磷酸二酯酶4抑制剂对人巨噬细胞噬菌能力的影响

摘要

Objective To investigate the influence of phosphodiesterases 4 inhibitor on the phagocytosis of non-biologi-cal particles and gram-positive bacteria,gram-negative bacteria by human alveolar macrophages. Methods Peripheral blood mononuclear cells (PBMCs)were isolated from venous blood from 12 healthy volunteers using Ficoll-Hypaque density gradients.Monocytes were incubated with media containing 2 ng/ml GM-CSF for 12d to allow full differentiation into macrophage (MDM), a functionally equivalent model of human AM.MDMwere pretreated with Rolipram overnight (16-18h),phagocytosis of fluores-cent labeled beads and H.influenzae,S.aureus by MDMwas measured using a fluostar optima fluorimeter.Cell viability was as-say with MTT. Results MDMphagocytosis of beads and bacteria was a time-dependant process.Rolipram in the concentration of 10-8-10-5Mdidn't inhibit or promote phagocytosis of beads and bacteria by MDM,and didn't affect the cell viability. Conclu-sion Phosphodiesterases 4 inhibitor would not affect the human macrophage phagocytic capacity of non-biological particles and bacteria associated with enhanced intracellular cAMP level.%目的：探讨磷酸二酯酶4抑制剂对人肺泡巨噬细胞（AM）吞噬非生物性颗粒及革兰阳性菌、阴性菌能力的影响。方法使用 Ficolll-Hypaque 密度梯度法将外周血单核细胞分离的静脉血，在含有2 ng／m GM-CSF 的培养液中经12天诱导培养成 AM替代细胞模型—单核细胞源性巨噬细胞（MDM）。用酶标仪检测 MDM经磷酸二酯酶4抑制剂 Rolipram 预处理过夜（16～18 h）后吞噬荧光标记的非生物颗粒 Beads 和热灭活的流感嗜血杆菌（H.influenzae）、金黄色葡萄球菌（S.aureus）量的改变，另使用 MTT 法检测细胞活性。结果成功建立的 MDM细胞模型对 Beads 和细菌的吞噬呈时效关。Rolipram 在实验浓度（10～8～10-5 M）下对 MDM吞噬 Beads、H.influenzae 和S.aureus 能力无明显促进或抑制作用，也不影响 MDM的活性。结论磷酸二酯酶4抑制剂不会因升高巨噬细胞内cAMP 水平而影响其吞噬非生物颗粒和细菌的能力。