Figure 9: In vivo studies with transgenic mice overexpressing barr2 in pancreatic β-cells.

(a–c) In vivo metabolic tests carried out with RIPII-barr2 Tg mice (Tg) and wild-type (wt) littermates maintained on RC. (a) Glucose-induced insulin secretion (GSIS; 2 g glucose per kg i.p.). (b) IGTT (2 g glucose per kg i.p.). (c) ITT (0.75 U insulin per kg i.p.). (d–f) RIPII-barr2 Tg mice (Tg) and wt littermates were maintained on a HFD and subjected to the same tests as in a–c. In d and e, the glucose dose was reduced to 1 g kg⁻¹ (i.p.). In f, the insulin dose was 1 U kg⁻¹ (i.p.). Mice were maintained on the HFD for at least 8 weeks. All studies were carried out with male mice (mouse age: RC, 8–12 weeks; HFD, 16–20 weeks). Data are given as means±s.e.m. (6–9 mice per group); *P<0.05, **P<0.01, as compared with the corresponding wt value (Student’s t-test). (g) Barr2 expression is reduced in islets of mice maintained on a HFD. Total RNA was prepared from pancreatic islets of wt mice (16-week-old male C57BL/6NTac mice; eight mice per group). Mice were maintained on either RC or on a HFD (for 8 weeks). Gene expression levels were determined via real-time qRT-PCR. Transcript levels were normalized relative to the expression of β-actin (for primer sequences, see Methods). Data represent means±s.e.m. (***P<0.001; Student’s t-test). (h,i) Reduced β-arrestin expression in human islets cultured in glucose-rich medium (16 mM) in the presence of 0.5 mM palmitic acid (PA) or a 2:1 mixture of PA and oleic acid (PA+OA; total fatty acid concentration: 0.5 mM) for 3 days. Relative BARR2 (h) and BARR1 (i) expression levels were determined by qRT-PCR (internal control gene: β-ACTIN). BARR2 and BARR1 expression levels determined with islets that had not been exposed to PA or PA+OA were set equal to 100 in each individual experiment. Data are given as means±s.e.m. (n=3 per group; islets were prepared from six different donors). **P<0.01, Student’s t-test. (j,l) Impaired insulin secretion in human islets cultured in glucose-rich medium (16 mM) in the presence of 0.5 mM PA or a 2:1 mixture of PA and OA (total fatty acid concentration: 0.5 mM) for 3 days. Insulin release in response to 4 and 8 mM glucose (G4 and G8, respectively) was monitored using an islet perifusion system. Note that isolated human islets are far more sensitive to glucose than mouse or rat islets explaining why both G4 and G8 strongly promote insulin release in control islets. Data are given means±s.e.m. of three independent perifusion experiments. (k,m) Quantification of the data shown in (j,l). Insulin release following stimulation wth G4 or G8 is expressed as AUC. Data represent means±s.e.m. of three independent perifusion experiments (*P<0.05, ***P<0.001, Student’s t-test). AUC, area under the curve.