Silva J P, Winterhalter K H, Richter C
Laboratory of Biochemistry I, Swiss Federal Institute of Technology (ETH), Zürich, Switzerland.
Redox Rep. 1997 Oct-Dec;3(5-6):331-41. doi: 10.1080/13510002.1997.11747131.
Rat liver mitochondria have a specific Ca2+ release pathway which operates when NAD+ is hydrolysed to nicotinamide and ADPribose. NAD+ hydrolysis is Ca(2+)-dependent and inhibited by cyclosporine A (CSA). Mitochondrial Ca2+ release can be activated by the prooxidant t-butylhydroperoxide (tbh) or by gliotoxin (GT), a fungal metabolite of the epipolythiodioxopiperazine group. Tbh oxidizes NADH to NAD+ through an enzyme cascade consisting of glutathione peroxidase, glutathione reductase, and the energy linked transhydrogenase, whereas GT oxidizes some vicinal thiols to the disulfide form, a prerequisite for NAD+ hydrolysis. We report now that rat skeletal muscle mitochondria also contain a specific Ca2+ release pathway activated by both tbh and GT. Ca2+ release increases with the mitochondrial Ca2+ load, is completely inhibited in the presence of CSA, and is paralleled by pyridine nucleotide oxidation. In the presence of tbh and GT, mitochondria do not lose their membrane potential and do not swell, provided continuous release and re-uptake of Ca2+ ('Ca2+ cycling') is prevented. These data support the notion that both tbh- and GT-induced Ca2+ release are not the consequence of an unspecific increase of the inner membrane permeability ('pore' formation). Tbh induces Ca2+ release from rat skeletal muscle less efficiently than from liver mitochondria indicating that the coupling between tbh and NADH oxidation is much weaker in skeletal muscle mitochondria. This conclusion is corroborated by a much lower glutathione peroxidase activity in skeletal muscle than in liver mitochondria. The prooxidant-dependent pathway promotes, under drastic conditions (high mitochondrial Ca2+ loads and high tbh concentrations), Ca2+ release to about the same extent and rate as the Na+/Ca2+ exchanger. This renders the prooxidant-dependent pathway relevant in the pathophysiology of mitochondrial myopathies where its activation by an increased generation of reactive oxygen species probably results in excessive Ca2+ cycling and damage to mitochondria.
大鼠肝脏线粒体具有一种特定的Ca2+释放途径，当NAD+水解为烟酰胺和ADP核糖时该途径发挥作用。NAD+水解依赖于Ca(2+)，并受到环孢素A（CSA）的抑制。线粒体Ca2+释放可被促氧化剂叔丁基过氧化氢（tbh）或由表硫代二氧哌嗪类真菌代谢产物麦角硫因（GT）激活。tbh通过由谷胱甘肽过氧化物酶、谷胱甘肽还原酶和能量偶联转氢酶组成的酶级联反应将NADH氧化为NAD+，而GT将一些邻位硫醇氧化为二硫键形式，这是NAD+水解的前提条件。我们现在报告，大鼠骨骼肌线粒体也含有一种由tbh和GT激活的特定Ca2+释放途径。Ca2+释放随着线粒体Ca2+负荷的增加而增加，在CSA存在时完全被抑制，并且与吡啶核苷酸氧化平行。在tbh和GT存在的情况下，只要Ca2+的持续释放和再摄取（“Ca2+循环”）被阻止，线粒体就不会失去其膜电位且不会肿胀。这些数据支持这样一种观点，即tbh和GT诱导的Ca2+释放不是内膜通透性非特异性增加（“孔”形成）的结果。与肝脏线粒体相比，tbh诱导大鼠骨骼肌Ca2+释放的效率较低，这表明在骨骼肌线粒体中tbh与NADH氧化之间的偶联要弱得多。骨骼肌中谷胱甘肽过氧化物酶活性远低于肝脏线粒体，这一结论得到了进一步证实。在极端条件下（高线粒体Ca2+负荷和高tbh浓度），促氧化剂依赖性途径促进Ca2+释放的程度和速率与Na+/Ca2+交换器大致相同。这使得促氧化剂依赖性途径在线粒体肌病的病理生理学中具有相关性，在这种疾病中，活性氧生成增加对其激活可能导致Ca2+过度循环和线粒体损伤。
