Extrinsic acidosis suppresses glycolysis and migration while increasing network formation in pulmonary microvascular endothelial cells
Extrinsic acidosis suppresses glycolysis and migration while increasing network formation in pulmonary microvascular endothelial cells
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DOI：
10.1152/ajplung.00544.2018
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发表时间：
2019-08-01
影响因子：
4.9
通讯作者：
Stevens, Troy
中科院分区：
文献类型：
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作者：
Lee, Ji Young;Onanyan, Mher;Stevens, Troy
酸中毒在危重患者中很常见，但目前纠正酸中毒的方法并不能改善疾病结局。在全身性酸中毒期间，细胞要么被动地暴露在其他细胞产生的胞外酸中毒(外源性酸中毒)中，要么暴露在它们产生的酸中并输出到细胞外空间(内源性酸中毒)。虽然已经研究了内源性酸中毒后的内皮修复，但外源性酸中毒对迁移和血管生成的影响尚不清楚。我们假设外源性酸中毒抑制肺微血管内皮细胞(PMVECs)的代谢和迁移，但促进毛细血管样网络的形成。外源性酸中毒通过滴定介质pH模拟。比较了两种类型的内源性酸中毒，包括通过化学抑制碳酸氢酶(CAS)IX和XII(SLC-0111)而增加细胞新陈代谢和低氧。在外源性和内源性酸中毒的情况下，PMVECs维持细胞内基础pH达24小时。外源性酸中毒可使全细胞CA IX蛋白表达降低，但不受缺氧的影响。当胞外pH相同时，外源性酸中毒抑制糖酵解，而内源性酸中毒不抑制糖酵解。外源性酸中毒抑制了迁移，但增加了Matrigel网络主连接和总片段长度。CRISPR-Cas9 CA IX基因敲除的PMVECs显示CA IX在促进糖酵解中的独立作用，因为CA IX的单独缺失伴随着己糖激酶I和丙酮酸脱氢酶L1a表达的降低和迁移的减少。2-脱氧-D-葡萄糖对细胞迁移无明显影响，但能显著抑制网络形成，增加N-钙粘附素的表达。因此，我们报告，虽然外源性酸中毒抑制内皮细胞糖酵解和迁移，但它促进了网络的形成。
Acidosis is common among critically ill patients, but current approaches to correct pH do not improve disease outcomes. During systemic acidosis, cells are either passively exposed to extracellular acidosis that other cells have generated (extrinsic acidosis) or they are exposed to acid that they generate and export into the extracellular space (intrinsic acidosis). Although endothelial repair following intrinsic acidosis has been studied, the impact of extrinsic acidosis on migration and angiogenesis is unclear. We hypothesized that extrinsic acidosis inhibits metabolism and migration but promotes capillary-like network formation in pulmonary microvascular endothelial cells (PMVECs). Extrinsic acidosis was modeled by titrating media pH. Two types of intrinsic acidosis were compared, including increasing cellular metabolism by chemically inhibiting carbonic anhydrases (CAs) IX and XII (SLC-0111) and with hypoxia. PMVECs maintained baseline intracellular pH for 24 h with both extrinsic and intrinsic acidosis. Whole cell CA IX protein expression was decreased by extrinsic acidosis but not affected by hypoxia. When extracellular pH was equally acidic, extrinsic acidosis suppressed glycolysis, whereas intrinsic acidosis did not. Extrinsic acidosis suppressed migration, but increased Matrigel network master junction and total segment length. CRISPR-Cas9 CA IX knockout PMVECs revealed an independent role of CA IX in promoting glycolysis, as loss of CA IX alone was accompanied by decreased hexokinase I and pyruvate dehydrogenase lila expression and decreasing migration. 2-deoxy-D-glucose bad no effect on migration but profoundly inhibited network formation and increased N-cadherin expression. Thus, we report that while extrinsic acidosis suppresses endothelial glycolysis and migration, it promotes network formation.