Expression of the renin-angiotensin system in a human placental cell line.

BACKGROUND: The renin-angiotensin system (RAS) is present in human placental tissue and participates in regulation of maternal-fetal blood flow during pregnancy. RAS expression in placental tissue is regulated by various hormones and is altered in various disease conditions. An in vitro system is needed to further investigate regulation of the placental RAS. To this end, we studied RAS expression in the human placenta-derived cell line, CRL-7548. METHODS: CRL-7548 cells were cultured in plastic plates. Total RNA was extracted, reverse transcribed, and amplified by polymerase chain reaction (PCR) with specific primers. Angiotensin II peptide in the culture media was measured by radioimmunoassay. Renin activity was detected by radioimmunoassay measuring angiotensin I generated. Angiotensin receptor type I was detected by Western blot. RESULTS: Specific mRNA for angiotensin, renin, angiotensin converting enzyme, and angiotensin receptor type I was detected by real-time PCR. Renin activity was detected in the placental cell lysate, and angiotensin II peptide, the final product of the RAS system, was detected in cell culture media by radioimmunoassay. Angiotensin receptor type I was identified as a 41 kDa protein in cell lysates by Western blot. CONCLUSIONS: These results demonstrate that all necessary components of the classic RAS are expressed in the human placental cell line CRL-7548. This cell line may prove useful as an in vitro system for studying RAS regulation in the placenta.

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.