Hypercapnia Impairs ENaC Cell Surface Stability by Promoting Phosphorylation, Polyubiquitination and Endocytosis of β-ENaC in a Human Alveolar Epithelial Cell Line.
Hypercapnia Impairs ENaC Cell Surface Stability by Promoting Phosphorylation, Polyubiquitination and Endocytosis of β-ENaC in a Human Alveolar Epithelial Cell Line.
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DOI：
10.3389/fimmu.2017.00591
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发表时间：
2017
影响因子：
7.3
通讯作者：
Vadász I
中科院分区：
文献类型：
--
作者：
Gwoździńska P;Buchbinder BA;Mayer K;Herold S;Morty RE;Seeger W;Vadász I
急性肺损伤与导致气体交换受损的肺水肿的形成有关。急性呼吸窘迫综合征（ARDS）患者需要机械通气来改善氧合;然而，使用相对较低的潮气量（以尽量减少肺的进一步损伤）通常会导致二氧化碳的进一步积聚（高碳酸血症）。高碳酸血症已被证明会损害肺泡液体清除率（AFC），从而导致肺水肿潴留，并可能导致更差的结局;然而，其潜在的分子机制仍不完全清楚。AFC严重依赖于上皮钠通道（ENaC），其驱动Na+跨肺泡上皮的矢量转运。因此，在目前的研究中，我们研究了高碳酸血症影响肺泡上皮细胞（AEC）中ENaC细胞表面稳定性的机制。升高的CO2水平导致AEC中β-ENaC的多泛素化和随后的α/β-ENaC复合物的内吞作用，这通过沉默E3泛素连接酶Nedd 4 -2来防止。高碳酸血症诱导的ENaC的泛素化和细胞表面修复严重依赖于β-ENaC的Thr 615残基的磷酸化，这是由细胞外信号调节激酶（ERK）1/2介导的。此外，ERK 1/2的激活导致随后的AMP活化蛋白激酶（AMPK）和c-Jun N-末端激酶（JNK）1/2的激活，这又使Nedd 4 -2在Thr 899残基处磷酸化。重要的是，Thr 899突变为Ala显著抑制了CO2诱导的β-ENaC的多泛素化，并恢复了ENaC复合物的细胞表面稳定性，突出了Nedd 4 -2磷酸化状态在靶向ENaC中的关键作用。总的来说，我们的数据表明，CO2水平升高促进人AEC系中ERK/AMPK/JNK轴的激活，其中ERK 1/2磷酸化β-ENaC，而JNK介导Nedd 4 -2的磷酸化，从而促进通道-连接酶相互作用。高碳酸血症诱导的ENaC功能障碍可能导致肺泡水肿清除受损，因此，干预这些分子机制可能会改善肺泡液体平衡，并导致ARDS患者的结局更好。
Acute lung injury is associated with formation of pulmonary edema leading to impaired gas exchange. Patients with acute respiratory distress syndrome (ARDS) require mechanical ventilation to improve oxygenation; however, the use of relatively low tidal volumes (to minimize further injury of the lung) often leads to further accumulation of carbon dioxide (hypercapnia). Hypercapnia has been shown to impair alveolar fluid clearance (AFC), thereby causing retention of pulmonary edema, and may lead to worse outcomes; however, the underlying molecular mechanisms remain incompletely understood. AFC is critically dependent on the epithelial sodium channel (ENaC), which drives the vectorial transport of Na+ across the alveolar epithelium. Thus, in the current study, we investigated the mechanisms by which hypercapnia effects ENaC cell surface stability in alveolar epithelial cells (AECs). Elevated CO2 levels led to polyubiquitination of β-ENaC and subsequent endocytosis of the α/β-ENaC complex in AECs, which were prevented by silencing the E3 ubiquitin ligase, Nedd4-2. Hypercapnia-induced ubiquitination and cell surface retrieval of ENaC were critically dependent on phosphorylation of the Thr615 residue of β-ENaC, which was mediated by the extracellular signal-regulated kinase (ERK)1/2. Furthermore, activation of ERK1/2 led to subsequent activation of AMP-activated protein kinase (AMPK) and c-Jun N-terminal kinase (JNK)1/2 that in turn phosphorylated Nedd4-2 at the Thr899 residue. Importantly, mutation of Thr899 to Ala markedly inhibited the CO2-induced polyubiquitination of β-ENaC and restored cell surface stability of the ENaC complex, highlighting the critical role of Nedd4-2 phosphorylation status in targeting ENaC. Collectively, our data suggest that elevated CO2 levels promote activation of the ERK/AMPK/JNK axis in a human AEC line, in which ERK1/2 phosphorylates β-ENaC whereas JNK mediates phosphorylation of Nedd4-2, thereby facilitating the channel–ligase interaction. The hypercapnia-induced ENaC dysfunction may contribute to impaired alveolar edema clearance and thus, interfering with these molecular mechanisms may improve alveolar fluid balance and lead to better outcomes in patients with ARDS.