Gantner F, Tenor H, Gekeler V, Schudt C, Wendel A, Hatzelmann A
Byk Gulden, Biochemistry Department, Konstanz, Germany.
J Allergy Clin Immunol. 1997 Oct;100(4):527-35. doi: 10.1016/s0091-6749(97)70146-5.
Several previous reports have suggested an increased activity of cAMP phosphodiesterases (PDEs) and a higher sensitivity of these enzymes toward PDE inhibitors in leukocytes of patients suffering from atopic dermatitis.
The purpose of the present study was to comprehensively analyze and compare the PDE expression and activity profile of highly purified populations of leukocytes from normal and atopic blood donors. In addition, the influence of PDE inhibitors on function of leukocytes from normal and atopic individuals was investigated.
Density gradient centrifugation, elutriation, and magnetic cell sorting techniques were used to purify eosinophils, monocytes, and B and T lymphocytes from peripheral human blood. Complementary DNA-polymerase chain reaction was used to analyze PDE4 subtype messenger RNA (mRNA) expression levels in addition to PDE isoenzyme activities. PDE4 inhibitor sensitivity was determined in monocyte homogenates from both groups. Functionally, suppression of lipopolysaccharide-induced synthesis of tumor necrosis factor-alpha in monocytes as well as phytohemagglutinin-induced T cell proliferation in peripheral blood mononuclear cell fractions by PDE4 and PDE3/4 inhibitors was compared.
Identical PDE activities and mRNA expression profiles were found in all cells from normal and atopic donors except that there was an increase in the mRNA levels of PDE4A and PDE4B2 in atopic T cells, which was, however, not reflected in overall PDE4A activity. In addition, no differences in sensitivity of the functional responses to PDE inhibitors were noted. The mixed PDE3/4 inhibitor zardaverine was a more potent inhibitor of T cell proliferation than rolipram, a selective PDE4 inhibitor.
No evidence for alterations of PDE activities in atopy is provided by our findings.
先前的几份报告表明，特应性皮炎患者白细胞中cAMP磷酸二酯酶（PDEs）的活性增加，且这些酶对PDE抑制剂的敏感性更高。
本研究的目的是全面分析和比较来自正常和特应性献血者的高度纯化白细胞群体的PDE表达和活性谱。此外，还研究了PDE抑制剂对正常和特应性个体白细胞功能的影响。
采用密度梯度离心、淘洗和磁性细胞分选技术从人外周血中纯化嗜酸性粒细胞、单核细胞以及B和T淋巴细胞。除了PDE同工酶活性外，还使用互补DNA聚合酶链反应分析PDE4亚型信使核糖核酸（mRNA）表达水平。在两组的单核细胞匀浆中测定PDE4抑制剂敏感性。在功能上，比较了PDE4和PDE3/4抑制剂对单核细胞中脂多糖诱导的肿瘤坏死因子-α合成的抑制作用以及对植物血凝素诱导的外周血单个核细胞组分中T细胞增殖的抑制作用。
在正常和特应性献血者的所有细胞中发现了相同的PDE活性和mRNA表达谱，只是特应性T细胞中PDE4A和PDE4B2的mRNA水平有所增加，但这并未反映在总体PDE4A活性中。此外，未观察到功能反应对PDE抑制剂敏感性的差异。混合PDE3/4抑制剂扎达维林比选择性PDE4抑制剂咯利普兰更有效地抑制T细胞增殖。
我们的研究结果未提供特应性中PDE活性改变的证据。
