背景超氧阴离子(O2-)对肺实质改变细胞增殖有多重作用,细胞代谢,和气道平滑肌(ASM)收缩。细胞内钙([Ca2+]i)浓度在调节ASM收缩中起着重要作用,放松,扩散,和基因表达。目的研究O2-对激动剂刺激的ASM细胞[Ca2]i变化的影响。设计/方法Fura-2AM加载,新鲜分离的猪ASM(PASM)细胞用于检查响应乙酰胆碱(ACh)的[Ca2]i释放,组胺,内皮素,咖啡因,和thapsigargin(TPG)在存在或不存在细胞外Ca2+的情况下。结果PASM细胞暴露于黄嘌呤和黄嘌呤氧化酶(XXO)导致O2-的时间依赖性产生,被超氧化物歧化酶(SOD)抑制。用X+XO预孵育PASM细胞15或45分钟抑制对ACh的净[Ca2+]i反应,组胺,咖啡因,和TPG与对照细胞相比。用SOD预处理PASM细胞30分钟减轻了XXO处理对ACh诱导的Ca2升高的抑制作用,提示O2-的作用。X+XO处理还抑制咖啡因和TPG诱导的Ca2+升高,表明O2-对[Ca2+]i释放和再摄取机制的影响。结论超氧化物减弱[Ca2+]i的释放,再摄取,并可能干扰ASM细胞的生理功能。
Background
Superoxide anions (O 2 - ) have multiple effects on pulmonary parenchyma altering cell proliferation, cellular metabolism, and airway smooth muscle (ASM) contraction. Intracellular calcium ([Ca 2+ ] i ) concentration plays a significant role in the regulation of ASM contraction, relaxation, proliferation, and gene expression. Objective We investigated the effects of O 2 - on agonist-stimulated changes in [Ca 2+ ] i in ASM cells. Design/Methods Fura-2 AM-loaded, freshly isolated porcine ASM (PASM) cells were used to examine [Ca 2+ ] i release in response to acetylcholine (ACh), histamine, endothelin, caffeine, and thapsigargin (TPG) in the presence or absence of extracellular Ca 2+ . Results Exposure of PASM cells to xanthine and xanthine oxidase (X + XO) resulted in a time-dependent generation of O 2 - , inhibited by
superoxide dismutase (SOD). Preincubating PASM cells with X + XO for 15- or 45-minute inhibited net [Ca 2+ ] i responses to ACh, histamine, caffeine, and TPG compared with control cells. Pretreating PASM cells with SOD for 30 minutes mitigated the inhibitory effect of X + XO treatment on ACh-induced Ca 2+ elevation suggesting role of O 2 - . X + XO treatment also inhibited caffeine- and TPG-induced Ca 2+ elevation suggesting effect of O 2 - on [Ca 2+ ] i release and reuptake mechanisms. Conclusion
Superoxide attenuates [Ca 2+ ] i release, reuptake, and may interfere with physiological functions of ASM cells.