Effects of hypoxia-reoxygenation on superoxide production and change of intra-cellular Ca^<++> of neutrophils

用细胞色素C还原法研究了缺氧-复氧对中性粒细胞生成超氧阴离子（O_2^-）的影响。从健康成人外周静脉血中分离出中性粒细胞。对照组采用空气PO_2 （220mmHg）治疗。缺氧组给予中性粒细胞N_2处理30 min。复氧组中性粒细胞在n2预处理30 min后再进行氧处理15 min。用细胞色素C还原法测定刺激和不刺激肉豆蔻酸佛酚后的超氧阴离子生成量。缺氧导致O_2^-产量显著下降。再氧化使未受刺激的中性粒细胞产生的O_2^-显著增加。这些结果表明，中性粒细胞被缺氧再氧化激活。第二项研究考察了缺氧-复氧对NADPH氧化酶活性的影响以及细胞内中性粒细胞的变化。对照组用空气（PO_2 220mmHg）处理中性粒细胞细胞膜。缺氧组用氮气（PO_2 28mmHg）处理细胞膜30 min。复氧组在氮气预处理30 min （PO_2 480mmHg）后，对细胞膜进行氧处理15 min。氧合组给予氧疗15 min （PO_2 480mmHg）。用细胞色素C还原法测定NADPH氧化酶活性。缺氧导致NADPH氧化酶活性显著降低。复氧引起NADPH氧化酶活性显著升高。这些结果表明，NADPH氧化酶被缺氧再氧化激活。细胞内钙离子随着再氧化而增加。这表明细胞内钙离子的增加激活了中性粒细胞中的NADPH氧化酶。

A study was made on the effects of hypoxia-reoxygenation on superoxide anion (O_2^-) production in neutrophils with the cytochrome C reduction method. Neutrophils were separated from the peripheral venous blood of healthy adults. The control group was treated with air PO_2 220mmHg). In the hypoxia group, neutrophils were treated with N_2 for 30 min. In the reoxygenation group, neutrophils were treated with oxygen for 15 min after pretreatment with N_2 for 30 min. Superoxide anion production was measured with the cytochrome C reduction method, after stimulation of phorbol myristate acetate or without such stimulation. Hypoxia caused a significant decrease of O_2^- production. Reoxygenation caused a significant increase of O_2^- production from unstimulated neutrophils. These results suggest that neutrophils were activated by hypoxia-reoxygenation.As the second study, the effects of hypoxia-reoxygenation on NADPH oxidase activity and the change of intracellular Ca ion neutrophils were investigated. In the control group cell membranes of neutrophils were treated with air(PO_2 220mmHg). In the hypoxia group, cell membranes was treated with N_2 for 30 min (PO_2 28mmHg). In the reoxygenation group, cell membranes were treated with oxygen for 15 min after pretreatment with N_2 for 30 min (PO_2 480mmHg. In the oxygenation group, that was treated with oxygen for 15 min (PO_2 480mmHg). NADPH oxidase activity was measured with the cytochrome C reduction method. Hypoxia caused a significant decrease of NADPH oxidase activity. Reoxygenation caused significant increase of NADPH oxidase activity. These results suggest that NADPH oxidase was activated by hypoxia-reoxygenation.Intracellular Ca ion was increased with reoxygenation. This finding suggested the increased intracellular Ca ion activates NADPH oxidase in the neutrophils.