有免疫调节特性的铁蛋白异构体（p43-PLF、黑色素瘤来源的H链铁蛋白）具有同源性的酸性异铁蛋白在体外从肝细胞中释放出来，并且能够刺激p53上调，介导涉及Fas（CD95）信号传导的凋亡以及激活内在的线粒体促凋亡途径。
