The mechanism of protection of apoptosis by the mitochondrial PHGPx

本课题旨在探讨线粒体型磷脂氢过氧化物谷胱甘肽过氧化物酶（PHGPx）保护细胞凋亡的机制。线粒体过表达PHGPx可完全抑制2脱氧葡萄糖引起的细胞凋亡。细胞色素c （cyt.c）的释放提供了细胞凋亡的不可逆信号。过表达线粒体PHGPx抑制线粒体中细胞c的释放。cyt的解放有两个步骤。C来自线粒体。第一步是细胞c与线粒体内膜的分离，第二步是游离细胞的传代。细胞C通过腺嘌呤核苷酸转运子（adenine nucleotide translocator， ANT）调控开放的通透性过渡孔（PT孔），优先结合线粒体内特异性磷脂——心磷脂单层。细胞胞浆细胞与细胞胞浆单层的结合被细胞胞浆细胞的自氧化作用显著降低。在自由基引发剂作用下，细胞c可从含CL的脂质体中释放，提示细胞c可从线粒体内膜中释放。诱导细胞凋亡显著降低了ANT活性。ANT的失活与cyt的释放很好地对应。C来自线粒体。CLOOH与分离的线粒体融合后，线粒体的ANT活性明显受到抑制，提示CLOOH可以抑制ANT活性。在重组蛋白脂质体中加入cloh可抑制重组蛋白脂质体的ANT活性。这些结果表明，作为线粒体特异性磷脂的CL的过氧化可能是启动细胞凋亡的关键步骤。

In this project, we try to find out the mechanism of the protection of apoptosis mitochondrial type of phospholipid hydroperoxide glutathione peroxidase (PHGPx). Apoptosis caused by 2deoxyglucose was completely suppressed by the overexpression of mitochondrial PHGPx. The release of cytochrome c (cyt.c) provide the irreversible signal of apoptosis. Overexpression of mitochondrial PHGPx inhibit the release of cyt.c from mitochondria. There are two steps for the liberation of cyt. C from mitochondria. First step is the dissociation of cyt.c from inner membrane of mitochondria and second step is the passage of free cyt. C through the permeability transition pore (PT pore) which regulates the opening by adenine nucleotide translocator (ANT).Cyt.c preferentially binds to the monolayer of cardiolipin which is the specific phospholipid in mitochondria. Binding of cyt.c to CL monolayer significantly decreased by the autooxidation of CL. Cyt.c released from liposome containing CL by the peroxidation with radical initiator, these results suggest that cytc could release from inner membrane of mitochondoria.ANT activity was significantly decreased by the induction of apoptosis. Inactivation of ANT was well correspond to the liberation of cyt. C from mitochondria. ANT activity of mitochondria markedly inhibited by the fusion of CLOOH with isolated mitochondria suggesting that CLOOH could inhibit ANT activity. ANT activity of reconstituted proteoliposome was suppressed by the addition of CLOOH in proteoliposome.These results suggest that peroxidation of CL as a mitochondrial specific phospholipid might be a key step to initiate the apoptosis.