Kouzan S, Nolan R D, Fournier T, Bignon J, Eling T E, Brody A R
Laboratory of Pulmonary Pathobiology, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709.
Am Rev Respir Dis. 1988 Jan;137(1):38-43. doi: 10.1164/ajrccm/137.1.38.
In previous studies on arachidonic acid (AA) metabolism by pulmonary macrophages in vitro, we observed that the presence of serum in the culture medium influenced the profile of AA metabolites released. To further characterize this phenomenon, rat alveolar macrophages were placed in plastic tissue culture dishes and allowed to adhere in the presence or absence of 7.5% fetal bovine serum (FBS) for 1 h. Adherent cells were then maintained in medium (equilibration) with or without FBS for 3.5 h before stimulation with the calcium ionophore A23187. The release of thromboxane B2 (TXB2) (the stable metabolite of TXA2) and leukotriene B4 (LTB4) during culture was measured by radioimmunoassay and reverse-phase high pressure liquid chromatography, respectively, at the end of each culture step. Cell adhesion to the plastic substrate in FBS-free medium induced an intense stimulation of AA metabolism, with the release of both TXB2 and LTB4. Adhesion and the accompanying TXB2 release appear to be mediated by trypsin-sensitive components since trypsin-pretreated macrophages showed a dramatic reduction in both adherence and TXB2 synthesis. The presence of FBS during the attachment phase of culture reduced both adhesion and release of TXB2 and LTB4 by more than 50%. On the other hand, addition of FBS to cells that had completed adhesion in serum-free medium stimulated release of both metabolites. When challenged with calcium ionophore after 4.5 h of culture, macrophages that had adhered in FBS-free medium released a much smaller amount of TXB2 than did macrophages that had been cultured in the presence of FBS.(ABSTRACT TRUNCATED AT 250 WORDS)
在之前关于肺巨噬细胞体外花生四烯酸（AA）代谢的研究中，我们观察到培养基中血清的存在会影响AA代谢产物的释放谱。为了进一步表征这一现象，将大鼠肺泡巨噬细胞置于塑料组织培养皿中，在有或无7.5%胎牛血清（FBS）的情况下使其贴壁1小时。然后，在无血清或有血清的培养基（平衡）中维持贴壁细胞3.5小时，之后用钙离子载体A23187刺激。在每个培养步骤结束时，分别通过放射免疫测定法和反相高压液相色谱法测量培养过程中血栓素B2（TXB2）（TXA2的稳定代谢产物）和白三烯B4（LTB4）的释放。在无FBS培养基中细胞与塑料底物的贴壁诱导了AA代谢的强烈刺激，伴随着TXB2和LTB4的释放。贴壁以及随之而来的TXB2释放似乎由胰蛋白酶敏感成分介导，因为经胰蛋白酶预处理的巨噬细胞在贴壁和TXB2合成方面均显著降低。培养贴壁阶段FBS的存在使TXB2和LTB4的贴壁及释放减少超过50%。另一方面，向在无血清培养基中完成贴壁的细胞添加FBS会刺激两种代谢产物的释放。在培养4.5小时后用钙离子载体刺激时，在无FBS培养基中贴壁的巨噬细胞释放的TXB2量比在有FBS存在下培养的巨噬细胞少得多。（摘要截短于250字）
