EGFR kinase regulates volume-sensitive chloride current elicited by integrin stretch via PI-3K and NADPH oxidase in ventricular myocytes.
EGFR kinase regulates volume-sensitive chloride current elicited by integrin stretch via PI-3K and NADPH oxidase in ventricular myocytes.
复制标题
EGFR激酶可调节整合素通过PI-3K伸展而引起的体积敏感的氯化物电流，在心室心肌细胞中通过PI-3K和NADPH氧化酶。
DOI：
10.1085/jgp.200509366
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发表时间：
2006-03
影响因子：
3.8
通讯作者：
Baumgarten, Clive M
中科院分区：
文献类型：
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作者：
Browe, David M;Baumgarten, Clive M
β1 整合素的延伸通过 AT1 受体、NADPH 氧化酶和活性氧激活向外整流的他莫昔芬敏感的 Cl− 电流 (Cl− SAC)，并且 Cl− SAC 类似于体积敏感的 Cl− 电流 (ICl,swell)。表皮生长因子受体 (EGFR) 激酶在拉伸、整合素接合和 AT1 受体激活时发生反式激活，进而刺激 NADPH 氧化酶。因此，我们测试了 Cl− SAC 是否受 EGFR 激酶信号调节，并且是否具有体积敏感性。将涂有 β1 整合素单克隆抗体的顺磁珠附着到肌细胞上，并用电磁体拉动。拉伸激活 Cl− SAC，在 +40 mV 时为 1.13 ± 0.10 pA/pF。 AG1478 (10 μM) 是一种 EGFR 激酶阻断剂，可抑制 93 ± 13% 的 Cl− SAC，并且用 1 μM AG1478 进行细胞内预处理可显着抑制 Cl− SAC 激活。 EGF (3.3 nM) 直接激活向外整流的 Cl− 电流（+40 mV 时为 0.81 ± 0.05 pA/pF），该电流被 10 μM 他莫昔芬（一种 ICl 膨胀阻断剂）完全阻断。磷脂酰肌醇 3-激酶 (PI-3K) 是 EGFR 激酶的下游。 PI-3K 阻断剂渥曼青霉素 (500 nM) 和 LY294002 (100 μM) 分别抑制 Cl− SAC 67 ± 6% 和 91 ± 25%，并且 EGF 诱导的 Cl− 电流也被 LY294002 完全阻断。此外，gp91ds-tat (500 nM) 是一种可渗透细胞的嵌合肽，可特异性阻断 NADPH 氧化酶组装，可显着抑制 EGF 诱导的 Cl− 电流。无活性的渗透性和活性的不渗透性对照肽没有效果。高渗沐浴介质引起的心肌细胞收缩可分别抑制 Cl− SAC 和 EGF 诱导的 Cl− 电流 88 ± 9% 和 127 ± 11%。这些结果表明，β1 整合素拉伸通过 EGFR、PI-3K 和 NADPH 氧化酶激活 Cl− SAC，并且 Cl− SAC 和 EGF 诱导的 Cl− 电流很可能是体积敏感的 Cl− 电流，ICl,swell。
Stretch of β1 integrins activates an outwardly rectifying, tamoxifen-sensitive Cl− current (Cl− SAC) via AT1 receptors, NADPH oxidase, and reactive oxygen species, and Cl− SAC resembles the volume-sensitive Cl− current (ICl,swell). Epidermal growth factor receptor (EGFR) kinase undergoes transactivation upon stretch, integrin engagement, and AT1 receptor activation and, in turn, stimulates NADPH oxidase. Therefore, we tested whether Cl− SAC is regulated by EGFR kinase signaling and is volume sensitive. Paramagnetic beads coated with mAb for β1 integrin were attached to myocytes and pulled with an electromagnet. Stretch activated a Cl− SAC that was 1.13 ± 0.10 pA/pF at +40 mV. AG1478 (10 μM), an EGFR kinase blocker, inhibited 93 ± 13% of Cl− SAC, and intracellular pretreatment with 1 μM AG1478 markedly suppressed Cl− SAC activation. EGF (3.3 nM) directly activated an outwardly rectifying Cl− current (0.81 ± 0.05 pA/pF at +40 mV) that was fully blocked by 10 μM tamoxifen, an ICl,swell blocker. Phosphatidylinositol 3-kinase (PI-3K) is downstream of EGFR kinase. Wortmannin (500 nM) and LY294002 (100 μM), blockers of PI-3K, inhibited Cl− SAC by 67 ± 6% and 91 ± 25% respectively, and the EGF-induced Cl− current also was fully blocked by LY294002. Furthermore, gp91ds-tat (500 nM), a cell-permeable, chimeric peptide that specifically blocks NADPH oxidase assembly, profoundly inhibited the EGF-induced Cl− current. Inactive permeant and active impermeant control peptides had no effect. Myocyte shrinkage with hyperosmotic bathing media inhibited the Cl− SAC and EGF-induced Cl− current by 88 ± 9% and 127 ± 11%, respectively. These results suggest that β1 integrin stretch activates Cl− SAC via EGFR, PI-3K, and NADPH oxidase, and that both the Cl− SAC and the EGF-induced Cl− currents are likely to be the volume-sensitive Cl− current, ICl,swell.