Friedlaender M M, Jain D, Ahmed Z, Hart D, Barnett R L, Nord E P
Department of Medicine, School of Medicine, Hadassah University Hospital, Jerusalem, Israel.
Am J Physiol. 1993 May;264(5 Pt 2):F845-53. doi: 10.1152/ajprenal.1993.264.5.F845.
Previous work from this laboratory has identified an endothelin (ET) type A (ETA) receptor on cultured rat renal medullary interstitial cells (RMIC), coupled to phosphatidylinositol-specific phospholipase C (PI-PLC), dihydropyridine-insensitive receptor-operated Ca2+ channels, and phospholipase A2. The current studies explored a role for ET stimulation of phosphatidylcholine-specific phospholipase D (PC-PLD) in intracellular signaling of this cell type. ET stimulated PLD activation, as measured by phosphatidic acid (PA) or phosphatidylethanol (PEt) accumulation, in a time- and concentration-dependent manner. Inhibition of diacylglycerol (DAG) kinase by ethylene glycol dioctanoate or 6-(2)4-[(4-fluorophenyl)-phenylmethylene]-1-piperadinyl]ethy l-7-methyl-5H - thiaxolo-[3,2-alpyrimidin]-5-one (R 59022) failed to blunt PA accumulation, indicating that PLD, and not DAG, was the source of PA. Inhibition of PA phosphohydrolase (PAP) by propranolol increased late accumulation of PA, suggesting that the prevailing metabolic flow was in the direction of PA to DAG. Phorbol 12-myristate 13-acetate (PMA) augmented ET-evoked PEt accumulation, whereas downregulation of protein kinase C (PKC) obviated agonist-induced PEt production. PMA augmentation of PLD activity proceeded independent of cytosolic free Ca2+ concentration. Ca2+ derived from either intracellular or extracellular sources enhanced ET-related PEt accumulation but was without effect in PKC-downregulated cells. Collectively, these observations indicate that ET stimulates PLD production in RMIC. PKC is the major regulator of this process, with Ca2+ playing a secondary, modulatory role. In addition, these data suggest that PC-PLD is coupled to the ETA receptor.
该实验室之前的研究已在培养的大鼠肾髓质间质细胞（RMIC）上鉴定出一种内皮素（ET）A型（ETA）受体，其与磷脂酰肌醇特异性磷脂酶C（PI-PLC）、二氢吡啶不敏感的受体操纵性Ca²⁺通道及磷脂酶A₂偶联。当前研究探讨了ET刺激磷脂酰胆碱特异性磷脂酶D（PC-PLD）在这种细胞类型的细胞内信号传导中的作用。ET以时间和浓度依赖性方式刺激PLD激活，这可通过磷脂酸（PA）或磷脂酰乙醇（PEt）积累来衡量。用乙二醇二辛酸酯或6-（2）4-[(4-氟苯基）-苯基亚甲基]-1-哌啶基]乙基-7-甲基-5H-噻唑并-[3,2-a]嘧啶-5-酮（R 59022）抑制二酰基甘油（DAG）激酶未能减弱PA积累，表明PA的来源是PLD而非DAG。普萘洛尔抑制PA磷酸水解酶（PAP）可增加PA的后期积累，提示主要的代谢流向是从PA到DAG。佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA）增强ET诱发的PEt积累，而蛋白激酶C（PKC）的下调消除了激动剂诱导的PEt产生。PMA增强PLD活性的过程独立于胞质游离Ca²⁺浓度。源自细胞内或细胞外的Ca²⁺增强了ET相关的PEt积累，但对PKC下调的细胞无作用。总体而言，这些观察结果表明ET刺激RMIC中PLD的产生。PKC是这一过程的主要调节因子，Ca²⁺起次要的调节作用。此外，这些数据表明PC-PLD与ETA受体偶联。
