Agonist-specific regulation of inositol phosphate metabolism in cardiac endothelial cells.

The actions of nucleotides and hormones at endothelial cell (EC) receptors are known to result in the release of ATP that acts as a local hormone to facilitate release of mediators such as NO and PGI2. Stimulation of ECs with the P2Y1 receptor agonist 2-MeSATP leads to the rapid release of Ca2+ from stores consistent with a role for inositol trisphosphate (Ins-1,4,5-P3) in mediating the action of extracellular nucleotides. Guinea pig ECs were grown in primary culture. [3H]d-myo-inositol (30 Ci/mmol) labeling studies revealed maximal incorporation of radioactivity into [3H]Ins-1,4,5-P3 when glucose in the labeling buffer was lowered to 1 mM and non-radioactive inositol was added at 10 microM. Stimulation of EC for one sec led to the dose-dependent accumulation of [3H]Ins-1,4,5-P3 as well as [3H]IP4, [3H]IP5, and [3H]IP6. Unexpectedly, the metabolism of [3H]Ins-1,4,5-P3 to IP1 was disparat in stimulated versus un-stimulated cells. In [3H]d-Ins labeled stimulated EC or in homogenates derived from unlabeled, stimulated EC, dephosphorylation of [3H]Ins-1,4,5-P3 led to the exclusive formation of [3H]Ins-4-P1. Addition of on-nucleotide agonists such as bradykinin gave the same results suggesting that the dephosphorylation pathway for IP formation in EC is dependent on agonist stimulation and may be correlated with regulation of agonist responsiveness or heretofore unrecognized actions of IP isomers in stimulated versus unstimulated cells.

已知核苷酸和激素在内皮细胞（EC）受体上的作用会导致ATP释放，ATP作为一种局部激素促进一氧化氮（NO）和前列环素（PGI2）等介质的释放。用P2Y1受体激动剂2 - 甲硫基腺苷三磷酸（2 - MeSATP）刺激内皮细胞会导致钙离子（Ca2 +）从储存库中迅速释放，这与肌醇三磷酸（Ins - 1,4,5 - P3）在介导细胞外核苷酸作用中的角色相符。豚鼠内皮细胞在原代培养中生长。[3H] - D - 肌醇（30居里/毫摩尔）标记研究显示，当标记缓冲液中的葡萄糖降低到1毫摩尔且加入10微摩尔非放射性肌醇时，放射性最大程度地掺入到[3H]Ins - 1,4,5 - P3中。刺激内皮细胞1秒会导致[3H]Ins - 1,4,5 - P3以及[3H]IP4、[3H]IP5和[3H]IP6呈剂量依赖性积累。出乎意料的是，[3H]Ins - 1,4,5 - P3代谢为IP1在受刺激细胞和未受刺激细胞中是不同的。在[3H]D - Ins标记的受刺激内皮细胞中，或者在来自未标记的受刺激内皮细胞的匀浆中，[3H]Ins - 1,4,5 - P3的去磷酸化仅形成[3H]Ins - 4 - P1。加入非核苷酸激动剂如缓激肽得到相同结果，这表明内皮细胞中IP形成的去磷酸化途径依赖于激动剂刺激，并且可能与激动剂反应性的调节或受刺激细胞与未受刺激细胞中IP异构体迄今未被认识的作用相关。