Profiling of functional phosphodiesterase in mesangial cells using a CRE-SEAP-based reporting system
Profiling of functional phosphodiesterase in mesangial cells using a CRE-SEAP-based reporting system
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DOI：
10.1038/sj.bjp.0706785
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发表时间：
2006-07-01
影响因子：
7.3
通讯作者：
Kitamura, Masanori
中科院分区：
文献类型：
--
作者：
Zhu, Ying;Yao, Jian;Kitamura, Masanori
1磷酸二酯酶（PDE）在肾小球系膜细胞功能调节中起重要作用，但其在系膜细胞中的功能分布尚不清楚。在这项研究中，我们研究了单个PDE通过cAMP途径调节肾小球系膜细胞行为的作用。2在cAMP反应元件（CRE）的控制下表达分泌型碱性磷酸酶（SEAP）的报告系膜细胞在cAMP存在或不存在的情况下暴露于选择性PDE抑制剂，CRE的活性，CRE调节蛋白的表达，检测有丝分裂和细胞存活。3将报告细胞暴露于cAMP升高剂导致CRE的时间和浓度依赖性活化。单独用任何PDE抑制剂处理细胞不会诱导CRE活化。在8-溴-cAMP或8-溴-cGMP刺激下，PDE 2、PDE 3、PDE 4和PDE 5的抑制剂增强CRE的激活。PDE 1或PDE 6的抑制不影响CRE的激活。4在测试的不同组合中，仅PDE 3和PDE 4的抑制剂协同增加细胞内cAMP的水平、蛋白激酶A的活性、CRE的激活和CRE调节的蛋白、连接蛋白43。5同时抑制PDE 3和PDE 4减弱有丝分裂原诱导的细胞外信号调节激酶的激活和细胞增殖。在血清剥夺条件下，联合抑制PDE 3和PDE 4仅引起caspase-3的激活和细胞凋亡。6目前的数据阐明，PDE 3和PDE 4在系膜细胞功能的调节中发挥关键作用。PDE 3和PDE 4被鉴定为支持系膜细胞存活的新型抗凋亡机制。
1 Phosphodiesterases (PDEs) are critically implicated in the regulation of mesangial cell function, but profile of functional PDEs in mesangial cells is still unclear. In this study, we investigated roles of individual PDEs in the regulation of mesangial cell behavior by the cAMP pathway.2 Reporter mesangial cells that express secreted alkaline phosphatase (SEAP) under the control of the cAMP response element (CRE) were exposed to selective PDE inhibitors in the presence or absence of cAMP, and activity of CRE, expression of CRE-regulated protein, mitogenesis and cell survival were examined.3 Exposure of reporter cells to cAMP-elevating agents resulted in time- and concentration-dependent activation of CRE. Treatment of the cells with any PDE inhibitors alone did not induce CRE activation. Under stimulation with 8-bromo-cAMP or 8-bromo-cGMP, however, inhibitors of PDE2, PDE3, PDE4 and PDE5 enhanced activation of CRE. Inhibition of PDE1 or PDE6 did not affect the CRE activation.4 Among different combinations tested, only inhibitors of PDE3 and PDE4 cooperatively increased the level of intracellular cAMP, activity of protein kinase A, activation of CRE, and CRE-regulated protein, connexin43.5 Concomitant inhibition of PDE3 and PDE4 attenuated mitogen-induced activation of extracellular signal-regulated kinases and cell proliferation. Under serum deprivation, combinational inhibition of PDE3 and PDE4 exclusively caused activation of caspase-3 and apoptosis.6 The present data elucidated that PDE3 and PDE4 play critical roles in the regulation of mesangial cell function. PDE3 and PDE4 were identified as the novel, antiapoptotic machinery that supports survival of mesangial cells.