Effect of hypoxic exposure on Na+/H+ antiport activity, isoform expression, and localization in endothelial cells
Effect of hypoxic exposure on Na+/H+ antiport activity, isoform expression, and localization in endothelial cells
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DOI：
10.1152/ajplung.1998.275.3.l442
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发表时间：
1998-09-01
影响因子：
4.9
通讯作者：
Sun, AM
中科院分区：
文献类型：
--
作者：
Cutaia, MV;Parks, N;Sun, AM
关于长期缺氧暴露对膜离子运输活性的影响，我们所知甚少。Na+/H+反端口是调节哺乳动物细胞内pH值的离子转运位点。我们测定了长时间缺氧暴露对人肺动脉内皮细胞抗转运活性、基因表达和定位的影响。单层细胞在缺氧或常氧条件下孵育72小时。以细胞内酸中毒的恢复率来测定抗转运活性。采用RT-PCR、Northern和Western blots检测逆转录异构体鉴定和基因表达。免疫荧光染色确定反端口定位和f -肌动蛋白细胞骨架组织。与常氧对照细胞相比，长时间的低氧暴露降低了抗氧化活性，但细胞活力没有变化。在人肺动脉内皮细胞中存在一种定位于基底外侧细胞表面的反端口异构体[Na+/H+交换异构体（NHE） 1]。低氧暴露对NHE1 mRNA转录物表达无影响，但NHE1蛋白表达上调。免疫荧光染色显示缺氧后f -肌动蛋白细胞骨架有明显改变，但NHE1定位没有变化。这些结果表明，长期缺氧暴露后NHE1活性的降低与基因表达的改变无关。NHE1活性的改变可能对血管功能有重要影响。
Little is known about the effects of prolonged hypoxic exposure on membrane ion transport activity. The Na+/H+ antiport is an ion transport site that regulates intracellular pH in mammalian cells. We determined the effect of prolonged hypoxic exposure on human pulmonary arterial endothelial cell antiport activity, gene expression, and localization. Monolayers were incubated under hypoxic or normoxic conditions for 72 h. Antiport activity was determined as the rate of recovery from intracellular acidosis. Antiport isoform identification and gene expression were determined with RT-PCR and Northern and Western blots. Antiport localization and F-actin cytoskeleton organization were defined with immunofluorescent staining. Prolonged hypoxic exposure decreased antiport activity, with no change in cell viability compared with normoxic control cells. One antiport isoform [Na+/H+ exchanger isoform (NHE) 1] that was localized to the basolateral cell surface was present in human pulmonary arterial endothelial cells. Hypoxic exposure had no effect on NHE1 mRNA transcript expression, but NHE1 protein expression was upregulated. Immunofluorescent staining demonstrated a significant alteration of the F-actin cytoskeleton after hypoxic exposure but no change in NHE1 localization. These results demonstrate that the decrease in NHE1 activity after prolonged hypoxic exposure is not related to altered gene expression. The change in NHE1 activity may have important consequences for vascular function.