Abstract 16731: Megalin Inhibition Regulates Angiotensinogen and Angiotensin II in Mice
Objective: Angiotensinogen (AGT) is the unique precursor for generation of angiotensin II (AngII). Megalin interacts with both AGT and AngII in the kidneys. The purpose of this study was to determine whether inhibition of megalin influences systemic and local AGT and AngII regulation. We also started to explore the potential structural mechanisms of AGT interacting with megalin.
Methods and Results: Male C57BL/6J mice were fed a normal laboratory diet and injected intraperitoneally with PBS or megalin antisense oligonucleotides (ASO; 120 mg/kg/week) for 5 weeks. Urine was collected using metabolic cages 2 days prior to termination. At termination, plasma was collected with either EDTA alone or protease inhibitor cocktail to measure plasma AGT and renin, and angiotensin peptides, respectively. Megalin ASO administration reduced megalin mRNA by 80% in the kidneys, and led to striking increases of AGT and renin concentrations in urine (PBS versus megalin ASO for AGT: 12 ± 1 versus 3876 ± 509 ng/ml, P < 0.001; for renin: 30 ± 24 versus 1758 ± 509 ng AngI/ml/hr, P < 0.001). Surprisingly, megalin inhibition also modestly increased plasma AGT concentrations (~17%), accompanied by increased mRNA expression of AGT in the liver and kidneys. Plasma renin and AngII concentrations did not differ between PBS and megalin ASO injected mice. Renal AngII, but not AngI, was reduced by megalin inhibition. These findings demonstrate that megalin regulates liver and renal AGT expression as well as renal AngII production. Our bioinformatic analysis identified several highly conserved sequences of AGT including the loop formed by residues 291-301 containing strictly conserved solvent accessible hydrophobic residues W292, V299 and S298. Surface plasmin resonance demonstrated that mutation of residue 292 from W to A diminished its binding to megalin.
Conclusions: Megalin regulates systemic and renal AGT in addition to renal AngII production. A conserved residue W292 of AGT may contribute to AGT interaction with megalin.
Author Disclosures: H. Lu: None. D. Howatt: None. A. Balakrishnan: None. J. Xiao: None. D. Strickland: None. C. Vander Kooi: None. A. Mullick: Employment; Modest; Ionis Pharmaceuticals. M. Graham: Employment; Modest; Ionis Pharmaceuticals. A. Danser: None. A. Daugherty: None.
- © 2016 by American Heart Association, Inc.