Specific receptor for angiotensinogen on human renal cells.

BACKGROUND: We recently demonstrated the existence of an angiotensinogen (AGT) receptor on placental cells. It has been established that there is a tissue-specific renin-angiotensin system (RAS) in the human kidney. This study focused on whether human renal proximal tubule epithelial cells possessed an AGT receptor. METHODS: Human renal proximal tubule epithelial cells were cultured in plastic wells. Binding assays were carried out by adding iodinated angiotensinogen ((125)I-AGT) to each culture well, with or without unlabeled AGT. The cells were washed, lysed, and the radioactivity was measured. RESULTS: Human renal proximal tubule epithelial cells bound (125)I-AGT in a time-dependent manner. This binding was competitively and specifically inhibited by unlabeled AGT. Bound (125)I-AGT was competitively displaced by AGT. Acid washing removed 30% at 8 h, indicating that 70% bound AGT had been internalized. Scatchard plot binding analysis showed that the identified AGT receptor possessed a single class of high-affinity binding sites (K(d)=1.73 nmol, B(max)=23.39 pmol/10(6) cells). CONCLUSION: The results of this study provide evidence for the presence of an AGT receptor on human renal proximal tubule epithelial cells. Our finding suggests that the AGT receptor may be an integral component of the renal RAS.

Detection of a receptor for angiotensinogen on placental cells.

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 ((125)I-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 x 10(4) cells/well and passed in Dulbecco's modified Eagle's medium free of fetal bovine serum before experimentation. A quantity of 3 microg of (125)I-AGT was added to each well at zero time. RESULTS: The cells rapidly bound (125)I-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 (125)I-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 degrees C removed 46% of the bound (125)I-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.