Studies on the molecular mechanisms of carbon monoxide on the induction or protection of cell death.

一氧化碳(CO)，无论是外源性的还是内源性的，都可以通过核因子-kB或线粒体ATP依赖的K~(++)通道来保护心肌和血管细胞免受缺血或内毒素的损伤。血红素加氧酶(HO)-1产生一氧化碳，从而保护细胞。我们已经证明，一种钙依赖的蛋白酶Calain通过α-fodrin蛋白分解促进缺氧条件下心源性H9c2细胞的坏死性死亡(Aki，T.，Yoshida，K.，和Fujimiya，T.(2002)J.Biochem)。132、921-926)。在这里，我们展示了第一条证据，一氧化碳抑制钙内流，通过荧光检测，这种抑制被L类型的钙通道激动剂BAYK8644增强。通过染料排斥、乳酸脱氢酶释放或碘化丙啶的通透性，缺血性死亡表现为坏死性。CO或L型钙通道阻断剂维拉帕米可抑制钙离子内流、蛋白印迹法检测的α-fodrin蛋白降解和缺血性死亡。正如JC-1检测到的那样，缺血也引起线粒体去极化，这一作用可被一氧化碳或维拉帕米抑制。此外，DCF、氢乙啶或Amplex Red过氧化氢检测到的ROS生成在缺血后增加，但不受复方氯化血红素处理的影响。Western blotting检测到缺氧细胞HO-1的表达增加。HO-1的诱导减少了缺血后钙离子内流和细胞死亡。因此，外源性和内源性CO通过抑制L型钙通道和钙蛋白酶激活抑制钙内流，从而保护心肌细胞免受缺血损伤。

Carbon monoxide (CO), either exogenous or endogenous, has been shown to protect the myocardial and vascular cells against injuries due to ischemia or lipopolysaccharide, through NF-kB or mitochondrial ATP-dependent K^+-channel. Heme-oxygenase (HO)-1, generates CO, thereby protecting the cells. We have shown that a Ca^<2+>-dependent protease calpain promotes necrotic death in the cardiogenic H9c2 cells under hypoxia through α-fodrin proteolysis (Aki, T., Yoshida, K., and Fujimiya, T. (2002) J.Biochem. 132, 921-926). Here, we show the first line of evidence that CO inhibits Ca^<2+>-influx, as detected by fluo-3 fluorescenece, which was enhanced by a L-type Ca^<2+>-channel agonist BAYK8644. The ischemic death was characterized as necrotic either by dye exclusion, LDH release, or propidium iodide permeabilization. The Ca^<2+>-influx, α-fodrin proteolysis, as detected by western blotting, and the ischemic death, were inhibited by CO ora L-type Ca^<2+.-channel inhibitor verapamil. Ischemia also induced mitochondrial depolarization, as detected by JC-1, which was inhibited by CO or verapamil. Additionally, reactive oxygen species (ROS) generation, as detected by DCF, hydroethidine, or Amplex Red Hydrogen Peroxide, was enhanced by ischemia, but unaffected by Co. Hemin treatment increased the HO-1 expression in the hypoxic cells, as detected by western blotting. The HO-1 induction reduced the Ca^<2+>-influx and cell death after ischemia. Thus, exogenous and endogenous CO protect the cardiomyogenic cells against ischemia by inhibiting Ca^<2+>-influx through L-type Ca^<2+> channel and calpain activation.