Sample Collection and Measurements
Blood samples were obtained from patients with diabetic foot ulcers before treatment was initiated. A total of 5 mL blood samples were taken from the volunteers: 3 mL blood samples were placed in tubes containing anticoagulants to obtain whole blood, and 2 mL blood samples were placed into dry tubes. Serum samples were obtained by centrifuging the dry tube blood samples at 3500 g for 10 minutes. Whole blood and serum samples were kept at -80oC until the study day. The body mass index (BMI) was calculated by dividing the body weight in kilograms by the square of the body height in meters (kg/m2).
In the whole blood samples, the amount of glycated haemoglobin (HbA1c) was determined on an analyzer (Arkray Adams HA-8160 from Japan) working with a high pressure liquid chromatography method. The serum levels of low-density lipoprotein (LDL), high-density lipoprotein (HDL), triglyceride, total cholesterol and glucose were measured by autoanalysis using photometric methods (Architect C 16200 Abbott Laboratories Inc., Abbott Park, IL, USA). The levels of CRP were detected by a BNR II SYSTEM SN 442176 instrument.
The serum IMA levels in both groups were measured according to the method described by Bar-Or et al.19 A 200 µL serum sample was placed in a glass tube, 50 µL cobalt chloride added and the mixture then incubated for 10 min at 24oC before 50 µL dithiothreitol was added to the mixture. After 2 minutes, 1 mL of sodium chloride solution was added. The IMA concentrations were calculated by measuring the absorbance of the coloured complex at 470 nm wavelength. The serum IMA levels were expressed as absorubans (ABSU).