The purpose of this study was to gather additional evidence in irradiated rat lung on the relationship between annexin I and prostaglandin synthesis. The right hemithorax of the animal was exposed to a single dose of 0 or 30 Gy of X-rays, and the animals were killed 3 months postirradiation. Levels of annexin I and synthesis of prostacyclin (PGI2) were determined in lungs, in cell-free bronchoalveolar lavage (BAL) fluid, and in macrophages lavaged from those lungs. In addition, protein concentration, lactate dehydrogenase (LDH) activity and macrophage count in BAL fluid obtained from irradiated lung were compared with that from sham-irradiated (0 Gy) lung. Levels of annexin I, the putative inhibitor of phospholipase A2, in lung and cell-free BAL fluid were decreased in samples from irradiated animals. By contrast, the level of annexin I in macrophages lavaged from irradiated lung was higher than that in macrophages from sham-irradiated lung. The irradiated lung produced nearly 3.5 times more prostacyclin than did the control lung. However, prostacyclin synthesis by macrophages lavaged from irradiated lung was no different than that of macrophages from sham-irradiated lung. Protein, LDH and macrophage number in BAL fluid from irradiated lungs were significantly higher than in corresponding specimens from sham-irradiated lungs. These data demonstrate that reduced levels of annexin I, as well as increased protein concentration, LDH activity and macrophage numbers in irradiated rat lung are reflected in BAL fluid. Therefore, information obtained from BAL fluid, but not from BAL macrophages, reflects lung status, and may serve as a minimally invasive index of radiation pneumonitis in this model. In irradiated lung, increased PGI2 synthesis coupled with a decreased annexin I level are consistent with the hypothesis of an inhibitory role of annexin I in prostaglandin metabolism. However, this hypothesis is not supported by findings in BAL macrophages, where increased annexin I concentration is not accompanied by a decrease in PGI2 production. In view of the latter findings, and a previous study from our laboratory demonstrating that phospholipase activity in irradiated rat lung is in fact decreased, despite the reduction in annexin I concentration and the hyperproduction of prostanoids, it would seem unlikely that annexin I negatively modulates prostaglandin synthesis via inhibition of phospholipase in this model.