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. 2010 Dec;59(12):3032-40.
doi: 10.2337/db10-0074. Epub 2010 Sep 28.

Disruption of hepatic leptin signaling protects mice from age- and diet-related glucose intolerance

Frank K Huynh  1 , Jasna Levi, Heather C Denroche, Sarah L Gray, Peter J Voshol, Ursula H Neumann, Madeleine Speck, Streamson C Chua, Scott D Covey, Timothy J Kieffer
Affiliations

Disruption of hepatic leptin signaling protects mice from age- and diet-related glucose intolerance

Frank K Huynh et al. Diabetes. 2010 Dec.

Abstract

Objective: The liver plays a critical role in integrating and controlling glucose metabolism. Thus, it is important that the liver receive and react to signals from other tissues regarding the nutrient status of the body. Leptin, which is produced and secreted from adipose tissue, is a hormone that relays information regarding the status of adipose depots to other parts of the body. Leptin has a profound influence on glucose metabolism, so we sought to determine if leptin may exert this effect in part through the liver.

Research design and methods: To explore this possibility, we created mice that have disrupted hepatic leptin signaling using a Cre-lox approach and then investigated aspects of glucose metabolism in these animals.

Results: The loss of hepatic leptin signaling did not alter body weight, body composition, or blood glucose levels in the mild fasting or random-fed state. However, mice with ablated hepatic leptin signaling had increased lipid accumulation in the liver. Further, as male mice aged or were fed a high-fat diet, the loss of hepatic leptin signaling protected the mice from glucose intolerance. Moreover, the mice displayed increased liver insulin sensitivity and a trend toward enhanced glucose-stimulated plasma insulin levels. Consistent with increased insulin sensitivity, mice with ablated hepatic leptin signaling had increased insulin-stimulated phosphorylation of Akt in the liver.

Conclusions: These data reveal that unlike a complete deficiency of leptin action, which results in impaired glucose homeostasis, disruption of leptin action in the liver alone increases hepatic insulin sensitivity and protects against age- and diet-related glucose intolerance. Thus, leptin appears to act as a negative regulator of insulin action in the liver.

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Figures

FIG. 1.
FIG. 1.
Leprflox/flox Albcre tg+ mice have a liver-specific loss of the leptin receptor signaling domain. A: Genomic DNA from tissues of Leprflox/flox mice with and without the Albcre transgene was used as template for PCR of the Leprflox allele. The predicted product sizes are 1,369 bp for Leprflox and 952 bp for LeprΔ17. B: RNA was extracted from the livers of Leprflox/flox mice and used as template for RT-PCR with primers flanking exon 17. The predicted product sizes are 343 bp for Leprflox and 267 bp for LeprΔ17 transcripts. Arrows to the left mark the migration of molecular weight markers in bp.
FIG. 2.
FIG. 2.
Attenuation of hepatic leptin signaling does not alter body composition. A and C: Leprflox/flox Albcre tg+ mice and littermate controls were assessed for body weight as well as (B and D) body composition as measured by NMR at 6 and 16 weeks old. At 6 weeks old, n ≥ 7, and at 16 weeks old, n ≥ 5, for Leprflox/flox Albcre tg+ and littermate controls. E and F: Twenty-one-week-old male mice were fasted for 4 h and then the liver was harvested. Lipids were isolated by a chloroform:methanol extraction and reconstituted into Thesit micelles. Samples were then assayed for triglycerides and cholesterol, n ≥ 8 in each group. Data are mean ± SEM.
FIG. 3.
FIG. 3.
Loss of hepatic leptin signaling prevents age- and diet-related glucose intolerance. Oral glucose tolerance tests were performed on male (A–E) Leprflox/flox Albcre tg+ mice and littermate controls or (F) db/db mice and C57BL/6 controls at the indicated ages. E: Mice were fed a high-fat diet (HFD) for 20 weeks, fasted for 4 h, and gavaged with 1.22 mg/g glucose. Data are mean ± SEM and n ≥ 6. *P ≤ 0.05 vs. wild-type control.
FIG. 4.
FIG. 4.
Attenuation of hepatic leptin signaling increases glucose-stimulated insulin levels. A: Plasma insulin levels were monitored following a gavage of 1.5-mg/g body weight glucose to assess steady-state levels of glucose-stimulated insulin secretion in 16- to 20-week-old male Leprflox/flox Albcre tg+ and tg− mice, n ≥ 13 in each group. B: Insulin-positive area as a function of total pancreas area and (C) total pancreatic β-cell mass, n ≥ 5 mice per genotype and 3 sections measured per pancreas from 22-week-old male mice. All data are mean ± SEM.
FIG. 5.
FIG. 5.
Ablation of hepatic leptin signaling increases insulin sensitivity. Insulin tolerance tests were performed on (A) 19-week-old females (n ≥ 4) and (B) 21-week-old males (n ≥ 6) with and without the Albcre transgene. All data are mean ± SEM. *P ≤ 0.05 vs. littermate controls.
FIG. 6.
FIG. 6.
Loss of hepatic leptin signaling enhances liver insulin sensitivity. Male Leprflox/flox Albcre tg+ mice and littermate controls (16 to 20 weeks old) were used in a hyperinsulinemic-euglycemic clamp as described in Research Design and Methods, n ≥ 6. A and B: Plasma insulin and blood glucose levels during basal and hyperinsulinemic states. C: Whole-body glucose utilization (BGU), endogenous glucose production (EGP), and glucose infusion rate (GIR). Data are mean ± SEM.
FIG. 7.
FIG. 7.
Attenuation of hepatic leptin signaling results in increased insulin-stimulated phosphorylation of Akt in the liver. Following the hyperinsulinemic-euglycemic clamp, liver tissues were harvested and flash-frozen. Liver lysates were prepared and Western blots performed for phosphorylated and total Akt levels. Representative blots from two Leprflox/flox Albcre tg− and two Leprflox/flox Albcre tg+ mice are shown in A. Quantification of all samples by densitometry is shown in B, n ≥ 8. Data are mean ± SEM. p-Akt, phosphorylated Akt.
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