内毒素耐受对人中性粒细胞抗炎反应的影响

Effects of endotoxin tolerance on anti-inflammatory responses in human neutrophils

目的:探讨脂多糖(lipopolysaccharide,LPS)诱导的耐受对人中性粒细胞分泌抗炎因子白介素(interleukin,IL)-10及细胞凋亡水平的影响。方法:采用1μg/m L牙龈卟啉单胞菌(Porphyromonasgingivalis,P.gingivalis)LPS或者1μg/m L大肠杆菌(Escherichia coli,E.coli)LPS刺激人中性粒细胞12 h,诱导细胞耐受。洗脱后,分别采用2种LPS再次刺激20 h,采用ELISA技术检测细胞分泌IL-10水平。同法诱导细胞耐受,洗脱后,再次刺激3 h,采用AV-PI法检测细胞凋亡水平。结果:P.gingivalis LPS或E.coli LPS单次刺激人中性粒细胞后,IL-10分泌水平均较刺激前明显增加(P<0.05),凋亡水平则无明显改变(P>0.05)。2种LPS分别重复刺激之后,IL-10分泌水平均较单次刺激后显著增加(P<0.05)。P.gingivalis LPS重复刺激3 h后,中性粒细胞短期内凋亡水平与单次刺激后无明显改变(P>0.05),而E.coli LPS重复刺激3 h后,细胞短期内凋亡水平较单次刺激后明显下降(P<0.05)。结论:内毒素耐受能促进人中性粒细胞分泌IL-10,但不同LPS诱导的耐受对细胞凋亡水平的影响可能存在差异。

Objective： To observe the effects of endotoxin tolerance induced by lipopolysaccharide （LPS） on the production of antiinflammatory cytokine IL-10 and the level of apoptosis in human neutrophils. Methods： Neutrophils were pretreated with 1 μg/mL Porphyromonasgingivalis （P.gingivalis） LPS or 1 μg/mL Escherichia coli （E.coli） LPS for 12 h to induce cell tolerance. Then, the cells were washed and stimulated with the same LPS for additional 20 h （for ELISA） or 3 h （for flow cytometry）. Levels of IL-10 in supernatants and apoptosis of neutrophils were detected by ELISA and flow cytometry, respectively. Results： After 20 h, the amount of IL-10 secreted by neutrophils treated with P.gingivalis LPS or E.coli LPS was increased significantly compared with that without any stimulation （P〈0.05）, while no significant changes of apoptosis in neutrophils with LPS stimulation were observed after 3 h （P〉 0.05）. After restimulation with P.gingivalis LPS or E.coli LPS, IL-10 production was increased significantly compared with that secreted by neutrophils challenged with the same LPS only once （P〈0.05）. However, no significant changes of apoptosis in the short term were detected after P.gingivalis LPS retreatment for 3 h （P〉0.05）, and apoptosis levels in the short term were significantly decreased after E.coli LPS restimulations for 3 h （P〈0.05）. Conclusion： Endotoxin tolerance might contribute to the increased production of IL-10 in human neutrophils, but the effects of tolerance on apoptosis might depend on the types of LPS.