Extended Data Figure 6. Effect of DPP4 silencing on insulin-induced p-AKT in primary hepatocytes, on NEFA in obese or lean mice and on metabolism in comparison with sitagliptin in obese mice.

a, Wild-type or DPP4-silenced primary hepatocytes were treated with or without 50 µM palmitate for 10 h with the last 5 h in serum-free medium, and then stimulated with 100 nM of insulin for 5 min. p-AKT and total AKT and DPP4 were assayed by immunoblot. The data are representative of two independent experiments. b, Plasma samples from the following mice were assayed for non-esterified fatty acids (NEFA) four weeks after intravenous injection with AAV8-H1-shDpp4 (H-shDpp4) or control AAV8-H1 vector (con): 16-week-old mice fed the DIO diet for the last 10 weeks, 5-weekold chow-fed ob/ob mice, and 6-week-old chow-fed wild-type lean mice. n = 5–6 mice per group; mean ± s.e.m.; *P < 0.05 and n.s., non-significant by two-tailed Student's t-test. c–h, After 10 weeks on high-fat diet, control DIO mice, H-shDpp4 DIO mice and DIO mice were administered sitagliptin (sita) in drinking water to achieve a dose of ~30–45 mg/kg/ day. After 4 weeks of treatment, the mice were analysed as follows. c, Body weight. d, Plasma and VAT DPP4 activity. e, Hepatic DPP4 immunoblot and hepatocyte DPP4 activity. f, Blood glucose and plasma insulin 5 h after food withdrawal. g, p-AKT and total AKT in VAT and liver after portal vein insulin injection. h, Plasma non-esterified fatty acids (NEFA). For all quantified data panels except e, n = 6–8 mice per group; for e, n = 4 mice per group; mean ± s.e.m.; *,^#P < 0.05 and n.s., non-significant by one-way ANOVA. For gel source data, see Supplementary Fig. 1. i, DIO mice were treated with AAV8-H1-con, AAV8-H1-shDpp4 or sitagliptin (+ AAV8- H1-con) exactly as above, except the mice were analysed after 11 weeks of treatment instead of after 4 weeks. n = 5 mice per group; mean ± s.e.m.; *,^# P < 0.05 and n.s., non-significant by one-way ANOVA.