The University of Virginia Department of Pathology, Charlottesville, VA, United States of America.
The University of Virginia Department of Medicine, Charlottesville, VA, United States of America.
PLoS One. 2018 Apr 11;13(4):e0189464. doi: 10.1371/journal.pone.0189464. eCollection 2018.
Salt sensitivity of blood pressure affects >30% of the hypertensive and >15% of the normotensive population. Variants of the electrogenic sodium bicarbonate cotransporter NBCe2 gene, SLC4A5, are associated with increased blood pressure in several ethnic groups. SLC4A5 variants are also highly associated with salt sensitivity, independent of hypertension. However, little is known about how NBCe2 contributes to salt sensitivity, although NBCe2 regulates renal tubular sodium bicarbonate transport. We hypothesized that SLC4A5 rs10177833 and rs7571842 increase NBCe2 expression and human renal proximal tubule cell (hRPTC) sodium transport and may be a cause of salt sensitivity of blood pressure.
To characterize the hRPTC ion transport of wild-type (WT) and homozygous variants (HV) of SLC4A5.
The expressions of NBCe2 mRNA and protein were not different between hRPTCs carrying WT or HV SLC4A5 before or after dopaminergic or angiotensin (II and III) stimulation. However, luminal to basolateral sodium transport, NHE3 protein, and Cl-/HCO3- exchanger activity in hRPTCs were higher in HV than WT SLC4A5. Increasing intracellular sodium enhanced the apical location of NBCe2 in HV hRPTCs (4.24±0.35% to 11.06±1.72% (P<0.05, N = 3, 2-way ANOVA, Holm-Sidak test)) as determined by Total Internal Reflection Fluorescence Microscopy (TIRFM). In hRPTCs isolated from kidney tissue, increasing intracellular sodium enhanced bicarbonate-dependent pH recovery rate and increased NBCe2 mRNA and protein expressions to a greater extent in HV than WT SLC4A5 (+38.00±6.23% vs HV normal salt (P<0.01, N = 4, 2-way ANOVA, Holm-Sidak test)). In hRPTCs isolated from freshly voided urine, bicarbonate-dependent pH recovery was also faster in those from salt-sensitive and carriers of HV SLC4A5 than from salt-resistant and carriers of WT SLC4A5. The faster NBCe2-specific bicarbonate-dependent pH recovery rate in HV SCL4A5 was normalized by SLC4A5- but not SLC4A4-shRNA. The binding of purified hepatocyte nuclear factor type 4A (HNF4A) to DNA was increased in hRPTCs carrying HV SLC4A5 rs7571842 but not rs10177833. The faster NBCe2-specific bicarbonate-dependent pH recovery rate in HV SCL4A5 was abolished by HNF4A antagonists.
NBCe2 activity is stimulated by an increase in intracellular sodium and is hyper-responsive in hRPTCs carrying HV SLC4A5 rs7571842 through an aberrant HNF4A-mediated mechanism.
血压的盐敏感性影响超过 30%的高血压患者和超过 15%的血压正常人群。电生钠碳酸氢盐共转运蛋白 NBCe2 基因 SLC4A5 的变体与多个种族群体的血压升高有关。SLC4A5 变体也与盐敏感性高度相关，与高血压无关。然而，尽管 NBCe2 调节肾管状钠碳酸氢盐转运，但对于 NBCe2 如何导致盐敏感性知之甚少。我们假设 SLC4A5 的 rs10177833 和 rs7571842 增加了 NBCe2 的表达和人肾近端小管细胞（hRPTC）的钠转运，可能是血压盐敏感性的一个原因。
描述携带野生型（WT）和纯合变体（HV）SLC4A5 的 hRPTC 的离子转运。
在多巴胺能或血管紧张素（II 和 III）刺激前后，携带 WT 或 HV SLC4A5 的 hRPTC 中 NBCe2 mRNA 和蛋白的表达没有差异。然而，HV SLC4A5 的 hRPTC 中腔侧至基底外侧的钠转运、NHE3 蛋白和 Cl-/HCO3-交换器活性更高。用 Total Internal Reflection Fluorescence Microscopy（TIRFM）测定，增加细胞内钠会增加 HV hRPTC 中 NBCe2 的顶端位置（4.24±0.35%至 11.06±1.72%（P<0.05，N = 3，2 路 ANOVA，Holm-Sidak 检验））。在从肾组织分离的 hRPTC 中，增加细胞内钠会增强碳酸氢盐依赖性 pH 恢复率，并在 HV 比 WT SLC4A5 中更显著地增加 NBCe2 mRNA 和蛋白的表达（+38.00±6.23%与 HV 正常盐（P<0.01，N = 4，2 路 ANOVA，Holm-Sidak 检验））。在从新鲜排空的尿液中分离的 hRPTC 中，盐敏感性和 HV SLC4A5 携带者的碳酸氢盐依赖性 pH 恢复也更快。HV SCL4A5 中更快的 NBCe2 特异性碳酸氢盐依赖性 pH 恢复速率通过 SLC4A5 而不是 SLC4A4-shRNA 得到了正常化。HV SLC4A5 中增加的细胞内钠刺激 NBCe2 活性，并且通过异常的 HNF4A 介导的机制，HV SLC4A5 rs7571842 携带的 hRPTC 中 NBCe2 的反应更敏感。
通过异常的 HNF4A 介导机制，通过增加细胞内钠，HV SLC4A5 rs7571842 携带的 hRPTC 中 NBCe2 活性受到刺激，并且 NBCe2 特异性碳酸氢盐依赖性 pH 恢复率对盐的反应更敏感。
