Quercetin Stimulates Na+/K+/2Cl- Cotransport via PTK-Dependent Mechanisms in Human Airway Epithelium
Quercetin Stimulates Na+/K+/2Cl- Cotransport via PTK-Dependent Mechanisms in Human Airway Epithelium
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DOI：
10.1165/rcmb.2008-0338oc
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发表时间：
2009-12-01
影响因子：
6.4
通讯作者：
Marunaka, Yoshinori
中科院分区：
文献类型：
--
作者：
Asano, Junji;Niisato, Naomi;Marunaka, Yoshinori
我们研究了Cl-分泌的调节机制，在维持人气道上皮Calu-3细胞的表面液体中起着至关重要的作用。本研究报道，槲皮素（一种黄酮类化合物）刺激布美他尼敏感性Cl-分泌，同时降低顶端Cl-传导，表明槲皮素通过激活Cl-通过Na+/K+/2Cl（-）协同转运蛋白（NKCC 1）穿过基底外侧膜的进入步骤来刺激Cl-分泌。为了阐明槲皮素刺激NKCC 1的机制，我们证实了蛋白激酶（PK）A，PKC，蛋白酪氨酸激酶（PTK）和胞质Ca 2+依赖性途径的参与。PKA抑制剂（PKI-14-22酰胺），PKC抑制剂（Go 6983）或Ca 2+螯合剂不影响槲皮素刺激的Cl-分泌。另一方面，PTK抑制剂（AG 18）显着减少了槲皮素对Cl-分泌的刺激作用，而对顶端Cl-电导没有抑制作用，表明PTK介导的途径参与了槲皮素的刺激作用。槲皮素对Cl-分泌的作用被布雷菲德菌素A抑制（BFA，从ER到高尔基体的囊泡转运的抑制剂），并且在表皮生长因子受体（EGFR）激酶抑制剂（AG 1478）的存在下未观察到BFA敏感的Cl-分泌，这表明槲皮素通过引起EGFR激酶介导的NKCC 1或NKC 1-NKCC 2的易位来刺激Cl-分泌。在人气道上皮Calu-3细胞中活化因子对基底外侧膜的作用。然而，槲皮素不增加NKCC 1的表面密度，但槲皮素提高NKCC 1的活性。这些观察结果表明，槲皮素通过EGFR激酶依赖性途径激活NKCC 1激活因子的转位激活NKCC 1，从而刺激Cl-分泌。
We investigated regulatory mechanisms of Cl- secretion playing an essential role in the maintenance of surface fluid in human airway epithelial Calu-3 cells. The present study reports that quercetin (a flavonoid) stimulated bumetanide-sensitive Cl- secretion with reduction of apical Cl- conductance, suggesting that quercetin Stimulates Cl- secretion by activating an entry step of Cl- across the basolateral membrane through Na+/K+/2Cl(-) cotransporter (NKCC1). To clarify the mechanism stimulating NKCC1 by quercetin, we verified involvement of protein kinase (PK)A, PKC, protein tyrosine kinase (PTK), and cytosolic Ca2+-dependent pathways. A PKA inhibitor (PKI-14-22 amide), a PKC inhibitor (Go 6983) or a Ca2+ chelating agent did not affect the quercetin-stimulated Cl- secretion. On the other hand, a PTK inhibitor (AG18) significantly diminished the stimulatory action of quercetin on Cl- secretion without inhibitory effects on apical Cl- conductance, suggesting that a PTK-mediated pathway is involved in the stimulatory action of quercetin. The quercetin action on Cl- secretion was suppressed with brefeldin A (BFA, an inhibitor of vesicular transport from ER to Golgi), and the BFA-sensitive Cl- secretion was not observed in the presence of an epidermal growth factor receptor (EGFR) kinase inhibitor (AG1478), suggesting that quercetin stimulates Cl- secretion by causing the EGFR kinase-mediated translocation of NKCC1 or an NKC1-activating factor to the basolateral membrane in human airway epithelial Calu-3 cells. However, the surface density of NKCC1 was not increased by quercetin, but quercetin elevated the activity of NKCC1. These observations indicate that quercetin stimulates Cl- secretion by activating NKCC1 via translocation of an NKCC1-activating factor through an EGFR kinase-dependent pathway.