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. 2005 Dec 27;4(2):e31. doi: 10.1371/journal.pbio.0040031

Figure 4. UCP2 is Up-Regulated in Sirt1 Knockdown Cells and in Sirt1 KO Mice.

Figure 4

(A) Northern blot for the insulin gene INS-1 in cells with a control vector (pSUPER) or a SiRNA-Sirt1 vector (SiRNA Sirt1).

(B and C) Western blot analyses of targets involved in insulin synthesis and secretion using specific antibodies: cEBP/β, insulin receptor α and β, and kir6.2, one of the K+ channel receptor subunits.

(D) Measurement of ATP/ADP levels in INS-1 control cells (open bars) or Sirt1 knockdown cells (black bars) treated with 16.7 mM glucose (+) or 4 mM glucose (−) (n = 3 experiments done in triplicate, *p < 0.005 in the pSUPER experiment; ANOVA).

(E) Western blot analysis for UCP2 in INS-1 control cells (pSUPER) or knockdown cells (SiRNA Sirt1).

(F) Northern blot analysis for UCP2 in INS-1 control cells (pSUPER) or knockdown cells (SiRNA Sirt1).

(G) NADH levels in INS-1 cells after glucose addition as determined by autofluorescence [46] and expressed as arbitrary units. Cells stably transfected with control or Sirt1 SiRNA vectors were used (n = 2, *p < 0.05 compared with no glucose).

(H) UCP2 protein levels in isolated pancreatic islets of two wild-type or two Sirt1 KO mice. Tubulin or actin was used as loading control in all Western and Northern blots.