Effect of sildenafil on cyclic nucleotide phosphodiesterase activity, vascular tone and calcium signaling in rat pulmonary artery
Effect of sildenafil on cyclic nucleotide phosphodiesterase activity, vascular tone and calcium signaling in rat pulmonary artery
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DOI：
10.1038/sj.bjp.0705277
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发表时间：
2003-06-01
影响因子：
7.3
通讯作者：
Savineau, JP
中科院分区：
文献类型：
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作者：
Pauvert, O;Lugnier, C;Savineau, JP
1西地那非（伟哥）是一种强效的PDE 5抑制剂，因此是阴茎海绵体平滑肌的松弛药物。在目前的工作中，我们证明了PDE 5同工酶的存在，并研究了西地那非对大鼠主肺动脉（MPA）中特定环核苷酸磷酸二酯酶（PDE）活性、平滑肌张力和钙信号的影响。2测定了胞浆和微粒体组分中的PDE活性。总cAMP和cGMP-PDE活性主要存在于胞浆组分中。西地那非（0.1 μ M）使cGMP-PDE活性降低了72%，而扎匹司特（10 μ M），一种相对选择性的PDE 5抑制剂，使该活性降低了63%。西地那非（0.1 μ M）也显著抑制（22%）cAMP-PDE活性。3蛋白质印迹分析显示，PDE 5的表达主要在MPA的胞浆部分。西地那非浓度依赖性地抑制（IC 50 = 3.4 nM）通过HPLC部分纯化的MPA PDE 5的活性。4西地那非（0.1 nM - 50 μ M）浓度依赖性地松弛用苯丙氨酸（0.5 μ M）预收缩的MPA环。西地那非的效力（IC 50 = 11 nM）与一氧化氮供体硝普钠相似，但高于扎普司特（IC 50 = 600 nM）。西地那非的血管舒张作用不受内皮去除或在KT 5823（1 μ M）和H89（1 μ M）存在下的改变，KT 5823和H89分别是PKG和PKA的有效抑制剂。5在已加载钙荧光团indo-1的分离的MPA肌细胞中，昔多芬（10 - 100 μ M）拮抗ATP和内皮素-1诱导的钙振荡，但对咖啡因诱导的瞬时[Ca 2 +]（i）无影响。6本研究证明了大鼠MPA中存在功能性和高度西地那非敏感的PDE 5同工酶。这种同工酶的抑制主要是西地那非强效肺血管扩张作用的原因，这涉及三磷酸肌醇介导的钙信号通路的改变。
1 Sildenafil (viagra) is a potent PDE5 inhibitor and thus a relaxant drug in corpus carvernosum smooth muscle. In the present work, we evidenced the presence of PDE5 isozyme and investigated the effect of sildenafil on the specific cyclic nucleotide phosphodiesterase (PDE) activity, smooth muscle tone and calcium signaling in the rat main pulmonary artery (MPA).2 The PDE activity was measured in cytosolic and microsomal fractions. Total cAMP and cGMP-PDE activities were mainly present in the cytosolic fraction. Sildenafil (0.1 muM) reduced by 72% cGMP-PDE activity, whereas zaprinast (10 muM), a relatively selective PDE5 inhibitor, reduced this activity by 63%. Sildenafil (0.1 muM) also inhibited significantly (22%) the cAMP-PDE activity.3 Western blot analysis revealed the expression of PDE5 mainly in the cytosolic fraction of MPA. Sildenafil concentration-dependently inhibited (IC50 = 3.4 nM) the activity of MPA PDE5 partially purified by HPLC.4 Sildenafil (0.1 nM - 50 muM) concentration-dependently relaxed MPA rings precontracted with phenylephrine (0.5 muM). The potency of sildenafil (IC50 = 11 nM) was similar to that of a nitric oxide donor, sodium nitroprusside, but higher than that of zaprinast (IC50 = 600 nM). The vasorelaxant effect of sildenafil was not altered by endothelium removal or in the presence of KT 5823 (1 muM) and H89 (1 muM), potent inhibitors of PKG and PKA, respectively.5 In isolated MPA myocytes, which had been loaded with the calcium fluorophore indo-1, sildenafil (10 - 100 muM) antagonized ATP- and endothelin-1-induced calcium oscillations but had no effect on the transient caffeine-induced [Ca2+](i) response.6 This study demonstrates the presence of a functional and highly sildenafil-sensitive PDE5 isozyme in rat MPA. Inhibition of this isozyme mainly accounts for the potent pulmonary vasodilator action of sildenafil, which involves alteration in the inositol triphosphate-mediated calcium signaling pathway.