Egulated in obese subjects as indicated by increased levels of C-peptide (P = 0.03), glucagon (P = 0.048), leptin (P,0.0001) and PAI-1 (P = 0.013). Likewise, obese subjects displayed higher inflammatory response as measured by IP-10 and RANTES chemokines (P = 0.003 and P = 0.012, respectively), but they did not differ significantly in the rest of the inflammatory mediators as well as the oxidative stress markers (Table 2 and data not shown).RT2 Profiler PCR Array for Hsp-related genesDespite the central role that HSPs play against a variety of diseases, only a few studies that documented their role in obesity, insulin resistance and diabetes. In addition, these studies were primarily focused on HSP-25, HSP-72 and to a lesser extent; HSP60 [8,9,33,39]. Therefore, we decided to perform an expression profiling analysis on a selected set of Hsp-related genes using RT2Profiler PCR heat shock array. RNA samples prepared from PBMC of 6 non-diabetic subjects divided into lean and obese Table 2. Clinical and biochemical characteristics of subjects at baseline.ImmunohistochemistryFormalin fixed, paraffin embedded adipose tissue samples were prepared and used to make sections for immunohistochemical studies as described previously [68]. Briefly, sections were deparaffinized and the antigens were retrieved at high-temperature using antigen unmasking solution (Dako, Denmark). The endogenous peroxidase was quenched using 3 H2O2 (Merck Schuchardt, Gemany) for 60 min at RT. Sections were blocked with 5 fat-free milk for 60 min at RT followed by 1 BSA for another 60 min and then, incubated at 4uC for overnight with primary antibodies. After washing, sections were stained with horseradish conjugated secondary antibody (Dako, Denmark) for 60 minutes at RT. Colors were developed using DAB kit (Dako, Denmark) and sections were counterstained with hematoxylin (Sigma Aldrich, St. Louis, MO). Quantification of the immunohistochemical staining data was done using Aperio software version 6.3 (Molecular Devices, Downingtown, PA) with an established arbitrary threshold.Lean (n = 54) Obese (n = 66) Resting HR (beat/min) SBP (mmHg) DBP (mmHg) VO2,maxP-value0.Glycine 73 0.SS-208 01 0.13 0.03 0.31 ,0.0001 0.14 0.0002 0.17 0.056 0.03 0.048 0.68 0.PMID:23775868 12 ,0.0001 0.013 0.53 0.83 0.54 0.43 0.003 0.012 0.24 0.80.71614.51 113.00610.81 76.4366.33 21.6363.76 5.0360.92 1.4360.49 3.1360.85 0.9160.43 5.0260.64 5.4760.40 2.4460.71 0.6560.11 2.6260.83 2.3561.12 4.8263.03 3.1961.55 23.7069.30 1.2060.47 4.9962.18 1.9761.39 0.3960.14 1.2960.60 1.3860.37 1.2960.77.4368.15 127.50611.89 82.00610.14 17.4864.83 5.2661.02 1.1360.24 3.4360.96 1.5260.91 5.4560.92 5.9361.03 3.6862.31 0.7160.15 2.6861.54 4.5162.11 9.7266.91 4.1461.53 25.70613.20 1.3560.85 4.6861.87 2.3362.24 0.6060.32 1.7860.86 1.5860.63 1.5260.(ml/kg/min)Cholesterol (mmol/l) HDL (mmol/l) LDL (mmol/l) TG (mmol/l) Glucose (mmol/l) HBA1C ( ) C-peptide (ng/ml) Glucagon (ng/ml) GLP-1 (ng/ml)Statistical analysisStatistical analyses were performed with SAS version 9.2 (SAS Institute Inc, Cary, NC). Unless otherwise stated, all descriptive statistics for the variables in the study were reported as means 6 standard deviation (SD). Student’s t-test was used to determine significance of difference in means between the two groups. Correlations between variables were calculated with the Spearman’s rank correlation test. Differences were considered statistically significant at P-values less than 0.05.Insulin (ng/ml) Leptin (ng/ml) PAI-1 (ng/ml) TNF-a (pg/.