Angiotensinogen and AT1 antisense inhibition of osteopontin translation in rat proximal tubular cells

Antisense oligonucleotide inhibition of angiotensinogen and ANG II type 1 receptor (AT1) mRNA translation in rat proximal tubules (PT) was examined to provide direct evidence for a role of the renin-angiotensin system (RAS) in upregulated osteopontin expression observed following mechanical cell stretch. Male Sprague-Dawley rats underwent unilateral ureteral obstruction (UUO) under Brevital anesthesia. In situ hybridization and Western blot analysis demonstrated angiotensinogen mRNA and angiotensin converting enzyme (ACE) protein localized to PTs and upregulated in obstructed kidneys, respectively, confirming an increased expression of renal RAS in vivo. In vitro studies were performed to provide mechanistic insight into ANG II-dependent osteopontin expression following mechanical cell stretch, which putatively mimics the increased PT luminal pressure post-UUO. A cationic transfection method was used to introduce either angiotensinogen or AT1 antisense oligonucleotide into cultured rat PT cells prior to 1...