Institute of Cellular and Molecular Physiology, Friedrich-Alexander University Erlangen-Nürnberg, Erlangen, Germany.
Division of Endocrinology, Diabetology, Vascular Disease, Nephrology and Clinical Chemistry, Department of Internal Medicine, University Hospital Tübingen, Tübingen, Germany.
Acta Physiol (Oxf). 2018 Sep;224(1):e13060. doi: 10.1111/apha.13060. Epub 2018 Mar 25.
Recent work has demonstrated that activation of the epithelial sodium channel (ENaC) by aberrantly filtered serine proteases causes sodium retention in nephrotic syndrome. The aim of this study was to elucidate a potential role of plasma kallikrein (PKLK) as a candidate serine protease in this context.
We analysed PKLK in the urine of patients with chronic kidney disease (CKD, n = 171) and investigated its ability to activate human ENaC expressed in Xenopus laevis oocytes. Moreover, we studied sodium retention in PKLK-deficient mice (klkb1 ) with experimental nephrotic syndrome induced by doxorubicin injection.
In patients with CKD, we found that PKLK is excreted in the urine up to a concentration of 2 μg mL which was correlated with albuminuria (r = .71) and overhydration as assessed by bioimpedance spectroscopy (r = .44). PKLK increased ENaC-mediated whole-cell currents, which was associated with the appearance of a 67 kDa γ-ENaC cleavage product at the cell surface consistent with proteolytic activation. Mutating a putative prostasin cleavage site in γ-ENaC prevented channel stimulation by PKLK. In a mouse model for nephrotic syndrome, active PKLK was present in nephrotic urine of klkb1 but not of klkb1 mice. However, klkb1 mice were not protected from ENaC activation and sodium retention compared to nephrotic klkb1 mice.
Plasma kallikrein is detected in the urine of proteinuric patients and mice and activates ENaC in vitro involving the putative prostasin cleavage site. However, PKLK is not essential for volume retention in nephrotic mice.
最近的研究表明，异常滤过的丝氨酸蛋白酶激活上皮钠离子通道（ENaC）可导致肾病综合征中的钠潴留。本研究旨在阐明血浆激肽释放酶（PKLK）作为候选丝氨酸蛋白酶在这种情况下的潜在作用。
我们分析了慢性肾脏病（CKD，n=171）患者尿液中的 PKLK，并研究了其激活在非洲爪蟾卵母细胞中表达的人 ENaC 的能力。此外，我们研究了用阿霉素注射诱导实验性肾病综合征的 PKLK 缺陷小鼠（klkb1）中的钠潴留。
在 CKD 患者中，我们发现 PKLK 排泄到尿液中的浓度高达 2μg/mL，与蛋白尿（r=.71）和生物电阻抗谱评估的过度水化（r=.44）相关。PKLK 增加了 ENaC 介导的全细胞电流，这与细胞表面出现 67kDa γ-ENaC 切割产物有关，这与蛋白水解激活一致。在 γ-ENaC 中突变一个假定的前丝氨酸切割位点可防止 PKLK 对通道的刺激。在肾病综合征的小鼠模型中，活性 PKLK 存在于 klkb1 但不存在于 klkb1 小鼠的肾病性尿液中。然而，与肾病性 klkb1 小鼠相比，klkb1 小鼠并没有从 ENaC 激活和钠潴留中得到保护。
PKLK 在蛋白尿患者和小鼠的尿液中被检测到，并通过假定的前丝氨酸切割位点在体外激活 ENaC。然而，PKLK 对于肾病小鼠的容量潴留并非必需。
