Division of Nephrology, Department of Medicine, Stanford University, Palo Alto, California; and.
Am J Physiol Cell Physiol. 2014 Feb 1;306(3):C263-78. doi: 10.1152/ajpcell.00381.2012. Epub 2013 Nov 27.
Under conditions of high dietary salt intake, prostaglandin E2 (PGE2) production is increased in the collecting duct and promotes urinary sodium chloride (NaCl) excretion; however, the molecular mechanisms by which PGE2 increases NaCl excretion in this context have not been clearly defined. We used the mouse inner medullary collecting duct (mIMCD)-K2 cell line to characterize mechanisms underlying PGE2-regulated NaCl transport. When epithelial Na(+) channels were inhibited, PGE2 exclusively stimulated basolateral EP4 receptors to increase short-circuit current (Isc(PGE2)). We found that Isc(PGE2) was sensitive to inhibition by H-89 and CFTR-172, indicating that EP4 receptors signal through protein kinase A to induce Cl(-) secretion via cystic fibrosis transmembrane conductance regulator (CFTR). Unexpectedly, we also found that Isc(PGE2) was sensitive to inhibition by BAPTA-AM (Ca(2+) chelator), 2-aminoethoxydiphenyl borate (2-APB) (inositol triphosphate receptor blocker), and flufenamic acid (FFA) [Ca(2+)-activated Cl(-) channel (CACC) inhibitor], suggesting that EP4 receptors also signal through Ca(2+) to induce Cl(-) secretion via CACC. Additionally, we observed that PGE2 stimulated an increase in Isc through crosstalk between cAMP and Ca(2+) signaling; BAPTA-AM or 2-APB inhibited a component of Isc(PGE2) that was sensitive to CFTR-172 inhibition; H-89 inhibited a component of Isc(PGE2) that was sensitive to FFA inhibition. Together, our findings indicate that PGE2 activates basolateral EP4 receptors and signals through both cAMP and Ca(2+) to stimulate Cl(-) secretion in IMCD-K2 cells. We propose that these signaling pathways, and the crosstalk between them, may provide a concerted mechanism for enhancing urinary NaCl excretion under conditions of high dietary NaCl intake.
在高盐饮食条件下，前列腺素 E2（PGE2）在集合管中的产生增加，促进尿氯化钠（NaCl）排泄；然而，在这种情况下，PGE2 增加 NaCl 排泄的分子机制尚未明确界定。我们使用小鼠内髓集合管（mIMCD）-K2 细胞系来表征 PGE2 调节的 NaCl 转运的机制。当上皮钠通道被抑制时，PGE2 仅刺激基底外侧 EP4 受体以增加短路电流（Isc（PGE2））。我们发现 Isc（PGE2）对 H-89 和 CFTR-172 的抑制敏感，表明 EP4 受体通过蛋白激酶 A 信号传导，通过囊性纤维化跨膜电导调节剂（CFTR）诱导 Cl（-）分泌。出乎意料的是，我们还发现 Isc（PGE2）对 BAPTA-AM（Ca（2+）螯合剂）、2-氨基乙氧基二苯硼酸盐（2-APB）（三磷酸肌醇受体阻滞剂）和氟芬那酸（FFA）[Ca（2+）激活的 Cl（-）通道（CACC）抑制剂]的抑制敏感，表明 EP4 受体也通过 Ca（2+）信号传导，通过 CACC 诱导 Cl（-）分泌。此外，我们观察到 PGE2 通过 cAMP 和 Ca（2+）信号转导的串扰刺激 Isc 的增加；BAPTA-AM 或 2-APB 抑制对 CFTR-172 抑制敏感的 Isc（PGE2）的一部分；H-89 抑制对 FFA 抑制敏感的 Isc（PGE2）的一部分。总之，我们的研究结果表明，PGE2 激活基底外侧 EP4 受体，并通过 cAMP 和 Ca（2+）信号转导刺激 IMCD-K2 细胞中的 Cl（-）分泌。我们提出，这些信号通路及其相互作用可能为高盐饮食条件下增强尿 NaCl 排泄提供协同机制。
