Phosphodiesterase-4 inhibition improves corticosteroid insensitivity in pulmonary endothelial cells under oxidative stress
Phosphodiesterase-4 inhibition improves corticosteroid insensitivity in pulmonary endothelial cells under oxidative stress
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DOI：
10.1111/all.12055
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发表时间：
2013-01-01
期刊：
影响因子：
12.4
通讯作者：
Cortijo, J.
中科院分区：
文献类型：
--
作者：
Ortiz, J. L.;Milara, J.;Cortijo, J.
研究背景：多项临床研究表明，哮喘和慢性阻塞性肺疾病患者的吸烟与皮质类固醇治疗无效密切相关。在这项工作中，我们分析了糖皮质激素不敏感性的人肺动脉内皮细胞（HPAECs）下香烟烟雾提取物（CSE）暴露以及可能的组合治疗与磷酸二酯酶（PDE）-4 inhibitors.Methods白细胞介素（IL）-8测定细胞上清液中的ELISA。组蛋白去乙酰化酶（HDAC），组蛋白乙酰化酶（HAT）和细胞内cAMP水平分别测定比色法和酶免疫法。结果PDE 4抑制剂咯利普兰剂量依赖性地抑制CSE诱导的IL-8分泌。地塞米松1 μ M对IL-8分泌无抑制作用。10 nM的亚有效咯利普兰浓度的组合将地塞米松的抑制作用增加至类似于45%的抑制。香烟烟雾提取物5%抑制HDAC活性并增加HAT活性，产生糖皮质激素不敏感性。Rolipram不改变HDAC活性，但在1 μ M时部分抑制HAT活性的增加。PDE 4同种型被CSE 5%上调，随后cAMP下调。地塞米松降低所有PDE 4同种型的表达，并显示与咯利普兰增加cAMP水平的累加效应。此外，咯利普兰增强GR-α的表达和抑制GR-β诱导CSE.Conclusions在糖皮质激素不敏感的条件下，咯利普兰和地塞米松的组合显示HPAECs的相加特性。这些结果可能对未来使用PDE 4抑制剂和糖皮质激素组合的抗炎治疗具有潜在价值。
Background Several clinical studies have shown that smoking in asthmatics and chronic obstructive pulmonary disease patients is closely associated with corticosteroid refractoriness. In this work, we have analyzed glucocorticoid insensitivity in human pulmonary artery endothelial cells (HPAECs) under cigarette smoke extract (CSE) exposure as well as the possible additive effects of the combination therapy with a phosphodiesterase (PDE)-4 inhibitor.Methods Interleukin (IL)-8 was measured in cell supernatants by ELISA. Histone deacetylase (HDAC), histone acetylase (HAT), and intracellular cAMP levels were measured by colorimetric assays and enzyme immunoassay, respectively. PDE4 isotypes and glucocorticoid receptor (GR)-alpha and beta expression were measured by real-time RT-PCR.Results The PDE4 inhibitor rolipram dose dependently inhibited the IL-8 secretion induced by CSE 5%. In contrast, dexamethasone 1 mu M did not show inhibitory effect on IL-8 secretion. Combination of subeffective rolipram concentrations at 10 nM increased the inhibitory effect of dexamethasone to similar to 45% of inhibition. Cigarette smoke extract 5% inhibited HDAC activity and increased HAT activity generating glucocorticoid insensitivity. Rolipram did not modify the HDAC activity, however partially inhibited the increase in HAT activity at 1 mu M. PDE4 isotypes were up-regulated by CSE 5% with the consequent cAMP down-regulation. Dexamethasone reduced all PDE4 isotypes expression and showed additive effects with rolipram enhancing cAMP levels. Furthermore, rolipram enhanced GR-alpha expression and inhibited the increase in GR-beta induced by CSE.Conclusions Combination of rolipram and dexamethasone shows additive properties in HPAECs under glucocorticoid insensitive conditions. These results may be of potential value in future anti-inflammatory therapies using combination of PDE4 inhibitors and glucocorticoids.