Department of Physiology and Cell Biology, University South Alabama, College of Medicine, Mobile, Alabama, United States of America.
Center for Lung Biology, University of South Alabama, College of Medicine, Mobile, Alabama, United States of America.
PLoS One. 2022 May 3;17(5):e0266890. doi: 10.1371/journal.pone.0266890. eCollection 2022.
Sodium-hydrogen exchangers (NHEs) tightly regulate intracellular pH (pHi), proliferation, migration and cell volume. Heterogeneity exists between pulmonary endothelial cells derived from different vascular segments, yet the activity and isoform expression of NHEs between these vascular segments has not been fully examined. Utilizing the ammonium-prepulse and recovery from acidification technique in a buffer lacking bicarbonate, pulmonary microvascular and pulmonary artery endothelial cells exhibited unique recovery rates from the acid load dependent upon the concentration of the sodium transport inhibitor, amiloride; further, pulmonary artery endothelial cells required a higher dose of amiloride to inhibit sodium-dependent acid recovery compared to pulmonary microvascular endothelial cells, suggesting a unique complement of NHEs between the different endothelial cell types. While NHE1 has been described in pulmonary endothelial cells, all NHE isoforms have not been accounted for. To address NHE expression in endothelial cells, qPCR was performed. Using a two-gene normalization approach, Sdha and Ywhag were identified for qPCR normalization and analysis of NHE isoforms between pulmonary microvascular and pulmonary artery endothelial cells. NHE1 and NHE8 mRNA were equally expressed between the two cell types, but NHE5 expression was significantly higher in pulmonary microvascular versus pulmonary artery endothelial cells, which was confirmed at the protein level. Thus, pulmonary microvascular and pulmonary artery endothelial cells exhibit unique NHE isoform expression and have a unique response to acid load revealed through recovery from cellular acidification.
钠氢交换器（NHEs）可严密调节细胞内 pH 值（pHi）、增殖、迁移和细胞体积。源自不同血管段的肺内皮细胞之间存在异质性，但这些血管段之间的 NHE 活性和同工型表达尚未得到充分研究。利用缓冲液中缺乏碳酸氢盐的铵脉冲和酸化恢复技术，肺微血管内皮细胞和肺动脉内皮细胞表现出独特的从酸化负荷中恢复的速率，这取决于钠转运抑制剂阿米洛利的浓度；此外，与肺微血管内皮细胞相比，肺动脉内皮细胞需要更高剂量的阿米洛利来抑制钠依赖性酸恢复，这表明不同内皮细胞类型之间存在独特的 NHE 互补体。虽然已在肺内皮细胞中描述了 NHE1，但并非所有 NHE 同工型都已被考虑在内。为了研究内皮细胞中的 NHE 表达，进行了 qPCR。使用双基因归一化方法，确定了 Sdha 和 Ywhag 用于 qPCR 归一化以及肺微血管和肺动脉内皮细胞之间 NHE 同工型的分析。两种细胞类型之间的 NHE1 和 NHE8 mRNA 表达水平相等，但 NHE5 的表达水平在肺微血管内皮细胞中明显高于肺动脉内皮细胞，这在蛋白质水平上得到了证实。因此，肺微血管和肺动脉内皮细胞表现出独特的 NHE 同工型表达，并通过从细胞酸化中恢复揭示了对酸负荷的独特反应。
