Plasma cystatin c

ELISA kits were used to measure plasma cystatin C (R&D Systems, Inc., Minneapolis, MN) and human VEGF protein in conditioned media harvested from cultured cells (R&D Systems, Inc., Minneapolis, MN) according to the manufacturer’s protocol.

Fasting blood glucose was measured from tail blood by glucometer (Medisense, Abbott Laboratories, Bedford, MA). Urine was collected from mice housed in metabolic cages for 6 h. Heparinized whole blood was collected by cardiac puncture in anesthetized animals for analysis of HbA₁c and isolation of heparinized plasma. Urine creatinine levels were determined by the Jaffe rate reaction method. ELISA kits were used to assess levels of urine albumin (Bethyl Laboratories, Montgomery, TX) and plasma cystatin C (R & D Systems, Minneapolis, MN). Albuminuria was measured by calculating the urine albumin/creatinine ratio (ACR). HbA1c was measured by DCA Vantage Analyzer (Siemens, Camberley, UK).