Regulation of L-arginine transport and nitric oxide release in superfused porcine aortic endothelial cells
Regulation of L-arginine transport and nitric oxide release in superfused porcine aortic endothelial cells
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DOI：
10.1113/jphysiol.1996.sp021138
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发表时间：
1996-01-01
影响因子：
5.5
通讯作者：
Mann, GE
中科院分区：
文献类型：
--
作者：
Bogle, RG;Baydoun, AR;Mann, GE
1.我们研究了细胞外离子组成的改变和底物剥夺对L-精氨酸转运和一氧化氮(NO)和前列环素(PGI(2))释放的影响。可饱和的L-精氨酸转运(K-m=0.14+/-0.03 mm；V-max=2.08+/-0.54nmolmin(-1)(5×10(6)cell)(-1))对pH不敏感，不受细胞外Na+或Ca~(2+)的影响。阳离子精氨酸类似物L-赖氨酸和L-鸟氨酸对L-精氨酸转运有抑制作用，而2-甲氨基异丁酸、beta-2-amino-bicyclo[2，2.1]-heptane-2-carboxylic酸、L-苯丙氨酸、6-重氮-5-氧代去甲亮氨酸、L-谷氨酰胺、L-半胱氨酸和L-谷氨酸的抑制作用较差。去L-精氨酸(30分钟至24小时)使细胞内游离L-精氨酸水平从0.87+/-0.07降至0.40+/-0.05 mM(P<；0.05)，并对L-精氨酸、L-赖氨酸和L-鸟氨酸转运产生40%的刺激。L-精氨酸和N-G-单甲基-L-精氨酸(L-NMMA)，而不是N-omega-硝基-L-精氨酸甲酯(L-NAME)，反式刺激L-[H-3]精氨酸的外流。70 mM K+去极化可减少L-精氨酸内流，阻止100 nM缓激肽对血管内皮细胞转运的刺激，但仍可检测到激动剂诱导的NO和PGI(2)释放。在名义上无钙溶液的细胞灌流过程中，L-精氨酸转运和NO释放的基础速率没有受到影响。缓激肽刺激的L-精氨酸转运对钙离子的清除不敏感，而激动剂诱导的NO释放则被取消。尽管缓激肽刺激的NO释放似乎与L-精氨酸转运的一过性增加没有直接的联系，但激动剂诱导的膜超极化或底物剥夺导致的通过y(+)系统的L-精氨酸内流速率的增加为增加L-精氨酸的供应以维持NO的产生提供了一个机制。
1. We have investigated whether changes in extracellular ion composition and substrate deprivation modulate basal and/or bradykinin-stimulated L-arginine transport and release of nitric oxide (NO) and prostacyclin (PGI(2)) in porcine aortic endothelial cells cultured and superfused on microcarriers.2. Saturable L-arginine transport (K-m = 0.14 +/- 0.03 mM; V-max = 2.08 +/- 0.54 nmol min(-1) (5 x 10(6) cells)(-1)) was pH insensitive and unaffected following removal of extracellular Na+ or Ca2+.3. Cationic arginine analogues, including L-lysine and L-ornithine, inhibited L-arginine transport, whilst 2-methylaminoisobutyric acid, beta-2-amino-bicyclo[2,2.1]-heptane-2-carboxylic acid, L-phenylalanine, 6-diazo-5-oxo-norleucine, L-glutamine, L-cysteine and L-glutamate were poor inhibitors.4. Deprivation of L-arginine (30 min to 24 h) reduced intracellular free L-arginine levels from 0.87 +/- 0.07 to 0.40 +/- 0.05 mM (P < 0.05) and resulted in a 40 % stimulation of L-arginine, L-lysine and L-ornithine transport.5. L-arginine and N-G-monomethyl-L-arginine (L-NMMA), but not N-omega-nitro-L-arginine methyl ester (L-NAME), trans-stimulated efflux of L-[H-3]arginine.6. Depolarization of endothelial cells with 70 mM K+ reduced L-arginine influx and prevented the stimulation of transport by 100 nM bradykinin, but agonist-induced release of NO and PGI(2) was still detectable.7. Basal rates of L-arginine transport and NO release were unaffected during superfusion of cells with a nominally Ca2+-free solution. Bradykinin-stimulated L-arginine transport was insensitive to removal of Ca2+, whereas agonist-induced NO release was abolished.8. Although bradykinin-stimulated NO release does not appear to be coupled directly to the transient increase in L-arginine transport, elevated rates of L-arginine influx via system y(+) in response to agonist-induced membrane hyperpolarization or substrate deprivation provide a mechanism for enhanced L-arginine supply to sustain NO generation.