Zinc Regulation & Toxicity in Cellular Energy Metabolism

描述（由申请人提供）：最近越来越多的报道试图将细胞内游离Zn++的变化与病理过程联系起来，特别是在神经系统中。细胞内Zn++升高与线粒体功能障碍、线粒体防御功能丧失、ROS生成增加和细胞死亡相关。我们已经证明1)升高的Zn++干扰线粒体的能量利用（呼吸），部分原因是抑制了在碳水化合物代谢中起作用的酶，特别是α -酮戊二酸脱氢酶。2) Zn++降低线粒体膜电位，诱导线粒体通透孔打开，释放Ca2+，抑制ATP生成。3) Zn++还能抑制吡啶核苷酸依赖的硫醇还原酶的活性，这对维持线粒体氧化还原防御至关重要。4) Zn++激活线粒体酶脂酰胺脱氢酶nadh氧化酶活性。这加速了超氧化物（O2）和过氧化氢（H2O2）的生成。5) Zn++诱导线粒体细胞色素c的释放，细胞色素c是细胞内凋亡的繁殖因子。

DESCRIPTION (provided by applicant): Recently a growing number of reports have attempted to link changes intracellular free Zn++ to pathological processes, particularly in the nervous system. Increased intracellular Zn++ is correlated with mitochondria dysfunction, loss of mitochondria defenses, increased production of ROS and cellular death. We have demonstrating that 1) Elevated Zn++ interferes with mitochondria energy utilization (respiration), due in part to inhibition of enzymes that play a role in carbohydrate metabolism, in particular alpha-ketoglutarate dehydrogenase. 2) Zn++ lowers mitochondria membrane potential and induces the opening of the mitochondria permeability pore, which releases Ca2+ and inhibits ATP generation. 3) Zn++ also inhibits the activity of pyridine nucleotide-dependent thiol reductases, which are critical to maintenance of mitochondria redox defenses. 4) Zn++ activates the NADH-oxidase activity of the mitochondria enzyme lipoamide dehydrogenase. This accelerates generation of superoxide (O2) and hydrogen peroxide (H2O2). 5) Zn++ induces release of mitochondria cytochrome c, which is a cytosolic propagator of apoptosis. Our hypotheses are that: Mitochondria and metallothioneins form a physiological feedback loop that utilizes Zn2+ as signal to regulate the balance of production between ATP, (ROS) and reduced thiols. Elevated intracellular Zn++ (is transported into mitochondria and) reduces cellular energy utilization directly by inhibiting enzymes involved in carbohydrate metabolism and electron transport and indirectly by interfering with mitochondrial redox balance. Excess (pathological) levels of cytosolic Zn2+ reduce mitochondria protection and destabilize mitochondria integrity (membrane permeability). Zn++ also induces mitochondria release of ROS and cytochrome c. Specifically, we will: 1) Characterize the spectrum of reactive products of the NADH-dehydrogenase activity of LADH that are stimulated by Zn++. 2) Characterize the inhibitory effect of Zn++ on thiol-redox activity at both the enzyme and mitochondrial level. 3) Establish the order of mitochondria events following exogenous addition of Zn++ in rat liver mitochondria. 4) Redox balance and other Zn++ -induced changes discovered and characterized in liver mitochondria in Aims 2 and 3 will be confirmed in rat brain mitochondria.