Adiponectin Adipose Tissue Expression And Gene Polymorphism Associated With Diabetes Research

Objective:The aim of this study was to probe single nucleotide polymorphyism (SNP) of human adiponectin gene by using allelic 3'terminal specific primes Real-time PCR, to analyze the correlation between concentration of adiponectin in sera and apM1 gene , to study the relationship between SNP in apM1 gene and insulin resistance, and to investigate the factors which effect level of adiponectin in patients with T2DM and obesity.Subjects and Methods:1 Subjects:All subjects Han nationality with no consanguinity, were from Jinan district in Shandong province. Three groups: T2DM (DM) group, obese (OB) group and normal control (NC) group. The T2DM group was again divided into two groups: T2DM with obesity (DM-OB) group, and non-obese T2DM (DM-N-OB) group. T2DM group: all patients were diagnosed according tothe 1999 WHO criteria and exclued renal disease. 69 patients were included, 35 male and 34 female. The mean age was (57.2 ±10.5) years old. The course of type 2 diabetes was 2 to 17 years. Obese group: All patients were diagnosed according to the 1998 WHO criteria. The mean body mass index (BMI) of all subjects were over 25 kg/m2, 52 subjects were chosen, 28 male and 22 female, the mean age was (39.4+11.0) years old. All patients were excluded heart disease, cerebrovascular disease and hyperpressure, no diabetes history. Normal control(NC) group consisted of 66 subjects, 35 male and 28 female, the mean age was (35.2 + 12.0) years old. All subjects were excluded disease history of heart, kidney, cerebrovascular disease and diabetes. All paticipants gave informed consent before participation.2 Methods:Fasting 12h over night venous blood was collected. Sera was saved for measurement of fasting blood glucose (FBG, glucose oxidase assay), insulin (FINS, time-resolved immunofluorometrc assay, TRIFMA), blood lipid (TG, TC, HDL) levels and Adiponectin concentration (ELISA). Homa-IR were calculated according to FBG and FINS. CV of intra-group and inter-group were 2.7% and 3.8% (FINS) and <2.5%, <3.2% respectively for adiponectin. All samples were analyzed in duplicate and the mean value were defined as the final value for every subjects.apMl gene SNP analysis:DNA was extracted from whole blood with EDTA-K2anticoagulant using the Promega kit.SNP +45 T/G and SNP +276 G/T were genotype using the allelic 3'terminal specific primers and perform Real-time PCR using SYBR Green dye, SNP genotype was carried out using melting curve analysis.3 Statistical analysis:All statistic were performed by using SPSS software (version 10.0). Skewes variables (blood lipid, insulin, Homa-IR and adiponectin) were log transformed to normalize their distribution before statistical analyses. The numerical data are expressed as mean + SD. ApMl gene +45T/G, +276G/T genotype frenquencies and alleles frenquencies were calculated. A conditional logistic regression analysis, t-test, analysis of variance(ANOVA),Chi-squared test were performed according to the different aims and data. Threshold for significance was set as p<0.05.Results:(1) Comparison of clinical biochemistric charateristics among three groups:1) There was difference of BMI, WHR, FBG, TG, TC, and Homa-IR between OB and NC group (p <0.05); HDL and adiponectin concentrations were significantly lower in OB group than in NC group (p <0.001).2) No statistical difference of BMI, WHR, TG, TC, HDL, FINS was observed between OB group and DM-OB group; Homa-IR was greater in DM-OB group than in OB group (p <0.05); adiponectin level was significantly lower inDM-OB group than in OB group (p <0.01).3) We found no statistical difference of HDL, Homa-IR, adiponectin concentration between OB group and DM-N-OB group; TG was greater in OB group than in DM-N-OB group (p <0.05).4) WHR, FBG, TG, Homa-IR were much lower in NC group than in DM-N-OB group (p<0.05, p<0.001) ; adiponectin was greater in NC group than in DM-N-OB group (p <0.05, p <0.001).(2) Serum adiponectin concentration was significantly negatively correlated with BMI, WHR and Homa-IR (p<0.001), the correlation was more significant in OB and DM group.(3) The results of apMl gene SNP genotype:1) The genotype apMl gene at +45 site was significantly correlated with T2DM. T/T genotype in DM group was significantly greater than in OB and NC group (p <0.001). The risk of T2DM incidence in persons with TT genotype was 4.6 times than G ( G/T + GG) genotypes.2) No difference of apMl gene at +276 site was found among three groups.3) The SNP of apMl gene at +45 and +276 sites had no correlation with obesity.4) In NC group, Homa-IR in subjects with G/G genotype was significantly greater than with T/T genotype (p <0.05). Both adiponectin concentration and HDL were significantly lower than that with T/T genotype (p <0.05).Conclusion:1. apMl gene SNPs exist in exon 2 and intron 2 in type 2 diabetes of Han population in Jinan district.2. The genotype apMl gene at +45 site was significantly correlated with T2DM.3. The SNP of apMl gene at +45 and +276 sites had no correlation with obesity and normal population.4. Adiponectin concentration in OB and T2DM was significantly lower than NC. Serum adiponectin content was significantly negatively correlated with BMI, WHR and Homa-IR.