Abstract 747: A Novel Aminopeptidase N - Ang IV Signaling Pathway Regulates Basolateral Na/K ATPase in Adaptation to High Salt
We have reported that aminopeptidase N/CD13 (Anpep), which cleaves an amino-terminal arginine from angiotensin III (Ang III) to yield the natriuretic hexapeptide angiotensin IV (Ang IV), exhibits greater renal tubular expression in the Dahl salt-resistant (SR) rat than its Dahl salt-sensitive (SS) counterpart and maps to a QTL for SS hypertension (Hypertension 43:282–5, 2004). Based on this, we have hypothesized that renal tubule Anpep plays a mechanistic role in adaptation to high salt. In this work, regulation of basolateral Na/K ATPase by Anpep was examined in LLC-PK1 cells, a proximal tubule (PT) renal epithelial cell line. Transfection of cells with siRNA to Anpep and Anpep over-expression (OE) increased and decreased, respectively, basolateral Na/K ATPase, measured by biotinylation and confocal immunohistochemistry, and oubain-inhibitable ATPase activity. In transgenic mice over-expressing Anpep in epithelial cells, there was reduced expression of basolateral Na/K ATPase in PT and distal tubules (DT). Together, these findings indicate that Anpep inhibits basolateral Na/K ATPase. To probe the relevant secondary signaling by Anpep, regulation of cellular Ang IV by Anpep was examined in LLC-PK1 cells. Anpep OE increased (p < 0.05) intracellular Ang IV abundance, measured by quantitative immunohistochemistry (QIHC) using a polyclonal Ab to Ang IV. siRNA to the Ang IV receptor (AT4 or insulin-regulated aminopeptidase (IRAP)) prevented the decrease in Na/K ATPase associated with Anpep OE. Together, these results suggest that Anpep reduces basolateral Na/K ATPase in nephrons and that the signaling pathway is via Ang IV and the AT4 receptor. This identifies a novel signaling pathway that may play a mechanistic role in adaptation to high salt and serve as a therapeutic target for hypertension.