Basu D K, Karmazyn M
J Pharmacol Exp Ther. 1987 Aug;242(2):673-85.
This study was designed to evaluate the effect of an exogenous free radical generating system consisting of purine plus xanthine oxidase on the isolated rat heart and in particular to assess the possible contribution of arachidonic acid or its metabolites to toxicity produced by this drug combination. Purine plus xanthine oxidase produced a time-dependent depression in cardiac contractility which was associated with stimulated release of lactate dehydrogenase (LDH). Electron microscopic analysis revealed a distinct separation of the glycocalyx from the sarcolemmal membrane with no apparent intracellular defects. Purine plus xanthine oxidase was a potent stimulus for 6-keto-prostaglandin F1 alpha (6K-PGF1 alpha) synthesis but leukotriene production was undetectable under any condition. Eicosatetraynoic acid, which totally prevents the metabolism of arachidonic acid, accelerated the loss in force and increased LDH release invoked by purine plus xanthine oxidase, but produced no noticeable change in sarcolemmal ultrastructure. Cyclooxygenase inhibitors produced little influence although pretreatment with either acetylsalicylic acid or ibuprofen decreased contractility toward the end of purine plus xanthine oxidase perfusion. Nordihydroguarietic acid, a purported inhibitor of 5'-lipoxygenase accelerated the loss in force produced by purine plus xanthine oxidase. The nordihydroguarietic acid effects were associated with reduced 6K-PGF1 alpha efflux but LDH release was unaffected. We also examined whether modification of arachidonic acid release through changes in calcium concentration was associated with altered response to purine plus xanthine oxidase. Lowering the calcium concentration to 0.41 mM (from 1.25 mM control) reduced markedly 6K-PGF1 alpha, efflux as well as LDH release. Although the latter is suggestive of protection, hypocalcemic perfusion resulted in a greater loss in force due to free radical generation. Furthermore, cells from these hearts exhibited a greater degree of glycocalyx separation. Increasing the calcium concentration to 2.50 mM produced no further toxic manifestations in the response to purine plus xanthine oxidase, although the release of 6K-PGF1 alpha was increased. Our results suggest complex toxicity induced by an exogenously generated free radical system. The injury produced by this method is restricted to sarcolemmal changes, the latter being dependent on the external calcium concentration. The study further suggests that accumulation of intracellular unesterified arachidonic acid, which may result from peroxidation of membrane lipids, increases tissue injury caused by exogenous free radicals.
本研究旨在评估由嘌呤加黄嘌呤氧化酶组成的外源性自由基生成系统对离体大鼠心脏的影响,尤其要评估花生四烯酸或其代谢产物对该药物组合产生的毒性可能的作用。嘌呤加黄嘌呤氧化酶使心脏收缩性出现时间依赖性降低,这与乳酸脱氢酶(LDH)的释放增加有关。电子显微镜分析显示糖萼与肌膜明显分离,细胞内无明显缺陷。嘌呤加黄嘌呤氧化酶是6-酮-前列腺素F1α(6K-PGF1α)合成的有力刺激因素,但在任何条件下均未检测到白三烯的产生。完全阻止花生四烯酸代谢的二十碳四炔酸加速了嘌呤加黄嘌呤氧化酶引起的力量丧失并增加了LDH释放,但对肌膜超微结构无明显影响。环氧合酶抑制剂影响不大,尽管用乙酰水杨酸或布洛芬预处理在嘌呤加黄嘌呤氧化酶灌注末期降低了收缩性。去甲二氢愈创木酸,一种据称的5'-脂氧合酶抑制剂,加速了嘌呤加黄嘌呤氧化酶引起的力量丧失。去甲二氢愈创木酸的作用与6K-PGF1α流出减少有关,但LDH释放未受影响。我们还研究了通过改变钙浓度来改变花生四烯酸释放是否与对嘌呤加黄嘌呤氧化酶的反应改变有关。将钙浓度从对照的1.25 mM降至0.41 mM显著降低了6K-PGF1α流出以及LDH释放。尽管后者提示有保护作用,但低钙灌注由于自由基生成导致力量丧失更大。此外,这些心脏的细胞表现出更大程度的糖萼分离。将钙浓度增加到2.50 mM在对嘌呤加黄嘌呤氧化酶的反应中未产生进一步的毒性表现,尽管6K-PGF1α的释放增加了。我们的结果提示外源性自由基系统诱导的复杂毒性。这种方法产生的损伤仅限于肌膜变化,后者取决于细胞外钙浓度。该研究进一步提示,细胞内未酯化花生四烯酸的积累可能源于膜脂质的过氧化,会增加外源性自由基引起的组织损伤。