Department of Internal Medicine, Hanyang University College of Medicine, Seoul, Republic of Korea.
Institute of Biomedical Science, Hanyang University College of Medicine, Seoul, Republic of Korea.
Am J Physiol Renal Physiol. 2021 Mar 1;320(3):F418-F428. doi: 10.1152/ajprenal.00282.2020. Epub 2021 Feb 1.
Urinary calcium and magnesium wasting is a characteristic feature of metabolic acidosis, and this study focused on the role of the thick ascending limb of Henle's loop in metabolic acidosis-induced hypercalciuria and hypermagnesiuria because thick ascending limb is an important site of paracellular calcium and magnesium reabsorption. Male Sprague-Dawley rats were used to determine the effects of acid loading (by adding NHCl, 7.2 mmol/220 g body wt/day to food slurry for 7 days) on renal expression of claudins and then to evaluate whether the results were reversed by antagonizing calcium-sensing receptor (using NPS-2143). At the end of each animal experiment, the kidneys were harvested for immunoblotting, immunofluorescence microscopy, and quantitative PCR (qPCR) analysis of claudins and the calcium-sensing receptor. As expected, NHCl loading lowered urinary pH and increased excretion of urinary calcium and magnesium. In NHCl-loaded rats, renal protein and mRNA expression of claudin-16, and claudin-19, were decreased compared with controls. However, claudin-14 protein and mRNA increased in NHCl-loaded rats. Consistently, the calcium-sensing receptor protein and mRNA were up-regulated in NHCl-loaded rats. All these changes were reversed by NPS-2143 coadministration and were confirmed using immunofluorescence microscopy. Hypercalciuria and hypermagnesiuria in NHCl-loaded rats were significantly ameliorated by NPS-2143 coadministration as well. We conclude that in metabolic acidosis, claudin-16 and claudin-19 in the thick ascending limb are down-regulated to produce hypercalciuria and hypermagnesiuria via the calcium-sensing receptor. This study found that the thick ascending limb of Henle's loop is involved in the mechanisms of hypercalciuria and hypermagnesiuria in metabolic acidosis. Specifically, expression of claudin-16/19 and claudin-14 was altered via up-regulation of calcium-sensing receptor in NHCl-induced metabolic acidosis. Our novel findings contribute to understanding the regulatory role of paracellular tight junction proteins in the thick ascending limb.
尿钙和尿镁丢失是代谢性酸中毒的特征性表现，本研究集中于探讨 Henle 袢升支粗段在代谢性酸中毒诱导的高钙尿和高镁尿中的作用，因为 Henle 袢升支粗段是细胞旁钙和镁重吸收的重要部位。本研究使用雄性 Sprague-Dawley 大鼠来确定酸化负荷（通过在食物糊剂中添加 7.2mmol/220g 体重/天的 NHCl 持续 7 天）对 Claudin 表达的影响，然后评估通过拮抗钙敏感受体（使用 NPS-2143）是否可以逆转这些结果。在每个动物实验结束时，采集肾脏进行免疫印迹、免疫荧光显微镜和 Claudin 和钙敏感受体的定量 PCR（qPCR）分析。正如预期的那样，NHCl 负荷降低了尿 pH 值并增加了尿钙和镁的排泄。在 NHCl 负荷大鼠中，Claudin-16 和 Claudin-19 的肾蛋白和 mRNA 表达与对照组相比降低。然而，Claudin-14 蛋白和 mRNA 在 NHCl 负荷大鼠中增加。一致地，钙敏感受体蛋白和 mRNA 在 NHCl 负荷大鼠中上调。NPS-2143 共给药逆转了所有这些变化，并通过免疫荧光显微镜得到证实。NHCl 负荷大鼠的高钙尿和高镁尿也明显改善。我们得出结论，在代谢性酸中毒中，Henle 袢升支粗段的 Claudin-16 和 Claudin-19 下调，通过钙敏感受体产生高钙尿和高镁尿。本研究发现，Henle 袢升支粗段在代谢性酸中毒中的高钙尿和高镁尿机制中发挥作用。具体而言，在 NHCl 诱导的代谢性酸中毒中，Claudin-16/19 和 Claudin-14 的表达通过钙敏感受体的上调而改变。我们的新发现有助于理解细胞旁紧密连接蛋白在 Henle 袢升支粗段中的调节作用。
