Yannick Le Marchand-Brustel

The stomach is a source of leptin

The circulating peptide leptin, which is the product of the ob gene, provides feedback information on the size of fat stores to central Ob receptors, that control food intake and body-weight homeostasis. Leptin has so far been reported to be secreted only by adipocytes and the placenta. Here we show that leptin messenger RNA and leptin protein are present in rat gastric epithelium, and that cells in the glands of the gastric fundic mucosa are immunoreactive for leptin. The physiological function of this previously unsuspected source of leptin is unknown. However, both feeding and administration of CCK-8 (the biologically active carboxy-terminal end of cholecystokinin) result in a rapid and large decrease in both leptin cell immunoreactivity and the leptin content of the fundic epithelium, with a concomitant increase in the concentration of leptin in the plasma. These results indicate that gastric leptin may be involved in early CCK-mediated effects activated by food intake, possibly including satiety.

Hypoinsulinaemia, glucose intolerance and diminished beta-cell size in S6K1-deficient mice

Insulin controls glucose homeostasis by regulating glucose use in peripheral tissues, and its own production and secretion in pancreatic β cells. These responses are largely mediated downstream of the insulin receptor substrates, IRS-1 and IRS-2 (refs 4,5,6,7,8), through distinct signalling pathways. Although a number of effectors of these pathways have been identified, their roles in mediating glucose homeostasis are poorly defined. Here we show that mice deficient for S6 kinase 1, an effector of the phosphatidylinositide-3-OH kinase signalling pathway, are hypoinsulinaemic and glucose intolerant. Whereas insulin resistance is not observed in isolated muscle, such mice exhibit a sharp reduction in glucose-induced insulin secretion and in pancreatic insulin content. This is not due to a lesion in glucose sensing or insulin production, but to a reduction in pancreatic endocrine mass, which is accounted for by a selective decrease in β-cell size. The observed phenotype closely parallels those of preclinical type 2 diabetes mellitus, in which malnutrition-induced hypoinsulinaemia predisposes individuals to glucose intolerance.

Metformin, independent of AMPK, induces mTOR inhibition and cell-cycle arrest through REDD1

Metformin is a widely prescribed antidiabetic drug associated with a reduced risk of cancer. Many studies show that metformin inhibits cancer cell viability through the inhibition of mTOR. We recently showed that antiproliferative action of metformin in prostate cancer cell lines is not mediated by AMP-activated protein kinase (AMPK). We identified REDD1 (also known as DDIT4 and RTP801), a negative regulator of mTOR, as a new molecular target of metformin. We show that metformin increases REDD1 expression in a p53-dependent manner. REDD1 invalidation, using siRNA or REDD1(-/-) cells, abrogates metformin inhibition of mTOR. Importantly, inhibition of REDD1 reverses metformin-induced cell-cycle arrest and significantly protects from the deleterious effects of metformin on cell transformation. Finally, we show the contribution of p53 in mediating metformin action in prostate cancer cells. These results highlight the p53/REDD1 axis as a new molecular target in anticancer therapy in response to metformin treatment.

The lipid phosphatase SHIP2 controls insulin sensitivity.

Insulin is the primary hormone involved in glucose homeostasis, and impairment of insulin action and/or secretion has a critical role in the pathogenesis of diabetes mellitus. Type-II SH2-domain-containing inositol 5-phosphatase, or ‘SHIP2’, is a member of the inositol polyphosphate 5-phosphatase family. In vitro studies have shown that SHIP2, in response to stimulation by numerous growth factors and insulin, is closely linked to signalling events mediated by both phosphoinositide-3-OH kinase and Ras/mitogen-activated protein kinase. Here we report the generation of mice lacking the SHIP2 gene. Loss of SHIP2 leads to increased sensitivity to insulin, which is characterized by severe neonatal hypoglycaemia, deregulated expression of the genes involved in gluconeogenesis, and perinatal death. Adult mice that are heterozygous for the SHIP2 mutation have increased glucose tolerance and insulin sensitivity associated with an increased recruitment of the GLUT4 glucose transporter and increased glycogen synthesis in skeletal muscles. Our results show that SHIP2 is a potent negative regulator of insulin signalling and insulin sensitivity in vivo.

Metformin in Cancer Therapy: A New Perspective for an Old Antidiabetic Drug?

Metformin is the most widely used antidiabetic drug in the world, and there is increasing evidence of a potential efficacy of this agent as an anticancer drug. First, epidemiological studies show a decrease in cancer incidence in metformin-treated patients. Second, metformin decreases insulin resistance and indirectly reduces insulin level, a beneficial effect because insulin promotes cancer cell growth. Third, several reports outline a direct inhibitory effect of metformin on cancer cell growth and an antitumoral action. Finally, metformin activates the AMP activated protein kinase (AMPK) pathway, a major sensor of the energetic status of the cell, which has been proposed as a promising therapeutic target in cancer. Mol Cancer Ther; 9(5); 1092–99. ©2010 AACR.

Targeting Cancer Cell Metabolism: The Combination of Metformin and 2-Deoxyglucose Induces p53-Dependent Apoptosis in Prostate Cancer Cells

Targeting cancer cell metabolism is a new promising strategy to fight cancer. Metformin, a widely used antidiabetic agent, exerts antitumoral and antiproliferative action. In this study, the addition of metformin to 2-deoxyglucose (2DG) inhibited mitochondrial respiration and glycolysis in prostate cancer cells leading to a severe depletion in ATP. The combination of the two drugs was much more harmful for cancer cells than the treatment with metformin or 2DG alone, leading to 96% inhibition of cell viability in LNCaP prostate cancer cells. In contrast, a moderate effect on cell viability was observed in normal prostate epithelial cells. At the cellular level, the combination of metformin and 2DG induced p53-dependent apoptosis via the energy sensor pathway AMP kinase, and the reexpression of a functional p53 in p53-deficient prostate cancer cells restored caspase-3 activity. In addition to apoptosis, the combination of metformin and 2DG arrested prostate cancer cells in G(2)-M. This G(2)-M arrest was independent of p53 and correlated with a stronger decrease in cell viability than obtained with either drug. Finally, metformin inhibited 2DG-induced autophagy, decreased beclin 1 expression, and triggered a switch from a survival process to cell death. Our study reinforces the growing interest of metabolic perturbators in cancer therapy and highlights the potential use of the combination of metformin and 2DG as an anticancerous treatment.

Potential role of protein kinase B in glucose transporter 4 translocation in adipocytes.

Phosphatidylinositol 3-kinase (PI 3-kinase) activation promotes glucose transporter 4 (Glut 4) translocation in adipocytes. In this study, we demonstrate that protein kinase B, a serine/threonine kinase stimulated by PI 3-kinase, is activated by both insulin and okadaic acid in isolated adipocytes, in parallel with their effects on Glut 4 translocation. In 3T3-L1 adipocytes, platelet-derived growth factor activated PI 3-kinase as efficiently as insulin but was only half as potent as insulin in promoting protein kinase B (PKB) activation. To look for a potential role of PKB in Glut 4 translocation, adipocytes were transfected with a constitutively active PKB (Gag-PKB) together with an epitope tagged transporter (Glut 4 myc). Gag-PKB was associated with all membrane fractions, whereas the endogenous PKB was mostly cytosolic. Expression of Gag-PKB led to an increase in Glut 4 myc amount at the cell surface. Our results suggest that PKB could play a role in promoting Glut 4 appearance at the cell surface following exposure of adipocytes to insulin and okadaic acid stimulation.

Hypoxia decreases insulin signaling pathways in adipocytes

OBJECTIVE—Obesity is characterized by an overgrowth of adipose tissue that leads to the formation of hypoxic areas within this tissue. We investigated whether this phenomenon could be responsible for insulin resistance by studying the effect of hypoxia on the insulin signaling pathway in adipocytes. RESEARCH DESIGN AND METHODS—The hypoxic signaling pathway was modulated in adipocytes from human and murine origins through incubation under hypoxic conditions (1% O2) or modulation of hypoxia-inducible factor (HIF) expression. Insulin signaling was monitored through the phosphorylation state of several key partners of the pathway and glucose transport. RESULTS—In both human and murine adipocytes, hypoxia inhibits insulin signaling as revealed by a decrease in the phosphorylation of insulin receptor. In 3T3-L1 adipocytes, this inhibition of insulin receptor phosphorylation is followed by a decrease in the phosphorylation state of protein kinase B and AS160, as well as an inhibition of glucose transport in response to insulin. These processes were reversible under normoxic conditions. The mechanism of inhibition seems independent of protein tyrosine phosphatase activities. Overexpression of HIF-1α or -2α or activation of HIF transcription factor with CoCl2 mimicked the effect of hypoxia on insulin signaling, whereas downregulation of HIF-1α and -2α by small interfering RNA inhibited it. CONCLUSIONS—We have demonstrated that hypoxia creates a state of insulin resistance in adipocytes that is dependent upon HIF transcription factor expression. Hypoxia could be envisioned as a new mechanism that participates in insulin resistance in adipose tissue of obese patients.

Retinoic acid activation of the ERK pathway is required for embryonic stem cell commitment into the adipocyte lineage.

Mouse embryonic stem (ES) cells are pluripotent cells that differentiate into multiple cell lineages. The commitment of ES cells into the adipocyte lineage is dependent on an early 3-day treatment with all-trans retinoic acid (RA). To characterize the molecular mechanisms underlying this process, we examined the contribution of the extracellular-signal-regulated kinase (ERK) pathway. Treatment of ES cell-derived embryoid bodies with RA resulted in a prolonged activation of the ERK pathway, but not the c-Jun N-terminal kinase, p38 mitogen-activated protein kinase or phosphoinositide 3-kinase pathways. To investigate the role of ERK activation, co-treatment of RA with PD98059, a specific inhibitor of the ERK signalling pathway, prevented both adipocyte formation and expression of the adipogenic markers, adipocyte lipid-binding protein and peroxisome-proliferator-activated receptor gamma. Furthermore, we show that ERK activation is required only during RA treatment. PD98059 does not interfere with the commitment of ES cells into other lineages, such as neurogenesis, myogenesis and cardiomyogenesis. As opposed to the controversial role of the ERK pathway in terminal differentiation, our results clearly demonstrate that this pathway is specifically required at an early stage of adipogenesis, corresponding to the RA-dependent commitment of ES cells.

Overexpression of a Constitutively Active Form of Phosphatidylinositol 3-Kinase Is Sufficient to Promote Glut 4 Translocation in Adipocytes

Insulin stimulates glucose transport in its target cells by recruiting the glucose transporter Glut 4 from an intracellular compartment to the cell surface. Previous studies have indicated that phosphatidylinositol 3-kinase (PI 3-kinase) is a necessary step in this insulin action. We have investigated whether PI 3-kinase activation is sufficient to promote Glut 4 translocation in transiently transfected adipocytes. Rat adipose cells were cotransfected with expression vectors that allowed transient expression of epitope-tagged Glut 4 and a constitutively active form of PI 3-kinase (p110*). The expression of p110* induced the appearance of epitope-tagged Glut 4 at the cell surface at a level similar to that obtained after insulin treatment, whereas a kinase-dead version of p110* had no effect. The p110* effect was observed over a wide range of the transfected cDNA. When subcellular fractionation of adipocytes was performed, p110* was found, similar to the endogenous PI 3-kinase, enriched in the low density microsomal compartment, which also contains the Glut 4 vesicles. This could suggest that a specific localization of PI 3-kinase in this compartment is required for the action on Glut 4. The observations made with PI 3-kinase are in contrast with those seen with the MAP kinase cascade. Indeed, a constitutively active form of MAP kinase kinase had no effect on Glut 4 translocation in basal conditions. At the highest degree of expression, the constitutively active form of MAP kinase kinase slightly inhibited the insulin stimulation of Glut 4 translocation. Taken together, our results indicate that Glut 4 translocation can be efficiently promoted by an active form of PI 3-kinase but not by the activation of the MAP kinase pathway.