Altered expression of cyclic nucleotide phosphodiesterase isozymes during culture of aortic endothelial cells
Altered expression of cyclic nucleotide phosphodiesterase isozymes during culture of aortic endothelial cells
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DOI：
10.1016/s0006-2952(97)00287-6
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发表时间：
1997-11-15
影响因子：
5.8
通讯作者：
Thompson, WJ
中科院分区：
文献类型：
--
作者：
Ashikaga, T;Strada, SJ;Thompson, WJ
牛主动脉内皮细胞（BAEC）的原代培养物表达PDE 2、PDE 4和PDE基因家族的环核苷酸磷酸二酯酶（CN PDE）同工酶。我们在这里报告的CN PDE的同工酶谱和每一个的量随BAEC培养物的传代次数而变化。通过阴离子交换色谱法和药理学标准进行表征，用于研究纯化BAEC早期（4-6）、中期（6-10）和晚期（>17）代的CN PDE。PDE 2和一小部分的PDES占环GMP水解在早期通道，但这两种同工酶都失去了细胞通道。在早期传代的内皮细胞中，PDE 2和PDE 4同工酶都能水解cAMP，但在高传代的内皮细胞中，PDE 4显著增加。在高传代细胞中还出现显著的PDE 1和较小的PDE 3活性。在所有传代中，细胞溶质与颗粒活性的比值相似。通过[H-3]-腺嘌呤预标记评估的完整细胞中的BAEC PDE亚型表明，心房肽素II降低了异丙肾上腺素诱导的环AMP在早期传代细胞中的蓄积，但不降低晚期传代细胞中的蓄积，这与PDE 2表达的丧失一致。rolipram，PDE 4抑制剂，异丙肾上腺素诱导的环AMP积累的增强，也大大减少在培养传代。CN PDE亚型表达和随之而来的环AMP营业额的变化验证了考虑细胞传代次数的重要性时，BAEC的培养物被用来研究在这个模型系统中的环核苷酸介导的内皮细胞环核苷酸代谢和过程的调节。(C)1997年爱思唯尔科学公司
Primary cultures of bovine aortic endothelial cells (BAEC) express cyclic nucleotide phosphodiesterase (CN PDE) isozymes of the PDE2, PDE4 and PDES gene families. We report here that the isozyme profiles of CN PDE and the amounts of each vary with the passage number of BAEC cultures. Characterization by anion-exchange chromatography and pharmacological criteria were used to study CN PDE in early (4-6), intermediate (6-10), and late (>17) passages of purified BAEC. PDE2 and a minor fraction of PDES accounted for cyclic GMP hydrolysis in early passages, but both isozymes were lost with cell passage. Cyclic AMP was hydrolyzed by both PDE2 and PDE4 isozymes in early passage endothelial cells, but PDE4 was increased dramatically in higher passage cells. Also appearing in the higher passage cells were prominent PDE1 and minor PDE3 activities. The ratios of cytosolic to particulate activities were similar at all passages. BAEC PDE isoforms in intact cells assessed by [H-3]-adenine prelabeling showed that atriopeptin II decreased isoproterenol-induced cyclic AMP accumulation in early but not later passage cells, consistent with the loss of PDE2 expression. Enhancement of isoproterenol-induced cyclic AMP accumulation by rolipram, a PDE4 inhibitor, was also greatly diminished during culture passages. Changes in CN PDE isoform expression and consequent cyclic AMP turnover validate the importance of considering cell passage number when cultures of BAEC are used to study the regulation of endothelial cell cyclic nucleotide metabolism and processes mediated by cyclic nucleotides in this model system. (C) 1997 Elsevier Science Inc.