Thrombin-promoted release of UDP-glucose from human astrocytoma cells
Thrombin-promoted release of UDP-glucose from human astrocytoma cells
复制标题
DOI：
10.1038/sj.bjp.0707692
复制
发表时间：
2008-04-01
影响因子：
7.3
通讯作者：
Lazarowski, E. R.
中科院分区：
文献类型：
--
作者：
Kreda, S. M.;Seminario-Vidal, L.;Lazarowski, E. R.
背景和目的：P2 Y（14）受体被UDP-糖激活，最有效地被UDP-葡萄糖激活，但不被游离核苷酸激活，表明UDP-葡萄糖是该受体的同源激动剂。然而，UDP-葡萄糖的调节释放的证据是稀缺的。本研究以1321 N1人星形细胞瘤细胞为研究对象，采用高效液相色谱法测定UDP-葡萄糖的释放和水解，并探讨UDP-葡萄糖的受体促进释放的机制。磷脂酶C激活和肌动蛋白细胞骨架重组进行了评估，通过测量磷酸肌醇形成和荧光共聚焦microscopics.Key结果：凝血酶和蛋白酶激活受体1（PAR 1）肽TFLLRNPNDK（PAR 1-AP）诱发释放UDP-葡萄糖和ATP，这是伴随着增强磷酸肌醇形成。虽然卡巴胆碱促进四倍以上的磷酸肌醇形成比凝血酶，它未能促进核苷酸的释放。凝血酶促进的核苷酸释放被BAPTA-AM、布雷菲德菌素A和细胞松弛素D抑制，并且对百日咳毒素和PI 3激酶抑制剂不敏感。凝血酶，而不是卡巴胆碱，诱导肌动蛋白细胞骨架重组，在1321 N1细胞中的Rho激活的标志。然而，PAR促进UDP-葡萄糖释放不受影响Rho激酶inhibition.Conclusions和影响：PAR 1诱发UDP-葡萄糖释放反映了钙离子依赖的机制，从事额外的信号独立的Gi和Rho激酶激活，需要一个功能性的肌动蛋白细胞骨架和高尔基体结构。我们的研究表明，发生的Ca-2+依赖性释放UDP-葡萄糖从星形细胞瘤细胞在生理相关的刺激，即G蛋白偶联受体激动剂。鉴于星形胶质细胞中存在P2 Y14受体，UDP-葡萄糖可能在脑中具有重要的自分泌/旁分泌功能。
Background and purpose: The P2Y(14) receptor is activated by UDP-sugars, most potently by UDP-glucose, but not by free nucleotides, suggesting that UDP-glucose is the cognate agonist for this receptor. However, evidence for regulated release of UDP-glucose is scarce. In the present study, the occurrence of receptor-promoted release of UDP-glucose was investigated, using 1321N1 human astrocytoma cells.Experimental approach: UDP-glucose release and hydrolysis were measured using HPLC-based techniques. Phospholipase C activation and actin cytoskeleton reorganization were assessed by measuring inositol phosphate formation and fluorescence confocal microscopy, respectively.Key results: Thrombin and the protease-activating receptor-1 (PAR1) peptide TFLLRNPNDK (PAR1-AP) evoked the release of UDP-glucose and ATP, which was accompanied by enhanced inositol phosphate formation. Although carbachol promoted fourfold greater inositol phosphate formation than thrombin, it failed to promote nucleotide release. Thrombin-promoted nucleotide release was inhibited by BAPTA-AM, brefeldin A and cytochalasin D, and was insensitive to Pertussis toxin and PI3- kinase inhibitors. Thrombin, but not carbachol, induced actin cytoskeleton reorganization, a hallmark of Rho activation in 1321N1 cells. However, PAR-promoted UDP-glucose release was not affected by Rho kinase inhibition.Conclusions and implications: PAR1-evoked UDP-glucose release reflected a Ca2+ dependent mechanism, engaging additional signalling independently of Gi and Rho kinase activation and requiring a functional actin cytoskeleton and Golgi structures. Our study demonstrates the occurrence of Ca-2+-dependent release of UDP-glucose from astrocytoma cells in response to a physiologically relevant stimulus, that is, a G-protein-coupled receptor agonist. Given the presence of P2Y14 receptors in astrocytes, UDP-glucose may have important autocrine/ paracrine functions in the brain.