Angii

All groups of rats were fed and maintained on a normal salt diet (NSD, 0.4% NaCl, Teklad, Harlan Laboratories, Madison, WI). Rats were fed a high-salt diet for 7 days (HSD, 4% NaCl, Teklad, Harlan Laboratories), or infused with AngII (AngII, 200 ng/kg/min (Moreno et al. 2003 (link)), Phoenix Pharmaceuticals, Burlingame, CA), or a combination of both. Osmotic mini-pumps containing AngII (model 2002; Alzet, Palo Alto, CA) were implanted subcutaneously under isoflurane anesthesia and sterile conditions according to the manufacturer's instructions as described previously in a subgroup of rats (Loria et al. 2010b (link)).

Adult male mice (25–30 g) were anesthetized with isofluorane and a 14-day osmotic mini-pump (Durect, Cupertino, CA) containing vehicle or AngII (Phoenix Pharmaceuticals, Burlingame, CA) to deliver at the rate of 1.4 mg/kg/d was placed subcutaneously in the mid-scapular region. Systolic blood pressure in conscious mice was measured by the tail-cuff method (CODA, Kent Scientific, Torrington, CT) after 2 weeks of AngII infusion. Mice were then anesthetized with pentobarbital (50 mg/kg i.p.) and both muscles of the diaphragms were cut. The thoracic aorta (from aortic arch to diaphragm) from each animal was removed, carefully cleaned and either used for reactivity or snap frozen in liquid nitrogen and stored at −80°C. The procedures used in this study were approved by the University of Michigan Committee on the Use and Care of Animals and the Albert Einstein College of Medicine Institutional Animal Care and Use Committee. The University of Michigan Unit for Laboratory Animal Medicine provided veterinary care for the animals reported in this study. Both the University of Michigan and Albert Einstein College of Medicine are accredited by the American Association of Laboratory Animal Care. The animal care and use programs conform to the standards in “The Guide for the Care and Use of Laboratory Animals,” Department of Health, Education, and Welfare Publication No. 86-23.

Pathological LVH was generated by angiotensin II (angII, Phoenix Pharmaceuticals Inc., CA, USA) infusion via subcutaneous Alzet osmotic minipumps (Durect Corporation, Cupertino, CA, USA) for two weeks (0.1 mg/kg/h). Five-to-seven-month-old male VEGFR-1 TK^−/− mice [12 (link)] and their littermate controls were anesthetised with isoflurane (induction: 4.5% isoflurane, 450 mL air, maintenance: 2.0% isoflurane, 200 mL air, Baxter 28 International Inc., Deerfield, IL, USA) for the operation and an analgesic (carprofen 5 mg/kg, s.c.; Rimadyl, Pfiser Inc., New York, NY, USA) was administered. The mice were kept in standard housing conditions in the National Laboratory Animal Centre of the University of Eastern Finland. Diet and water were provided ad libitum. Mice were euthanized with CO₂ and perfused with phosphate-buffered saline (PBS). All animal procedures were approved by the Animal Experiment Board in Finland and were carried out in accordance with the guidelines of the Experimental Animal Committee of the University of Eastern Finland. The animal licence number is ESAVI-2014-5089.