Gallazzini Morgan, Ferraris Joan D, Kunin Margarita, Morris Ryan G, Burg Maurice B
Laboratory of Kidney and Electrolyte Metabolism, National Heart Lung and Blood Institute, Department of Health and Human Services, Bethesda, MD 20892-1603, USA.
Proc Natl Acad Sci U S A. 2006 Oct 10;103(41):15260-5. doi: 10.1073/pnas.0607133103. Epub 2006 Oct 2.
Glycerophosphocholine (GPC) is an osmoprotective compatible and counteracting organic osmolyte that accumulates in renal inner medullary cells in response to high NaCl and urea. We previously found that high NaCl increases GPC in renal [Madin-Darby canine kidney (MDCK)] cells. The GPC is derived from phosphatidylcholine, catalyzed by a phospholipase that was not identified at that time. Neuropathy target esterase (NTE) was recently shown to be a phospholipase B that catalyzes production of GPC from phosphatidylcholine. The purpose of the present study was to test whether NTE contributes to the high NaCl-induced increase of GPC synthesis in renal cells. We find that in mouse inner medullary collecting duct cells, high NaCl increases NTE mRNA within 8 h and NTE protein within 16 h. Diisopropyl fluorophosphate, which inhibits NTE esterase activity, reduces GPC accumulation, as does an siRNA that specifically reduces NTE protein abundance. The 20-h half-life of NTE mRNA is unaffected by high NaCl. TonEBP/OREBP is a transcription factor that is activated by high NaCl. Knockdown of TonEBP/OREBP by a specific siRNA inhibits the high NaCl-induced increase of NTE mRNA. Further, the lower renal inner medullary interstitial NaCl concentration that occurs chronically in ClCK1-/- mice and acutely in normal mice given furosemide is associated with lower NTE mRNA and protein. We conclude that high NaCl increases transcription of NTE, likely mediated by TonEBP/OREBP, and that the resultant increase of NTE expression contributes to increased production and accumulation of GPC in mammalian renal cells in tissue culture and in vivo.
甘油磷酸胆碱（GPC）是一种具有渗透保护作用的相容性且能起抵消作用的有机渗透溶质，它在高氯化钠和尿素作用下会在肾髓质内层细胞中蓄积。我们之前发现高氯化钠会使肾[麦迪逊-达比犬肾（MDCK）]细胞中的GPC增加。GPC由磷脂酰胆碱衍生而来，由当时尚未明确的一种磷脂酶催化产生。最近研究表明，神经病变靶酯酶（NTE）是一种磷脂酶B，可催化从磷脂酰胆碱生成GPC。本研究的目的是测试NTE是否参与高氯化钠诱导的肾细胞中GPC合成增加。我们发现，在小鼠髓质内层集合管细胞中，高氯化钠在8小时内会使NTE mRNA增加，在16小时内会使NTE蛋白增加。抑制NTE酯酶活性的二异丙基氟磷酸酯会减少GPC的蓄积，特异性降低NTE蛋白丰度的小干扰RNA（siRNA）也会如此。NTE mRNA的20小时半衰期不受高氯化钠影响。TonEBP/OREBP是一种受高氯化钠激活的转录因子。用特异性siRNA敲低TonEBP/OREBP会抑制高氯化钠诱导的NTE mRNA增加。此外，ClCK1-/-小鼠长期以及正常小鼠急性给予呋塞米后肾髓质内层间质氯化钠浓度降低，这与较低的NTE mRNA和蛋白相关。我们得出结论，高氯化钠会增加NTE的转录，可能由TonEBP/OREBP介导，并且由此导致的NTE表达增加有助于在组织培养和体内的哺乳动物肾细胞中GPC的生成和蓄积增加。
