Leukotriene B4 production in human mononuclear phagocytes is modulated by interleukin-4-induced 15-lipoxygenase
Leukotriene B4 production in human mononuclear phagocytes is modulated by interleukin-4-induced 15-lipoxygenase
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DOI：
10.1124/jpet.300.3.868
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发表时间：
2002-03-01
影响因子：
3.5
通讯作者：
Vignola, AM
中科院分区：
文献类型：
--
作者：
Profita, M;Sala, A;Vignola, AM
本研究的目的是评估白细胞介素(IL)-4诱导的15-脂氧合酶（15-LO）的后果。白三烯B-4 （LTB4）在人单核细胞合成中的表达。人单核细胞与IL-4 （10 ng/ml）孵育24、48和72小时后，用Ca2+-离子载体A23187（钙霉素，5 muM）或调理酶san刺激。采用高效液相色谱/放射免疫法、液相色谱/串联质谱法（LC/MS/MS）或气相色谱/质谱法测定15(S)-羟基二碳四烯酸[15(S)-HETE]、LTB4和花生四烯酸（AA）的释放量。在aa处理的单核细胞中评估15-LO活性。用逆转录-聚合酶链反应分析15-LO、5-脂氧合酶（5-LO）和5-LO激活蛋白（FLAP）的表达。中性粒细胞趋化活性评价采用微致性室试验。IL-4 （10 ng/ml）作用48和72 h后，a23187诱导的15(S)-HETE的合成显著增加（p < 0.001）。同时观察到LTB4释放减少。与IL-4孵育72 h后（p < 0.001）。LC/MS/MS分析证实，il -4处理的单核细胞在调理酶san刺激后产生15(S)-HETE，并显著抑制LTB4的合成。IL-4处理诱导15-LO酶活性和15-LO mRNA，但不影响5-LO和FLAP mRNA在单核细胞中的表达。il -4处理单核细胞的上清液显示中性粒细胞趋化活性明显低于对照组。15(S)-HETE显著抑制a23187刺激的人单核细胞诱导的LTB4产生，而不影响AA的释放。il -4诱导的15-LO在单核细胞中的表达导致LTB4的产生显著减少。然而，这种影响并不反映5-LO和FLAP mRNA表达的变化，合成15(S)-HETE能够显著抑制LTB4的合成，而不影响AA的释放。
The aim of this study was to evaluate, the consequences of interleukin (IL)-4-induced 15-lipoxygenase (15-LO). expression on leukotriene B-4 (LTB4) synthesis in human monocytes. Human monocytes incubated for 24, 48, and 72 h with IL-4 (10 ng/ml) were stimulated with Ca2+-ionophore A23187 (calcimycin, 5 muM) or opsonized zymosan. 15(S)-hydroxyeicosatetraenoic acid [15(S)-HETE], LTB4, and arachidonic acid (AA) release were measured by high-performance liquid chromotography/radioimmunoassay, liquid chromotography/tandem mass spectrometry (LC/MS/MS), or gas chromatography/mass spectrometry. 15-LO activity was evaluated in AA-treated monocytes. 15-LO, 5-lipoxygenase (5-LO) and 5-LO activating protein (FLAP) expression were analyzed by reverse transcription-polymerase chain reaction. Neutrophil chemotactic activity was evaluated using a microtaxis chamber assay. A23187-induced synthesis of 15(S)-HETE was significantly increased after treatment with IL-4 (10 ng/ml) for 48 and 72 h (p < 0.001). Concomitant decrease of LTB4 release was observed. after 72 h of incubation with IL-4 (p < 0.001). LC/MS/MS analysis confirmed the production of 15(S)-HETE and the significant inhibition of LTB4 synthesis in IL-4-treated monocyte after challenge with opsonized zymosan. IL-4 treatment induced 15-LO enzymatic activity as well as 15-LO mRNA, but did not affect either 5-LO or FLAP mRNA expression in monocytes. Supernatant from IL-4-treated monocytes showed significantly lower neutrophil chemotactic activity than controls. 15(S)-HETE significantly inhibited LTB4 production induced by A23187-stimulated human monocytes without affecting AA release. IL-4-induced expression of 15-LO in monocytes caused a significant reduction of LTB4 production. Whereas this effect did not reflect changes in 5-LO and FLAP mRNA expression, synthetic 15(S)-HETE was able to significantly inhibit the synthesis of LTB4, without affecting AA release.