Characterization of angiotensin II-receptor subtypes in podocytes.
Glomerular podocytes play a key role in maintaining the integrity of the glomerular filtration barrier. This function may be regulated by angiotensin II (Ang II) through activation of cell-surface receptors. Although studies suggest that podocytes express receptors for Ang II, the Ang II binding site has not been characterized with radioligand binding techniques. We therefore used iodine 125-labeled Ang II to monitor Ang II-receptor density during differentiation of a mouse podocyte cell line. Scatchard analyses of equilibrium binding data revealed a single class of high-affinity binding sites (dissociation constant approximately 3 nmol/L) in both differentiated and nondifferentiated cells. During differentiation, the density of Ang II-receptor sites increased roughly 15-fold in differentiated podocytes (maximal density of specific binding sites 881 fmol/mg protein) compared with that in nondifferentiated cells (52 fmol/mg protein; P<.005). Glomerular podocytes expressed messenger RNA for AT1A, AT1B, and AT2 receptor subtypes, and competitive binding studies found that differentiated podocytes expressed mostly AT1 receptors (approximately 75%) with lesser amounts of AT2 (approximately 25%). Up-regulation of Ang II-receptor number was associated with increased Ang II-receptor responsiveness, as evidenced by enhanced Ang II-stimulated inositol phosphate (IP) generation and incorporation of tritiated thymidine. Both [3H]thymidine incorporation and IP generation were mediated by AT1-receptor activation. These data suggest that glomerular podocytes express a high-affinity binding site for Ang II with pharmacologic characteristics of both AT1 and AT2 receptors. This receptor site is up-regulated during podocyte differentiation, and receptor activation induces both IP generation and DNA synthesis by AT1-dependent mechanisms. We speculate that activation of podocyte Ang II receptors contributes to glomerular damage in disease states.
Wang
Liming
L
Division of Nephrology, Department of Medicine, Duke University Medical Center and Durham Veterans Affairs Medical Center, NC 27710, USA.
Flannery
Patrick J
PJ
Spurney
Robert F
RF
D000818:Animals; D002454:Cell Differentiation / Q000502:physiology; D002461:Cell Line, Transformed; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism*; D015870:Gene Expression; D007457:Iodine Radioisotopes; D007678:Kidney Glomerulus / Q000166:cytology*; D051379:Mice; D011869:Radioligand Assay; D044140:Receptor, Angiotensin, Type 1 / Q000235:genetics / Q000378:metabolism*; D044139:Receptor, Angiotensin, Type 2 / Q000235:genetics / Q000378:metabolism*
D007457:Iodine Radioisotopes; D044140:Receptor, Angiotensin, Type 1; D044139:Receptor, Angiotensin, Type 2