Chang Y J, Iwata S, Terada Y, Ozawa K
Intensive Care Unit, Shiga University of Medical Science, Otsu, Japan.
J Surg Res. 1996 Dec;66(2):91-9. doi: 10.1006/jsre.1996.0378.
High morbidity and mortality in surgical management for patients with obstructive jaundice was greatly attributed to the metabolic derangements in jaundiced liver mitochondria. Isolated liver mitochondria from jaundiced rat, produced by common bile duct ligation, were used to study the relationship among NADH level, oxygen consumption, and extramitochondrial calcium concentration. Alterations in NADH percentage and oxygen consumption were accomplished by incubating mitochondria with different substrates and monitoring oxygen consumption and NAD(P)H fluorescence simultaneously. In jaundiced liver mitochondria with glutamate + malate as substrate, respiration increased after the addition of exogenous Ca2+ at concentrations of 1 x 10(-7), 5 x 10(-7), and 1 x 10(-6) M. The maximal effect occurred at 5 x 10(-7) M. With different NADH-related substrates, the NAD(P)H fluorescence measurements (X axis) correlated linearly with state 3 respiration (Y axis), the slopes of the correlation curves being 2.27 and 0.79 in control and jaundiced mitochondria, respectively. After the addition of 5 x 10(-7) M Ca2+, the respirations of both control and jaundiced mitochondria increased and the slope for jaundiced mitochondria rose to 1.67. The matrix free Ca2+ concentration in jaundiced mitochondria, measured by fluo-3 loading, was higher than that in controls (162.1 +/- 16.7 nM, vs 129.7 +/- 12.6 nM, P < 0.01), while the matrix free/total Ca2+ ratio decreased from 34.9 +/- 6.0 (x10(-6)) to 27.2 +/- 4.4 (x10(-6), 9- < 0.05. The amplitude of the change in NAD(P)H fluorescence was reduced in jaundiced rat liver mitochondria and this correlated with the depression of respiration. A decrease in free/total Ca2+ ratio may be closely related to mitochondrial respiratory impairment in jaundice.
梗阻性黄疸患者外科治疗的高发病率和高死亡率很大程度上归因于黄疸肝脏线粒体的代谢紊乱。通过胆总管结扎制备的黄疸大鼠的离体肝线粒体，用于研究NADH水平、氧消耗和线粒体外钙浓度之间的关系。通过用不同底物孵育线粒体并同时监测氧消耗和NAD(P)H荧光来实现NADH百分比和氧消耗的改变。在以谷氨酸 + 苹果酸为底物的黄疸肝线粒体中，添加浓度为1×10(-7)、5×10(-7)和1×10(-6) M的外源Ca2+后呼吸增加。最大效应出现在5×10(-7) M。使用不同的与NADH相关的底物时，NAD(P)H荧光测量值（X轴）与状态3呼吸（Y轴）呈线性相关，对照和黄疸线粒体中相关曲线的斜率分别为2.27和0.79。添加5×10(-7) M Ca2+后，对照和黄疸线粒体的呼吸均增加，黄疸线粒体的斜率升至1.67。通过fluo-3负载测量，黄疸线粒体中的游离Ca2+浓度高于对照（162.1±16.7 nM，vs 129.7±12.6 nM，P < 0.01），而游离/总Ca2+比值从34.9±6.0（×10(-6)）降至27.2±4.4（×10(-6)），P < 0.05。黄疸大鼠肝线粒体中NAD(P)H荧光变化的幅度降低，这与呼吸抑制相关。游离/总Ca2+比值的降低可能与黄疸时的线粒体呼吸损伤密切相关。
