FLOW CYTOMETRIC DETECTION OF ABNORMAL FETAL ERYTHROPOIESIS - APPLICATION TO 5-FLUOROURACIL-INDUCED ANEMIA

We sought to determine whether flow cytometric analysis of circulating fetal blood cells could be used to rapidly detect perturbations of fe tal erythropoiesis. In addition, we wanted to determine whether this a pproach would allow sample collection by exsanguination instead of fet al cardiac puncture, a difficult technique used to prevent contaminati on of samples with maternal erythrocytes. To monitor fetal erythropoie sis from gestational day (GD) 14-20, we analyzed the cell size, RNA co ntent, and percentage of circulating liver-derived reticulocytes relat ive to yolk-sac-derived erythroblasts. As a model toxicant, we chose 5 -fluorouracil (5-FU), since we previously observed that maternal admin istration at 20-40 mg/kg on gestational day (GD) 14 produced fetal ane mia on GD 16-17, as evidenced by dose-dependent decreases in the cell counts, hematocrit, and hemoglobin content of fetal blood obtained by cardiac puncture. We report herein that 48 hr after maternal 5-FU admi nistration, both cardiac and peripheral blood samples exhibited a dose -dependent decrease in the relative percentage of reticulocytes, indic ating a reduced rate of reticulocyte release from the fetal liver. Mor eover, at 30 and 40 mg/kg, reticulocytes exhibited increased size and reduced RNA content on GD 16, but elevated RNA content (indicative of premature release) by GD 18. These data suggest that 5-FU inhibits bot h erythroid cell proliferation and RNA synthesis reversibly, resulting in an anemia that triggers compensatory release of immature reticuloc ytes. Thus, by using flow cytometry to analyze fetal blood, we were ab le to detect and characterize 5-FU-induced perturbations of fetal eryt hropoiesis. In so doing, flow cytometry afforded the advantages of rap id determination of individual cell type, size, and RNA content, and t he ability to exclude contaminating maternal erythrocytes from analysi s, thereby eliminating the need to acquire samples through the time-co nsuming technique of fetal cardiac puncture. (C) 1995 Wiley-Liss, Inc.