Purinergic axis in cardiac blood vessels. Agonist-mediated release of ATP from cardiac endothelial cells.
Purinergic axis in cardiac blood vessels. Agonist-mediated release of ATP from cardiac endothelial cells.
复制标题
心脏血管中的嘌呤能轴。
DOI：
10.1161/01.res.74.3.401
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发表时间：
1994
影响因子：
20.1
通讯作者：
Buxton,IL
中科院分区：
文献类型：
--
作者：
Yang,S;Cheek,DJ;Westfall,DP;Buxton,IL
通过酶灌注从豚鼠心脏分离的纯化内皮细胞在单层培养中生长，并用于测试各种血管活性剂刺激这些细胞释放ATP的能力。用肽激动剂缓激肽（1 nmol/L）、乙酰胆碱（1 mumol/L）、5-羟色胺（1 mumol/L）或腺苷5 '-二磷酸（10 mumol/L）刺激内皮细胞，导致孵育培养基中ATP的快速出现，用萤火虫荧光素酶测定ATP。加入毒蕈碱（阿托品，0.1 μ mol/L）和嘌呤能（苏拉明，100 μ mol/L;活性蓝-2，100 μ mol/L）受体的拮抗剂表明，这些细胞的ATP释放是受体介导的。缓激肽诱导的ATP释放是快速的（在3 nmol/L缓激肽时峰值< 30秒），具有剂量依赖性（EC 50，0.18 nmol/L），并随着激动剂的重复给药而减弱。对缓激肽的脱敏也影响乙酰胆碱诱导释放的能力，并且当细胞短期恢复生长条件时是可逆的。通过高效液相色谱法测量作为其荧光N6-乙烯基嘌呤衍生物释放的腺苷嘌呤，揭示了释放的嘌呤的来源是ATP，并证实了其快速去磷酸化。向内皮细胞培养物中添加嘌呤核苷酸类似物2-甲硫基-ATP（2-甲基-S-ATP）、ADP和β-γ-亚甲基ATP，导致在30秒时在浸泡细胞的培养基中测量的ATP出现剂量依赖性增加，表明存在ATP诱导的ATP释放。（250字处删节）
Purified endothelial cells isolated from guinea pig hearts by enzymatic perfusion were grown in monolayer culture and used to test the ability of a variety of vasoactive agents to stimulate ATP release from these cells. Stimulation of endothelial cells with the peptide agonist bradykinin (1 nmol/L), acetylcholine (1 mumol/L), serotonin (1 mumol/L), or adenosine 5'-diphosphate (10 mumol/L) resulted in the rapid appearance of ATP in the incubation medium determined with the firefly luciferase assay for ATP. Addition of antagonists for muscarinic (atropine, 0.1 mumol/L) and purinergic (suramin, 100 mumol/L; reactive blue-2, 100 mumol/L) receptors suggested that ATP release from these cells was receptor-mediated. Bradykinin-induced release of ATP was rapid (peak < 30 seconds at 3 nmol/L bradykinin), dose-dependent (EC50, 0.18 nmol/L), and diminished with repeated administration of agonist. Desensitization to bradykinin also affected the ability of acetylcholine to induce release and was reversible when cells were returned to growth conditions for short periods. Measurement of released adenyl purines as their fluorescent N6-ethenopurine derivatives by high-performance liquid chromatography revealed the origin of the purine released to be ATP and confirmed its rapid dephosphorylation. Addition of the purine nucleotide analogues 2-methylthio-ATP (2-methyl-S-ATP), ADP, and beta gamma-methylene ATP to endothelial cell cultures resulted in a dose-dependent increase in the appearance of ATP measured in the medium bathing the cells at 30 seconds, suggesting the presence of ATP-induced ATP release.(ABSTRACT TRUNCATED AT 250 WORDS)