Figure 3: Inhibition of p53 in β-cells improves glucose tolerance and insulin secretion in β-MDM2KO mice.
(a) Pancreatic islets were isolated from C57 mice intraperitoneally injected with AAV encoding FLAG-tagged dominant-negative form of p53 (AAV-DN-p53) or GFP control (AAV-GFP) at an indicated amount of genomic copy (gc) number per mouse for 2 weeks. The islets were subjected to immunoblotting using an antibody against MDM2, FLAG, p53, GFP or β-actin as indicated. The right panel is the densitometric analysis for the relative abundance of MDM2 and p53 normalized with β-actin (n=4). (b) IHC staining with an anti-FLAG antibody in pancreatic sections of C57 mice injected with AAV-GFP or AAV-DN-p53 (1 × 1012 gc per mouse) with original magnification at × 400. Scale bar, 50 μm. (c–d) Eight-week-old male β-MDM2KO mice and its WT littermates were injected with AAV-GFP or AAV-DN-p53 at a dose of 1 × 1012 gc per mouse for 2 weeks. (c) GTT. (d) Serum insulin levels during GTT in c. (e–g) Isolated islets from β-MDM2KO mice or its WT littermates were infected with adenovirus encoding luciferase (Adv-Luci) or FLAG-tagged DN-p53 (Adv-DN-p53) (e,f) or treated with PFTα (10 μM) or DMSO as vehicle control (g) for 24 h. (e) The infected islets were subjected to immunoblotting with an anti-p53, -FLAG or -β-actin antibody. (f,g) Static GSIS in the islets were measured at 30 min after glucose stimulation (20 mM) (n=5). Note that insulin content in the isolated islets was similar between different treatment groups. *P<0.05 (KO+AAV-GFP versus WT+AAV-GFP, n=6), #P<0.05 (KO+AAV-GFP versus KO+AAV-DN-p53, n=6), @P<0.05 (Student’s t-test). All experiments were repeated at least three times and representative immunoblots and IHC images are shown. All data are represented as the mean±s.e.m.
