CFTR-mediated chloride permeability is regulated by type III phosphodiesterases in airway epithelial cells.
CFTR-mediated chloride permeability is regulated by type III phosphodiesterases in airway epithelial cells.
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CFTR 介导的氯离子通透性由气道上皮细胞中的 III 型磷酸二酯酶调节。
DOI：
10.1165/ajrcmb.13.6.7576703
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发表时间：
1995
期刊：
影响因子：
--
通讯作者：
Drumm,ML
中科院分区：
文献类型：
--
作者：
Kelley,TJ;al-Nakkash,L;Drumm,ML
囊性纤维化跨膜传导调节剂（CFTR）的氯离子通道活性需要3'-5'-环腺苷单磷酸（cAMP）激活蛋白激酶A （PKA）。cAMP的水平受cAMP合成和腺苷酸环化酶和磷酸二酯酶（PDEs）水解之间的平衡控制。在Calu-3和16HBE细胞中，CFTR通道活性似乎对III型环核苷酸PDEs的活性最敏感，这两种细胞都来源于气道上皮，表达野生型CFTR。III型PDEs可通过其对特定抑制剂（如米立酮和氨立酮）的敏感性来识别。在Calu-3细胞中，III型PDE的特异性抑制使氯化物外排增加13.7倍，而罗利普兰、Ro20-1724 （IV型PDE抑制剂）和3-异丁基-1-甲基黄嘌呤（IBMX，一种非特异性PDE抑制剂）均未引起显著增加。这些化合物对总细胞cAMP水平都没有明显的影响，然而米立酮和氨立酮对氯离子外排的影响被rp -cAMP（一种cAMP类似物，在cAMP结合位点抑制PKA）处理的细胞阻断。同样，PKA抑制剂H-8也能减少米立酮刺激的氯离子外排，表明外排是通过cAMP/PKA途径介导的。全细胞膜片钳分析显示，米立酮产生的氯离子电导与CFTR的性质一致。在没有腺苷酸环化酶激动剂的情况下，米立酮以剂量依赖性的方式激发氯离子电流并诱导CFTR活性。这些数据表明III型PDE特异性参与气道上皮细胞中CFTR的激活，PDE对CFTR的调节可能涉及cAMP的亚细胞区室。
Chloride channel activity of cystic fibrosis transmembrane conductance regulator (CFTR) requires activation of protein kinase A (PKA) by 3'-5'-cyclic adenosine monophosphate (cAMP). The level of cAMP is controlled by the balance between cAMP synthesis and hydrolysis by adenylate cyclase and phosphodiesterases (PDEs), respectively. CFTR channel activity appears to be most sensitive to the activity of type III cyclic nucleotide PDEs in Calu-3 and 16HBE cells, both derived from airway epithelium and expressing wild-type CFTR. Type III PDEs can be identified by their sensitivity to specific inhibitors such as milrinone and amrinone. In Calu-3 cells, specific inhibition of type III PDEs increased chloride efflux up to 13.7-fold, whereas neither rolipram nor Ro20-1724 (type IV PDE inhibitors) nor 3-isobutyl-1-methylxanthine (IBMX, a nonspecific PDE inhibitor) elicited significant increases. None of these compounds had an appreciable effect on total cellular cAMP levels, yet the effects of milrinone and amrinone on chloride efflux were blocked by treatment of cells with Rp-cAMPS, a cAMP analog that inhibits PKA at the site of cAMP binding. Similarly, H-8, an inhibitor of PKA, reduced milrinone-stimulated chloride efflux, indicating that efflux is mediated through the cAMP/PKA pathway. Whole-cell patch clamp analysis revealed that milrinone generated chloride conductances with properties consistent with those of CFTR. Milrinone elicited chloride currents in a dose-dependent manner and induced CFTR activity in the absence of adenylate cyclase agonists. These data suggest that type III PDEs are specifically involved in CFTR activation in airway epithelial cells and that PDE regulation of CFTR may involve subcellular compartments of cAMP.