Patients
We recruited 68 drug-naïve and newly diagnosed diabetic patients (male and female) aged from 25 to 60 years. Thirty-one patients were lean with body mass index (BMI) less than 25 kg/m2 and 37 patients were obese with BMI at least 25 kg/m2. The patients eligible for enrollment had a fasting plasma glucose (FPG) level no more than 11.1 mmol/l. The patients were excluded if they had type 1 or type 2 diabetes with diabetic ketoacidosis (DKA), infection and other stress status, autoimmune disease, hepatic and renal diseases, severe heart failure (NYHA III and IV), symptomatic heart failure, established edema, or increased risk of fractures.
Study Design
Eight hospitals in six provinces and cities in China participated in the study. Euglycemic hyperinsulinemic clamp test was used to evaluate the status of insulin resistance. The acute insulin response (AIR) was adopted to determine β-cell function of first phase insulin secretion. In order to investigate the mechanism of glucose reduction, the changes of insulin resistance and pancreatic β-cell function were analyzed after normalization of glucose control which was achieved by the PIO therapy. Medications other than PIO such as those for controlling dyslipidemia and hypertension remained unchanged during entire the trial. Participants were asked to maintain their usual lifestyles including diet and physical activities throughout the study.
All procedures followed were in accordance with the ethical standards of the responsible committee on human experimentation (institutional and national) and with the 1964 Declaration of Helsinki, as revised in 2013. This study was approved by the Ethics Committee of Fuwai Hospital Chinese Academy of Medical Sciences. All patients gave written informed consent prior to data collection.
Study Procedures
Patients were required to fast overnight before undergoing an intravenous glucose tolerance test (IVGTT) the next morning. Blood samples were collected at 0, 3, 5, 7, and 10 min for the measurements of glucose and insulin. Glycated hemoglobin (HbA1c), hepatic and renal function, lipid profile, and urine tests were also examined under the fasting status. After that, the euglycemic hyperinsulinemic clamp test was performed on the same day. Insulin (Novolin R; NovoNordisk, Bagsvaerd, Denmark) in normal saline solution and 20% glucose solution were infused separately into the ulnar veins. The dorsum manus vein was used for extracting arterialized venous blood after warming the arm to 50–55 °C with an infrared heater. The insulin infusion rate was adjusted according to the target plasma glucose level and maintained at 1 mU/kg/min for the next 120 min, and blood was drawn every 5 min to measure plasma glucose concentration. Adjustment of the infusion rate of 20% glucose solution was made to keep the blood glucose approximately at the target value, 5.0 mmol/L. The mean glucose infusion rate (GIR) value was estimated to be equal to the GIR in the last 30 min of steady state. Insulin in the plasma samples was analyzed using a immunoradiometric assay (IRMA, CIS bio international, Bagnols, France). Additional baseline assessments included blood pressure, height, and body weight.
PIO treatment was initiated after the completion of the tests described above. A dose of 30 mg per day was administered initially and increased to 45 mg if needed. The duration of treatment was 16 weeks. The patients were followed monthly and PIO dosages were adjusted to achieve the target goal of plasma glucose control (FPG < 6.1 mmol/L and 2-h plasma glucose level, 2hPG < 7.8 mmol/L). During each follow-up visit, levels of FPG and 2hPG were measured, and urinalysis was performed. All measurements at baseline, including the IVGTT and the euglycemic hyperinsulinemic clamp test, were repeated at the end of the 16-week PIO treatment.
Measurements and Calculations
Insulin sensitivity was evaluated by GIR (mg/kg/min) based on the euglycemic hyperinsulinemic clamp test. The area under the curve of insulin during IVGTT was quantified as first phase insulin secretion (AIR, min × µU/ml). Glucose disposal index (DI), the products of GIR and AIR, was also calculated. Ratios of the area under the curve of insulin (AUC-Insulin) to the area under the curve of glucose (AUC-Glucose, Ratio AUC-I/G) were calculated to evaluate β-cell sensitivity to glucose. Plasma levels of leptin, adiponectin (APN), and high sensitivity C-reactive protein (hs-CRP) were also measured at baseline and at the end of the 16-week PIO treatment.
Statistical Analysis
Mean value and standard deviation (SD) were reported for continuous variables. Number and percentage (%) were used for categorical variables. For right-skewed distribution variables, a median, 25 percentile and 75 percentile of the measure were reported. The changes of insulin sensitivity, plasma insulin, and glucose DI levels from baseline to the end of PIO treatment in each subgroup (the obese group and the lean group) were analyzed by pairwise t tests. A mixed model for repeated measures (MMRM) analysis was used to compare the differences between the lean and the obese group. For variables not normally distributed, npar1way analysis was performed to find the differences between the lean and the obese group. The adverse events related to the study were assessed.