Quantichrom urea assay kit

BAECs at 80% confluence were given different treatment. The cell samples were collected, homogenized in 100 μl of distilled water and centrifuged at 12,000 rpm for 20 minutes at 4 °C and the supernatant was for urea measurement using QuantiChrom Urea Assay kit (Roche Diagnostics, Indianapolis, IN, USA). Results were expressed relative to protein content.

Adult WKYs and SHRs were adapted in metabolic cages (Harvard Apparatus, Holliston, MA, USA) for 3 days before a seven-day treatment. After rat bladder was emptied by gentle abdominal massage, urine was collected by spontaneous voiding every 24 hours. PU-14 (PU-14 was home synthesized with purity at 99% as determined by HPLC) at 50 mg/kg in 40% (g/ml) 2-hydroxypropyl-β-cyclodextrin was subcutaneously injected every 6 hours (0:30 a.m., 6:30 a.m., 12:30 p.m., and 6:30 p.m.) for one week16 (link). 40% 2-hydroxypropyl-β-cyclodextrin was used as a vehicle control. Two hours after the last administration, blood sample and thoracic aorta were collected under anesthesia with pentobarbital (1%) at 40 mg/kg body weight. The adequacy of anaesthesia was monitored based on the disappearance of the pedal with drawal reflex response to foot pinch. Urinary volume was measured by gravimetry, assuming a density of 1 g/ml. Urinary osmolality was measured by freezing point depression (Micro-osmometer, FISKER ASSOCIATES, Norwood, MA). Urea concentration was measured with QuantiChrom Urea Assay kit (Roche Diagnostics, Indianapolis, IN, USA).