Lung vascular effects of phorbol myristate acetate in awake sheep.
Lung vascular effects of phorbol myristate acetate in awake sheep.
复制标题
佛波醇肉豆蔻酸酯醋酸酯对清醒羊肺血管的影响。
DOI：
10.1152/jappl.1983.54.1.267
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发表时间：
1983
期刊：
影响因子：
--
通讯作者：
Brigham,KL
中科院分区：
文献类型：
--
作者：
Loyd,JE;Newman,JH;English,D;Ogletree,ML;Meyrick,BO;Brigham,KL
方法绵羊和人粒细胞的体外反应。通过Weening等人使用的改良方法分离用于体外研究的粒细胞。（30）.将肝素化血液在Ficoll-Hypaque垫上离心。收集通过梯度的细胞（粒细胞和红细胞），用等渗氯化铵裂解红细胞。将细胞洗涤两次，并以3.0 × 107中性粒细胞/ml的浓度重悬于Hanks平衡盐溶液中。通过台盼蓝排除法评估，最终制剂含有> 95%的活粒细胞。超氧化物（02.）PMA刺激的人和绵羊粒细胞的生成通过细胞色素c还原测定使用先前描述的方法（6）来测量。将粒细胞悬液、细胞色素c和缓冲液置于37”，并用400 ng/ml PMA刺激。对于基线值，使用BSS代替PMA。分光光度计空白另外含有50 μ g/ml超氧化物歧化酶，以特异性防止超氧化物还原细胞色素c。孵育20 min后，通过加入超氧化物终止Oy依赖的细胞色素c还原
METHODSIn vitro responses of sheep and human granulocytes. Granulocytes for in vitro studies were isolated by a modification of the method used by Weening et al.(30). Heparinized blood was centrifuged on cushions of Ficoll-Hypaque. Cells that passed through the gradient (granulocytes and erythrocytes) were collected, and the erythrocytes were lyzed with isotonic ammonium chloride. Cells were washed twice and resuspended in Hanks’ balanced salt solution at a concentration of 3.0 x lo7 neutrophils/mI. The final preparations contained> 95% viable granulocytes as assessed by trypan blue exclusion. Superoxide (02.) generation by PMA-stimulated human and sheep granulocytes was measured by the cytochrome c reduction assay using a previously described method (6). Granulocyte suspensions, cytochrome c, and buffer were brought to 37” and stimulated with 400 ng/ml of PMA. For base-line values, BSS was used in place of PMA. Spectrophotometric blanks contained in addition 50, ug/ml of superoxide dismutase to specifically prevent the reduction of cytochrome c by superoxide. After incubation for 20 min, Oy-dependent cytochrome c reduction was terminated by addition of superoxide