DETANO and nitrated lipids increase chloride secretion across lung airway cells
DETANO and nitrated lipids increase chloride secretion across lung airway cells
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DOI：
10.1165/rcmb.2008-0005oc
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发表时间：
2008-08-01
影响因子：
6.4
通讯作者：
Matalon, Sadis
中科院分区：
文献类型：
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作者：
Chen, Lan;Bosworth, Charles A.;Matalon, Sadis
我们在体外和体内研究了一氧化氮 (NO) 增加肺上皮细胞氯 (Cl-) 分泌的细胞机制。将 (Z)-1-[2-(2-氨基乙基)-N-(2-氨乙基)氨基] diazen-1-ium-1, 2-二醇 (DETANONOate [DETANO]; 1-1,000 mu M) 添加到含有 Calu-3 细胞的 Ussing 室的顶端隔室中，使短路电流 (I-sc) 从 5.2 +/- 0.8 增加到 15.0 +/- 2.1 μA/cm(2)（X +/- 1 SE；n = 7；P < 0.001）。由两种硝化脂质（硝基亚油酸和硝基油酸；1-10 μM）产生的 NO 也使 Is 增加约 100%。在透化的 Calu-3 细胞的基底外侧（但顶部没有）中发现了类似的效果。在用10μM 1H-[1,2,4)恶二唑并[4,3-a]喹喔啉-1-酮（可溶性鸟苷酸环化酶抑制剂）预处理的Calu-3细胞中，这些NO供体均未增加Is。 NO 的清除可以阻止或逆转 1,c 的增加。这些数据表明，NO 对可溶性鸟苷酸环化酶的刺激对于通过刺激顶端囊性纤维化跨膜调节因子 (CFTR) 来增加 I-SC 是充分且必要的。 Calu-3 和肺泡 II 型 (ATII) 细胞均含有 CFTR，蛋白激酶 (PK) A 对免疫沉淀的 CFTR 进行体外磷酸化证明了这一点。PKGII（但不是 PKGI）磷酸化了 Calu-3 细胞免疫沉淀的 CFTR。 ATII 细胞中的相应值低于检测阈值。此外，DETANO、8-Br-cGMP 或 8-(4-氯苯硫基)-cGMP（每种最高 2 mM）在体外不会增加阿米洛利处理的 ATII 细胞的 Cl 分泌。对麻醉小鼠鼻腔电位差的测量表明，用 DETANO 灌注鼻孔可激活格列本脲敏感的 Cl 分泌。这些发现表明，小浓度的 NO 供体可能有利于刺激气道细胞的 Cl- 分泌，而不促进肺泡水肿。
We investigated the cellular mechanisms by which nitric oxide (NO) increases chloride (Cl-) secretion across lung epithelial cells in vitro and in vivo. Addition of (Z)-1-[2-(2-aminoethyl)-N-(2-ammonioethyl)amino] diazen-1-ium-1, 2-diolate (DETANONOate [DETANO]; 1-1,000 mu M) into apical compartments of Ussing chambers containing Calu-3 cells increased short-circuit currents (I-sc) from 5.2 +/- 0.8 to 15.0 +/- 2.1 mu A/cm(2) (X +/- 1 SE; n = 7; P < 0.001). NO generated from two nitrated lipids (nitrolinoleic and nitrooleic acids; 1-10 mu M) also increased Is, by about 100%. Similar effects were noted across basolaterally, but not apically, permeabilized Calu-3 cells. None of these NO donors increased Is, in Calu-3 cells pretreated with 10 mu M 1H-[1,2,4)oxadiazolo[4,3-a]quinoxalin-1-one (an inhibitor of soluble guanylyl cyclase). Scavenging of NO either prevented or reversed the increase of 1,c. These data indicate that NO stimulation of soluble guanylyl cyclase was sufficient and necessary for the increase of I-SC via stimulation of the apical cystic fibrosis transmembrane regulator (CFTR). Both Calu-3 and alveolar type II (ATII) cells contained CFTR, as demonstrated by in vitro phosphorylation of immunoprecipitated CFTR by protein kinase (PK) A. PKGII (but not PKGI) phosphorylated CFTR immuniprecipitated from Calu-3 cells. Corresponding values in ATII cells were below the threshold of detection. Furthermore, DETANO, 8-Br-cGMP, or 8-(4-chlorophenylthio)-cGMP (up to 2 mM each) did not increase Cl- secretion across amiloride-treated ATII cells in vitro. Measurements of nasal potential differences in anesthetized mice showed that perfusion of the nares with DETANO activated glybenclamide-sensitive Cl- secretion. These findings suggest that small concentrations of NO donors may prove beneficial in stimulating Cl- secretion across airway cells without promoting alveolar edema.