丙酮酸乙酯对缺氧／复氧损伤小鼠腹腔巨噬细胞一氧化氮生成及白细胞介素-1和6表达的影响

目的：探讨丙酮酸乙酯（EP）对缺氧／复氧损伤小鼠腹腔巨噬细胞产生一氧化氮（N0）及白细胞介素-1（IL-1）、IL-6表达的影响。方法：采用原代培养小鼠腹腔巨噬细胞和缺氧／复氧诱导损伤模型，墨汁吞噬实验鉴定巨噬细胞纯度，于缺氧3h后复氧1、3、6、9h时收集上清液，硫代巴比妥酸法检测丙二醛（MDA）活性，采用噻唑兰法测定其存活率。在缺氧3h／复氧6h（H3／R6）损伤实验中，细胞分为正常对照组、损伤组和加入不同浓度EP（20、2、0．2mmol／L）后的H3／R6损伤组，取各组细胞上清，硝酸还原酶法测定其NO生成量，放射免疫分析法检测细胞上清中IL-1、IL-6的表达。结果：细胞培养5d后，墨汁吞噬实验鉴定细胞吞噬率达99％以上。与正常对照组相同时间点相比，缺氧3h后复氧1、3、6和9h时，细胞存活率明显下降，上清中MDA含量明显增加（P〈0．05），但复氧6h组与复氧9h组间差异均无显著性。与正常对照组相比，H3／H6损伤组NO生成量降低，IL-1、IL-6含量明显升高（P〈0．05）；不同浓度EP处理后结果显示，0．2和2mmol／LEP可明显促进N0的生成，降低IL-1和IL-6的含量（P〈0．05）；而20mmol／LEP处理可降低NO生成，提高IL-1和IL-6的表达。结论：低浓度EP对小鼠腹腔巨噬细胞缺氧／复氧损伤有保护作用，其途径可能是通过上调NO含量，下调IL-1和IL-6表达水平实现的。

Effects of ethyl pyruvate on the production of nitic oxide and expression of interleukin-1 and interleukin-6 in peritoneal macrophages after hypoxia-reoxygenation in mice

Objective：To study the effects of ethyl pyruvate （EP） on the production of nitric oxide （NO） and expression of interleukin-1 （IL-1） and IL-6 in peritoneal macrophages after hypoxia-reoxygenation injury in mice. Methods：Peritoneal macrophages of mice were harvested and cultured, and they were used to reproduce the hypoxia/reoxygenation injury model. The cells were identified by phagocytosis activity. At 1, 3,6 and 9 hours of reoxy- genation after 3-hour hypoxia, the activity of malondialdehyde （MDA） and survival rate of peritoneal macrophages were determined with TBA and MTT methods. In the experiment of 3-hour hypoxia/6-hour reoxygenation （H3/R6）, the macrophages were randomly divided into normal control group, H3/R6 group, H3/R6 4-20 mmol/L, 2 mmol/L or 0. 2 mmol/L of EP groups. The NO levels were detected by nitric acid reductase assay, and the IL-1 and IL-6 expression were analyzed by radioimmunoassay. Results：After a 5-day culture, the phagocytosis rate of the cells reached 99%. At 3- hour hypoxia and 1,3,6,9-hour reoxygernation, the survival rate of the macrophages was reduced significantly and MDA level increased significantly compared to those of the normal macrophages （P〈0. 05）, but the difference was not significant between 6- and 9-hour reoxygenation cells （P〉0. 05）. After H3/R6 injury, the NO content decreased and the expression of IL-1 and IL-6 increased compared to the normal control group （P〈0.05）. It was showed that 0. 2 and 2 mmol/L EP could improve NO level and decrease IL-1, IL-6 contents, but the effect of 20 mmol/L EP was vice versa. Conclusion： Low concentration of EP has protective effect on hypoxia/reoxygenation injury in mice. The modulation in NO and IL-1 ,IL-6 production in macrophage by EP may play an important role in immunoregulation.