The role of mouse Akt2 in insulin-dependent suppression of adipocyte lipolysis in vivo - PubMed (original) (raw)

The role of mouse Akt2 in insulin-dependent suppression of adipocyte lipolysis in vivo

Shlomit Koren et al. Diabetologia. 2015 May.

Abstract

Aim/hypothesis: The release of fatty acids from adipocytes, i.e. lipolysis, is maintained under tight control, primarily by the opposing actions of catecholamines and insulin. A widely accepted model is that insulin antagonises catecholamine-dependent lipolysis through phosphorylation and activation of cAMP phosphodiesterase 3B (PDE3B) by the serine-threonine protein kinase Akt (protein kinase B). Recently, this hypothesis has been challenged, as in cultured adipocytes insulin appears, under some conditions, to suppress lipolysis independently of Akt.

Methods: To address the requirement for Akt2, the predominant isoform expressed in classic insulin target tissues, in the suppression of fatty acid release in vivo, we assessed lipolysis in mice lacking Akt2.

Results: In the fed state and following an oral glucose challenge, Akt2 null mice were glucose intolerant and hyperinsulinaemic, but nonetheless exhibited normal serum NEFA and glycerol levels, suggestive of normal suppression of lipolysis. Furthermore, insulin partially inhibited lipolysis in Akt2 null mice during an insulin tolerance test (ITT) and hyperinsulinaemic-euglycaemic clamp, respectively. In support of these in vivo observations, insulin antagonised catecholamine-induced lipolysis in primary brown fat adipocytes from Akt2-deficient mice.

Conclusions/interpretation: These data suggest that suppression of lipolysis by insulin in hyperinsulinaemic states can take place in the absence of Akt2.

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Conflict of interest statement

Duality of interest

The authors declare that there is no duality of interest associated with this manuscript.

Figures

Fig. 1

Fig. 1

Akt2 null mice have mildly reduced adiposity and equivalent levels of adipokines to wild-type mice. All experiments were done in randomly fed, 10–12-week old, wild-type (black bars) or Akt2 null (white bars) male mice. (a) Weight in grams, _n_=19 for both genotypes. (b) Fat/lean mass % as measured by NMR, _n_=5 for both genotypes. (c) Inguinal fat and (d) epididymal fat pad weights in grams, _n_=5 for both genotypes. (e) Serum leptin, (f) adiponectin, (g) resistin and (h) TNF-α levels, _n_=19 for both genotypes. Data are expressed as means ± SEM. *p<0.05, as indicated. WT, wild-type

Fig. 2

Fig. 2

Altered glucose homeostasis with normal suppression of lipolysis in refed Akt2 null mice. Wild-type (black bars) or Akt2 null (white bars) male mice, 10–12 weeks old, were fasted overnight (16 h) and refed for 2 h. Serum levels of (a) glucose (_n_=20), (b) insulin (_n_=20), (c) glycerol (_n_=20), (d) NEFA (_n_=10), (e) TG (_n_=9–10) and (f) ketones (_n_=9–10) were measured and expressed as means ± SEM. *p<0.05, **p<0.01, as indicated. WT, wild-type

Fig. 3

Fig. 3

Akt2 null mice are glucose intolerant but have normal suppression of lipolysis. Wild-type (black lines/black bars [_n_=20]) or Akt2 null (dotted lines/white bars [_n_=14]), male mice, 10–12 weeks old, were fasted overnight (16 h). D-glucose (2 g/kg) was administered orally to conscious mice. (a) Blood glucose levels were sampled at the indicated times. (b) AUC of the glucose response. Serum levels of (c) insulin, (d) glycerol and (e) NEFA were sampled at baseline and at 15 min. Data are expressed as means ± SEM. *p<0.05, **p<0.01. WT, wild-type

Fig. 4

Fig. 4

Insulin partially inhibits lipolysis in Akt2 null mice after i.p. insulin injection. Wild-type (black line/black bars [_n_=10]) or Akt2 null (dotted line/white bars [_n_=10]) male mice, 10–12 weeks old, were fasted overnight (16 h). Insulin (1 U/kg) was administered by i.p. injection to conscious mice. (a) Blood glucose levels were sampled at the indicated times. (b) AUC of the glucose response. Serum levels of (c) insulin (d) glycerol and (e) NEFA were sampled before the injection and at 30 min. Data are expressed as means ± SEM. **p<0.01, as indicated. WT, wild-type

Fig. 5

Fig. 5

Akt2 null mice show selective insulin resistance by euglycaemic clamp. Wild-type (black lines/black bars [_n_=6]) or Akt2 null (dotted lines/white bars [_n_=8]), male mice, 10–12 weeks old, were fasted overnight (16 h). A hyperinsulinaemic–euglycaemic clamp procedure was conducted in conscious, unanaesthetised mice with a primed-continuous infusion of insulin (5 mU kg−1 min−1) and a variable infusion of 50% glucose to maintain euglycaemia. (a) Glucose levels during the clamp, (b) glucose infusion rate during the clamp, (c) insulin, (d) glycerol and (e) NEFA levels at baseline and at the end of the clamp were measured. Data are expressed as means ± SEM. *p<0.05, **p<0.01, as indicated. GIR, glucose infusion rate; WT, wild-type

Fig. 6

Fig. 6

Insulin inhibits isoprenaline-induced lipolysis in primary brown adipocytes of Akt2 null mice. Primary brown adipocytes were isolated from either wild-type or Akt2 null pups. The cells were treated with 1 nmol/l isoprenaline (Iso) ± 100 nmol/l insulin (INS). (a) Glycerol release assay and (b) NEFA release assay were performed. Data are presented as % of maximal response of wild-type cells to 1 nmol/l isoprenaline. (c) 2-Deoxyglucose uptake assay was performed with no treatment and with 100 nmol/l insulin. Insulin dose-response curves of (d) glycerol and (e) NEFA release were generated using 1 nmol/l isoprenaline stimulation. (f) Glycerol release assay in the presence of 1 nmol/l isoprenaline ± 100 nmol/l insulin ± 20 μmol/l AktiVIII (Akti) ±10 μmol/l PI-103 (PI3Ki). Data are expressed as means ± SEM from 2–4 experiments. *p<0.05, **p<0.01, as indicated. No Tx, no treatment, WT, wild-type

Fig. 7

Fig. 7

Protein levels of the different Akt isoenzymes. Primary brown adipocytes were lysed and lysates were immunoblotted for Akt1, Akt2, Akt3 and tubulin. The blot was developed using the Licor Odyssey system (Lincoln, NE, USA).

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