Obesity-Induced Methylation of Osteopontin Contributes to Adipogenic Differentiation of Adipose-Derived Mesenchymal Stem Cells

Figure 2

The methylation of the OPN promoter inhibits the transcription of OPN. (a) ASCs from HFD mice treated with 5 μmol 5-aza-2dC (HFD + 5-aza-2dC) for 2 days related to untreated (HFD) and control (CD) cells was determined by real-time PCR and western blot analysis. (b) Effect of siDNMT1 treatment on the relative abundance of transcript levels of DNMT1 and OPN. (c) Relative location for primers of MS-PCR. (d) MS-PCR analysis of OPN in ASC lines from CD and HFD mice. Same amount of DNA was amplified with primers specific to the unmethylated (U) or the methylated (M) CpG sites of the OPN gene after modification with sodium bisulfite. Methylation or unmethylation percentage in ASCs have been quantitated by ImageJ (National Institutes of Health) with normalization to positive (P) or negative (N) control. (e) MS-PCR analyzed the methylation patterns of OPN promoters in ASCs after treatment with 5-aza-2dC for 2 days. The schematic showed the methylation percentage in response of 5-aza-2dC treatment. P, methylated DNA; N, unmethylated DNA; B, blank control. Values are means ± SEM. ; ; . All experiments were repeated at least thrice.

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