Gallazzini Morgan, Karim Zoubida, Bichara Maurice
INSERM U426, Faculté de Médecine Xavier Bichat, et Université Paris 7, Paris, France.
Nephron Physiol. 2006;104(4):126-35. doi: 10.1159/000095855. Epub 2006 Sep 22.
The present study assessed the mechanisms by which hypertonicity caused by NaCl enhances the renal outer medullary potassium channel (ROMK) mRNA abundance in rat kidney medullary thick ascending limb (MTAL) and in cultured mouse TAL cells. Using the run-off technique, we observed that the ROMK gene transcription rate in nuclei isolated from MTAL fragments was enhanced approximately 40% by a high NaCl medium. In MTAL fragments, hypertonicity (450 mosm) caused by NaCl, not by mannitol or urea, enhanced both ROMK mRNA abundance and tonicity-responsive enhancer binding protein (TonEBP) total abundance and nuclear localization. In an immortalized mouse TAL cell culture in which ROMK is apically expressed, hypertonicity caused by both NaCl and mannitol, not urea, enhanced both ROMK mRNA abundance and TonEBP total abundance and nuclear localization. Confocal microscopy confirmed an increased nuclear translocation of TonEBP in response to NaCl-induced hypertonicity. Finally, inhibition of the p38 MAPK pathway by SB203580 and of the ERK pathway by PD98059 abolished the NaCl-induced stimulation of TonEBP and ROMK. These results establish that mRNA expression of ROMK is augmented in the MTAL by NaCl-induced hypertonicity through stimulation of ROMK gene transcription, and that TonEBP and the p38 MAPK and ERK pathways are involved in this effect.
本研究评估了由氯化钠引起的高渗性增强大鼠肾髓质厚升支（MTAL）和培养的小鼠TAL细胞中肾外髓质钾通道（ROMK）mRNA丰度的机制。使用径流技术，我们观察到，高氯化钠培养基使从MTAL片段分离的细胞核中的ROMK基因转录率提高了约40%。在MTAL片段中，由氯化钠而非甘露醇或尿素引起的高渗性（450 mosm）增强了ROMK mRNA丰度以及渗透压反应增强子结合蛋白（TonEBP）的总丰度和核定位。在顶端表达ROMK的永生化小鼠TAL细胞培养物中，由氯化钠和甘露醇而非尿素引起的高渗性增强了ROMK mRNA丰度以及TonEBP的总丰度和核定位。共聚焦显微镜证实，响应氯化钠诱导的高渗性，TonEBP的核转位增加。最后，SB203580对p38 MAPK途径的抑制以及PD98059对ERK途径的抑制消除了氯化钠诱导的对TonEBP和ROMK的刺激。这些结果表明，氯化钠诱导的高渗性通过刺激ROMK基因转录增强了MTAL中ROMK的mRNA表达，并且TonEBP以及p38 MAPK和ERK途径参与了这一效应。
