Wu Karen L, Khan Shenaz, Lakhe-Reddy Sujata, Wang Liming, Jarad George, Miller R Tyler, Konieczkowski Martha, Brown Arthur M, Sedor John R, Schelling Jeffrey R
Department of Medicine, and Rammelkamp Center for Education and Research, Case Western Reserve University School of Medicine, Cleveland, Ohio 44109, USA.
Am J Physiol Renal Physiol. 2003 Apr;284(4):F829-39. doi: 10.1152/ajprenal.00314.2002. Epub 2002 Nov 26.
Renal tubular epithelial cell (RTC) apoptosis causes tubular atrophy, a hallmark of renal disease progression. Apoptosis is generally characterized by reduced cell volume and cytosolic pH, but epithelial cells are relatively resistant to shrinkage due to regulatory volume increase, which is mediated by Na(+)/H(+) exchanger (NHE) 1. We investigated whether RTC apoptosis requires caspase cleavage of NHE1. Staurosporine- and hypertonic NaCl-induced RTC apoptosis was associated with cell shrinkage and diminished cytosolic pH, and apoptosis was potentiated by amiloride analogs, suggesting NHE1 activity opposes apoptosis. NHE1-deficient fibroblasts demonstrated increased susceptibility to apoptosis, which was reversed by NHE1 reconstitution. NHE1 expression was markedly decreased in apoptotic RTC due to degradation, and preincubation with peptide caspase antagonists restored NHE1 expression, indicating that NHE1 is degraded by caspases. Recombinant caspase-3 cleaved the in vitro-translated NHE1 cytoplasmic domain into five distinct peptides, identical in molecular weight to NHE1 degradation products derived from staurosporine-stimulated RTC lysates. In vivo, NHE1 loss-of-function C57BL/6.SJL-swe/swe mice with adriamycin-induced nephropathy demonstrated increased RTC apoptosis compared with adriamycin-treated wild-type controls, thereby implicating NHE1 inactivation as a potential mechanism of tubular atrophy. We conclude that NHE1 activity is critical for RTC survival after injury and that caspase cleavage of RTC NHE1 may promote apoptosis and tubular atrophy by preventing compensatory intracellular volume and pH regulation.
肾小管上皮细胞（RTC）凋亡导致肾小管萎缩，这是肾脏疾病进展的一个标志。凋亡通常表现为细胞体积减小和胞质pH值降低，但上皮细胞由于由钠/氢交换体（NHE）1介导的调节性容积增加而相对抵抗细胞收缩。我们研究了RTC凋亡是否需要半胱天冬酶切割NHE1。星形孢菌素和高渗氯化钠诱导的RTC凋亡与细胞收缩和胞质pH值降低相关，并且凋亡被阿米洛利类似物增强，提示NHE1活性对抗凋亡。NHE1缺陷的成纤维细胞对凋亡的敏感性增加，而NHE1的重新构建可使其逆转。由于降解，凋亡的RTC中NHE1表达明显降低，用肽半胱天冬酶拮抗剂预孵育可恢复NHE1表达，表明NHE1被半胱天冬酶降解。重组半胱天冬酶-3将体外翻译的NHE1胞质结构域切割成五个不同的肽段，其分子量与来自星形孢菌素刺激的RTC裂解物的NHE1降解产物相同。在体内，与阿霉素处理的野生型对照相比，患有阿霉素诱导的肾病的NHE1功能丧失的C57BL/6.SJL-swe/swe小鼠表现出RTC凋亡增加，从而提示NHE1失活是肾小管萎缩的潜在机制。我们得出结论，NHE1活性对于损伤后RTC的存活至关重要，并且RTC NHE1的半胱天冬酶切割可能通过阻止细胞内容积和pH的代偿性调节而促进凋亡和肾小管萎缩。
