Glucagon-Like Peptide-1 Gene Therapy in Obese Diabetic Mice Results in Long-Term Cure of Diabetes by Improving Insulin Sensitivity and Reducing Hepatic Gluconeogenesis

Serum triglyceride and FFA levels in rAd-GLP-1–treated ob/ob mice. Diabetic ob/ob mice were treated with rAd-GLP-1 (n = 3) or rAd-βgal (n = 4). Two weeks later, mice were not fed for 4 h. Serum triglyceride (A) and serum FFA (B) levels were measured. Untreated diabetic ob/ob, wild-type, lean, and pair-fed diabetic ob/ob mice (n = 3–5 per group) served as controls. Data are means ± SD. *P < 0.05 compared with rAd-βgal–treated mice.

Glucose tolerance tests in rAd-GLP-1–treated ob/ob mice. Diabetic ob/ob mice were treated with rAd-GLP-1 (n = 6) or rAd-βgal (n = 5). Two weeks later, mice were not fed for 4 h and injected with glucose, and blood glucose levels were measured. Untreated diabetic ob/ob mice, wild-type lean mice, and pair-fed diabetic ob/ob mice (n = 3–8 per group) served as controls. Data are means ± SD. *P < 0.05; **P < 0.01 compared with rAd-βgal–treated mice. □, untreated; ▴, rAd-βgal; •, rAd-GLP-1; ◊, wild type; ▵, pair fed.

rAd-GLP-1 treatment improved β-cell function in diabetic ob/ob mice. Diabetic ob/ob mice were treated with rAd-GLP-1 or rAd-βgal. Two weeks later, β-cell mass (A) and insulin content (n = 4–6 per group) (B) were measured. C: Mice were not fed for 4 h, and the expression of insulin mRNA in islets was determined by RT-PCR. Untreated diabetic ob/ob mice served as controls. Expression of GAPDH served as an internal control. A gel picture representative of three different experiments is shown. D: Serum insulin levels were measured at 0 and 30 min after glucose injection (2 g/kg body wt i.p.) by enzyme immunosorbent assay (n = 3–6 per group). □, 0 min; ▪, 30 min. Data are means ± SD. *P < 0.05; ** P < 0.01 compared with rAd-βgal–treated mice. ***P < 0.01 compared with insulin secretion at 0 min.

rAd-GLP-1 treatment improved insulin sensitivity in diabetic ob/ob mice. Diabetic ob/ob mice were treated with rAd-GLP-1 or rAd-βgal. A: Two weeks later, insulin (2 units/kg body wt) was injected and blood glucose levels were measured (n = 4–7 per group). Data are expressed as a percentage of the initial blood glucose level before insulin injection. □, untreated; ▴, rAd-βgal; ▵, pair fed; •, rAd-GLP-1; ⋄, wild type. B: Four weeks later, adipocytes were removed and incubated in the presence of insulin and the uptake of 2-deoxy-d-[1-³H]glucose was measured. Data are expressed as the percentage change from 2-deoxy-d-[1-³H]glucose uptake in the absence of insulin (n = 5–9 mice per group). Untreated diabetic ob/ob mice, wild-type lean mice, and pair-fed diabetic ob/ob mice served as controls. Data are means ± SD. *P < 0.05; ** P < 0.01 compared with rAd-βgal–treated mice.

Expression and phosphorylation of IRS-1 and activity of Akt and PKC in the muscle of rAd-GLP-1–treated diabetic ob/ob mice. Diabetic ob/ob mice were treated with rAd-GLP-1 or rAd-βgal. Four weeks later, mice were treated without (−) or with (+) insulin, and muscle tissue was sampled 15 min later. A: Tissue lysates were immunoprecipitated (IP) with anti–IRS-1 antibody and immunoblotted (IB) with anti–IRS-1 (IRS-1) or anti-phophotyrosine (PY) antibody. Representative immunoblots are shown (upper panel). IRS-1 protein in mice without insulin treatment is presented as a ratio of IRS-1 in wild-type lean mice (lower left panel). Phosphorylated IRS-1 was normalized with total IRS-1 protein and presented as a ratio of the phosphorylated IRS-1 level in insulin-stimulated wild-type mice (lower right panel). B: Tissue extracts were immunoblotted (IB) with anti-Akt (AKT) or anti–phospho-Akt (P-AKT) antibody. Representative immunoblots are shown (upper panel). Phosphorylated Akt was normalized with total Akt protein and presented as a ratio of the phosphorylated Akt level in insulin-stimulated wild-type mice (lower panel). C: PKC activity was determined. Untreated diabetic ob/ob mice and wild-type lean mice served as controls. Data are means ± SD. n = 3–7 per group. *P < 0.05 compared with rAd-βgal–treated mice. □, basal; ▪, insulin.

Expression and phosphorylation of IRS-1 and activity of Akt and PKC in the liver of rAd-GLP-1–treated diabetic ob/ob mice. Diabetic ob/ob mice were treated with rAd-GLP-1 or rAd-βgal. Four weeks later, mice were treated without or with insulin and liver tissue was sampled 15 min later. A–C: As for Fig. 6. Data are means ± SD. n = 3–7 per group. *P < 0.05; **P < 0.01 compared with rAd-βgal–treated mice. □, basal; ▪, insulin.