Högberg J, Orrenius S, Larson R E
Eur J Biochem. 1975 Jan 15;50(3):595-602. doi: 10.1111/j.1432-1033.1975.tb09900.x.
Intracellular lipid peroxidation was initiated by the addition of ADP-complexed ferric iron to isolated rat hepatocytes and the reaction monitored by the thiobarbituric acid method or by measurement of the formation of conjugated dienes. Both the production of malondialdehyde (thiobarbituric-acid-reacting substances) and of conjugated dienes was dependent, on the ADP-Fe-3+ concentration in a dose-related fashion. Malondialdehyde formation stopped spontaneously within 20 min after the initiation of the reaction and the plateau reached was also related to the ADP-Fe-3+ concentration. Control experiments revealed that more than 90% of the malondialdehyde accumulating during the incubation period could be ascribed to intracellular production. The cellular NADPH/NADP+ ratio was always high and only slightly decreased upon ADP-Fe-3+-induced lipid peroxidation which, however, was associated with a marked decrease in the cellular glutathione concentration. The rate of accumulation of malondialdehyde as well as the final level reached during ADP-Fe-3+-initiated lipid peroxidation was increased by the addition of chloral hydrate. This apparent stimulatory effect could, however, be ascribed to the inhibition of the mitochondrial oxidation of the malondialdehyde formed during cellular lipid peroxidation, thus allowing more malondialdehyde to accumulate during the process. ADP-Fe-3+-induced cellular lipid peroxidation was associated with a decrease in the concentration of glutathione. Also, lowering of the intracellular glutathione level by the addition of diethyl maleate or by simply preincubating the hepatocytes (up to 50 min) promoted the ADP-Fe-3+ malondialdehyde production and formation of conjugated dienes. Furthermore, when cellular glutathione concentration had been lowered by preincubation of the hepatocytes, significant malondialdehyde production could be observed even at ADP-Fe-3+ concentrations which were too low to induce measurable lipid peroxidation in fresh hepatocytes. It is thus concluded that glutathione has an important role in the cell defence against lipid peroxidation and suggested that the isolated hepatocytes provide a suitable experimental model system for the characterization of this and other possible cellular defence mechanisms and how they are affected by the nutritional status of the donor animal.
通过向分离的大鼠肝细胞中添加与ADP络合的三价铁引发细胞内脂质过氧化反应，并通过硫代巴比妥酸法或通过测量共轭二烯的形成来监测该反应。丙二醛（硫代巴比妥酸反应物质）和共轭二烯的产生均以剂量相关的方式依赖于ADP-Fe³⁺浓度。反应开始后20分钟内丙二醛的形成会自发停止，达到的平台期也与ADP-Fe³⁺浓度有关。对照实验表明，孵育期间积累的丙二醛中超过90%可归因于细胞内产生。细胞的NADPH/NADP⁺比值始终较高，在ADP-Fe³⁺诱导的脂质过氧化过程中仅略有下降，然而，这与细胞内谷胱甘肽浓度的显著降低有关。添加水合氯醛会增加丙二醛的积累速率以及在ADP-Fe³⁺引发的脂质过氧化过程中达到的最终水平。然而，这种明显的刺激作用可归因于对细胞脂质过氧化过程中形成的丙二醛的线粒体氧化的抑制，从而使更多的丙二醛在该过程中积累。ADP-Fe³⁺诱导的细胞脂质过氧化与谷胱甘肽浓度的降低有关。此外，通过添加马来酸二乙酯或简单地对肝细胞进行预孵育（长达50分钟）来降低细胞内谷胱甘肽水平，会促进ADP-Fe³⁺诱导的丙二醛产生和共轭二烯的形成。此外，当通过预孵育肝细胞降低细胞内谷胱甘肽浓度时，即使在ADP-Fe³⁺浓度过低以至于在新鲜肝细胞中无法诱导可测量的脂质过氧化的情况下，也能观察到显著的丙二醛产生。因此得出结论，谷胱甘肽在细胞抵御脂质过氧化中起重要作用，并表明分离的肝细胞为表征这种及其他可能的细胞防御机制以及它们如何受供体动物营养状况影响提供了合适的实验模型系统。
