Detection of a receptor for angiotensinogen on placental cells

Abstract Background Current evidence indicates that there may be a tissue-specific renin-angiotensin system (RAS) in the human placenta. To better define the placental RAS, this study sought to determine whether placental derived cells possessed a receptor for angiotensinogen (AGT), a rate limiting component of the RAS. Methods A human placenta–derived cell line, CRL-7548, a highly purified AGT and iodine-125–labeled angiotensinogen (125I-AGT) were used in this study. The cells were seeded in 35-mm diameter plastic wells, cultured for 2 days in fetal bovine serum–Dulbecco’s modified Eagle’s medium to reach about 80% to 90% confluence, 2 × 104 cells/well and passed in Dulbecco’s modified Eagle’s medium free of fetal bovine serum before experimentation. A quantity of 3 μg of 125I-AGT was added to each well at zero time. Results The cells rapidly bound 125I-AGT in a time dependent manner with saturation being achieved in 2 to 4 h. This binding was competitively inhibited by unlabeled AGT. Prior addition of a 100-fold excess of unlabeled AGT resulted in a 54% decrease in maximal binding. Bound AGT was also competitively displaced by AGT. Addition of a 200-fold excess of unlabeled AGT displaced 70% of the bound 125I-AGT. Thus, approximately 30% of the total binding can be attributed to nonspecific binding. An acid wash (0.2 mol/L acetic acid, 0.5 mol/L NaCl, pH 2.5) at 4°C removed 46% of the bound 125I-AGT. An acid wash is known to dissociate cell surface ligand-receptor complexes, but does not remove internalized ligand. Conclusion The results of this study provide evidence for the existence of an AGT receptor on placenta-derived cells. This is the first report of an AGT receptor.