Srebf1

To analyze the binding of SREBF1/HMGA2 protein to the GLUT4 gene promoter, adipocyte cells transfected with either SREBF1 or HMGA2 antibodies (Abcam) were used for ChIP analyses that were performed using a Pierce Agarose Chip Kit (EpiGentek Group, Inc., Farmingdale, NY, USA). Normal rabbit IgG was used as a negative control. Non-precipitated genomic DNA input was amplified as an input control. After purification, the DNAs were used for PCR analyses that were performed using primers (Sangon Biotech Co., Ltd., Shanghai, China) that encompassed the GLUT4 promoter region. The conditions used for PCR were as follows: denaturation at 95˚C for 2 min, followed by 40 cycles at 95˚C for 20 sec, 58˚C for 20 sec, and 72˚C for 20 sec.
Statistical analysis. All statistical analyses were performed using IBM SPSS Statistics for Windows, version 19.0 (IBM Corporation, Armonk, NY, USA). The levels of miR-33b-5p and gene expression are shown as the mean ± SD, and differences between groups were analyzed using Student's t-test. One-way ANOVA and Tukey's test were used to evaluate the levels of miR-33b-5p and gene expression in human adipocytes treated with different concentrations of glucose or insulin. The association between miR-33b-5p and SREBF1/HMGA2 expression was assessed using Pearson's correlation test. P-values <0.05 were regarded as statistically significant.