Yuan Qing, Ray Ramesh M, Johnson Leonard R
Department of Physiology, The University of Tennessee Health Science Center, Memphis, Tennessee 38163, USA.
Am J Physiol Cell Physiol. 2002 Jun;282(6):C1290-7. doi: 10.1152/ajpcell.00351.2001.
We have shown previously that depletion of polyamines delays apoptosis induced by camptothecin in rat intestinal epithelial cells (IEC-6). Mitochondria play an important role in the regulation of apoptosis in mammalian cells because apoptotic signals induce mitochondria to release cytochrome c. The latter interacts with Apaf-1 to activate caspase-9, which in turn activates downstream caspase-3. Bcl-2 family proteins are involved in the regulation of cytochrome c release from mitochondria. In this study, we examined the effects of polyamine depletion on the activation of the caspase cascade, release of cytochrome c from mitochondria, and expression and translocation of Bcl-2 family proteins. We inhibited ornithine decarboxylase, the first rate-limiting enzyme in polyamine synthesis, with alpha-difluoromethylornithine (DFMO) to deplete cells of polyamines. Depletion of polyamines prevented camptothecin-induced release of cytochrome c from mitochondria and decreased the activity of caspase-9 and caspase-3. The mitochondrial membrane potential was not disrupted when cytochrome c was released. Depletion of polyamines decreased translocation of Bax to mitochondria during apoptosis. The expression of antiapoptotic proteins Bcl-x(L) and Bcl-2 was increased in DFMO-treated cells. Caspase-8 activity and cleavage of Bid were decreased in cells depleted of polyamines. These results suggest that polyamine depletion prevents IEC-6 cells from apoptosis by preventing the translocation of Bax to mitochondria, thus preventing the release of cytochrome c.
我们之前已经表明，多胺的消耗会延迟喜树碱诱导的大鼠肠上皮细胞（IEC-6）凋亡。线粒体在哺乳动物细胞凋亡调控中起重要作用，因为凋亡信号诱导线粒体释放细胞色素c。后者与凋亡蛋白酶激活因子-1（Apaf-1）相互作用以激活半胱天冬酶-9，进而激活下游的半胱天冬酶-3。Bcl-2家族蛋白参与线粒体细胞色素c释放的调控。在本研究中，我们检测了多胺消耗对半胱天冬酶级联激活、线粒体细胞色素c释放以及Bcl-2家族蛋白表达和转位的影响。我们用α-二氟甲基鸟氨酸（DFMO）抑制鸟氨酸脱羧酶（多胺合成中的首个限速酶），以消耗细胞内的多胺。多胺的消耗可防止喜树碱诱导的线粒体细胞色素c释放，并降低半胱天冬酶-9和半胱天冬酶-3的活性。细胞色素c释放时线粒体膜电位未被破坏。多胺的消耗减少了凋亡过程中Bax向线粒体的转位。在经DFMO处理的细胞中，抗凋亡蛋白Bcl-x(L)和Bcl-2的表达增加。多胺消耗的细胞中半胱天冬酶-8活性和Bid的裂解减少。这些结果表明，多胺消耗通过阻止Bax向线粒体转位，从而防止细胞色素c释放，进而阻止IEC-6细胞凋亡。
