Evaluating Aortic Endothelial Function in Mice

Wildtype male mice (n = 4, 12–15 weeks of age) were euthanized by overdose inhalation of isoflurane. Descending thoracic aortas were removed from mice and placed in ice-cold Krebs Hepes buffer while cleaned of adherent fat and connective tissues. Krebs Hepes buffer (pH 7.4, 37 °C) was composed of 114 mM NaCl, 4.7 mM KCl, 0.8 mM KH₂PO4, 1.2 mM MgCl₂ 6H₂O, 2.5 mM, CaCl₂ 2H₂O, 11.0 mM D-Glucose, 20 mM NaHCO₃, and 5 mM Hepes hemisodium salt. Krebs buffer was bubbled continuously with 95% O₂/5% CO₂ during myograph experiments. In brief, we used DMT 620M myograph chambers with the methods and conditions described in (41 ) for continuous measuring and recording of isometric tension with mouse aortas. The aorta from each mouse was divided into two groups: control (vehicle, PBS) and treatment (rhFABP3, 45 ng/ml). We tested the viability of aorta preparations (1–3 mm lengths) using 90 mM KCl. Viable tissues contractions were >1 mN. We assessed acetylcholine-induced relaxations of phenylephrine-contracted aortas under isometric tension conditions as we described previously (41 ). Aortic rings mounted in the DMT620 M chambers were exposed to treatments for 20 min then contracted with phenylephrine (3 μM) and then acetylcholine dose–responses curves constructed.