Protein kinase B (PKB) is known to mediate a number of biological responses to insulin and growth factors, its role in glucose uptake being one of the most extensively studied. In this work, we have employed a recently described allosteric inhibitor of PKB, Akti, to clarify the role of PKB in lipid metabolism in adipocytes-a subject that has received less attention. Pretreatment of primary rat and 3T3L1 adipocytes with Akti resulted in dose-dependent inhibition of PKB phosphorylation and activation in response to insulin, without affecting upstream insulin signaling [insulin receptor (IR), insulin receptor substrate (IRS)] or the insulin-induced phosphoinositide 3-kinase (PI3K)-dependent activation of the ERK/p90 ribosomal kinase (RSK) pathway. PKB activity was required for the insulin-induced activation of phosphodiesterase 3B (PDE3B) and for the antilipolytic action of insulin. Moreover, inhibition of PKB activity resulted in a reduction in de novo lipid synthesis and in the ability of insulin to stimulate this process. The regulation of the rate-limiting lipogenic enzyme acetyl-CoA carboxylase (ACC) by insulin through dephosphorylation of S79, which is a target for AMP-activated protein kinase (AMPK), was dependent on the presence of active PKB. Finally, AMPK was shown to be phosphorylated by PKB on S485 in response to insulin, and this was associated with a reduction in AMPK activity. In summary, we propose that PKB is required for the positive effects of insulin on lipid storage and that regulation of PDE3B and AMPK by PKB is important for these effects.

Graphical abstract

AMP-activated protein kinase (AMPK) is an important regulator of cellular energy status. In adipocytes, stimuli that increase intracellular cyclic AMP (cAMP) have also been shown to increase the activity of AMPK. The precise molecular mechanisms responsible for cAMP-induced AMPK activation are not clear. Phosphodiesterase 3B (PDE3B) is a critical regulator of cAMP signalling in adipocytes. Here we investigated the roles of PDE3B, PDE4, protein kinase B (PKB) and the exchange protein activated by cAMP 1 (Epac1), as well as lipolysis, in the regulation of AMPK in primary rat adipocytes. We demonstrate that the increase in phosphorylation of AMPK at T172 induced by the adrenergic agonist isoproterenol can be diminished by co-incubation with insulin. The diminishing effect of insulin on AMPK activation was reversed upon treatment with the PDE3B specific inhibitor OPC3911 but not with the PDE4 inhibitor Rolipram. Adenovirus-mediated overexpression of PDE3B and constitutively active PKB both resulted in greatly reduced isoproterenol-induced phosphorylation of AMPK at T172. Co-incubation of adipocytes with isoproterenol and the PKA inhibitor H89 resulted in a total ablation of lipolysis and a reduction in AMPK phosphorylation/activation. Stimulation of adipocytes with the Epac1 agonist 8-pCPT-2’O-Me-cAMP led to increased phosphorylation of AMPK at T172. The general lipase inhibitor Orlistat decreased isoproterenol-induced phosphorylation of AMPK at T172. This decrease corresponded to a reduction of lipolysis from adipocytes. Taken together, these data suggest that PDE3B and PDE4 regulate cAMP pools that affect the activation/phosphorylation state of AMPK and that the effects of cyclic AMP on AMPK involve Epac1, PKA and lipolysis.

The incidence of obesity in the developed world is increasing at an alarming rate. Concurrent with the increase in the incidence of obesity is an increase in the incidence of type 2 diabetes. Cyclic AMP (cAMP) and cGMP are key second messengers in all cells; for example, when it comes to processes of relevance for the regulation of energy metabolism, cAMP is a key mediator in the regulation of lipolysis, glycogenolysis, gluconeogenesis and pancreatic β cell insulin secretion. PDE3B, one of several enzymes which hydrolyze cAMP and cGMP, is expressed in cells of importance for the regulation of energy homeostasis, including adipocytes, hepatocytes, hypothalamic cells, and β cells. It has been shown, using PDE3 inhibitors and gene targeting approaches in cells and animals, that altered levels of PDE3B result in a number of changes in the regulation of glucose and lipid metabolism and in overall energy homeostasis. This article highlights the complexity involved in the regulation of PDE3B by hormones, and in the regulation of downstream metabolic effects by PDE3B in several interacting tissues.

Aims/hypothesis Inhibition of the enzyme dipeptidyl peptidase 4 (DPP-4), which cleaves and inactivates glucagon-like peptide 1 (GLP-1), is a glucose-lowering strategy in type 2 diabetes. Since DPP-4 is a ubiquitously distributed enzyme, we examined whether it is expressed in islets and whether an islet effect to inhibit DPP-4 may result in stimulated insulin secretion. Methods We investigated DPP-4 expression and activity in the islets of mouse models of obesity as well as human islets from non-diabetic and type 2 diabetic donors. We further investigated whether inhibition with DPP-4 inhibitors could promote insulin secretion via islet GLP-1 in isolated islets.