Naringenin inhibits phosphoinositide 3-kinase activity and glucose uptake in 3T3-L1 adipocytes

Previous studies have shown that flavonoids inhibit glucose uptake in cultured cells. In this report, we show that the grapefruit flavanone naringenin inhibited insulin-stimulated glucose uptake in 3T3-L1 adipocytes in a dose-dependent manner. Naringenin acts by inhibiting the activity of phosphoinositide 3-kinase (PI3K), a key regulator of insulin-induced GLUT4 translocation. Although naringenin did not alter the phosphotyrosine status of the insulin receptor, insulin receptor substrate proteins, or PI3K, it did inhibit the phosphorylation of the downstream signaling molecule Akt. In an in vitro kinase assay, naringenin inhibited PI3K activity. A physiologically attainable dose of 6 microM naringenin reduced insulin-stimulated glucose uptake by approximately 20%. This inhibitory effect remained 24h after the removal of naringenin from the culture medium. Collectively, our findings suggest that the regular consumption of naringenin in grapefruit may exacerbate insulin resistance in susceptible individuals via impaired glucose uptake in adipose tissue.     

A class II phosphoinositide 3-kinase plays an indispensable role in hepatitis C virus replication

Phosphoinositides function as fundamental signaling molecules and play roles in diverse cellular processes. Certain types of viruses may employ host cell phosphoinositide signaling systems to facilitate their replication cycles. Here we demonstrate that the β isoform of class II PI3K (PI3K-C2β) plays an indispensable role in hepatitis C virus (HCV) propagation in human hepatocellular carcinoma cells. Knockdown of PI3K-C2β abrogated HCV propagation in the cell. Using an HCV replicon system, we found that knockdown of PI3K-C2β substantially repressed the full-genome replication, while showing relatively small reductions in sub-genome replication, in which structural proteins including core protein were deleted. We also found that HCV core protein showed the binding activity towards D4-phosphorylated phosphoinositides and overlapped localization with phosphatidylinositol 3,4-bisphosphate in the cell. These results suggest that the phosphoinositide generated by PI3K-C2β plays an indispensable role in the HCV replication cycle through the binding to HCV core protein.     

CD2-associated protein/phosphoinositide 3-kinase signaling has a preventive role in angiotensin II-induced podocyte apoptosis

Angiotensin II (Ang II) works as a paracrine or autocrine cytokine agent to regulate renal functions and promotes podocytes dysfunction directly or indirectly, causing proteinuria. The glomerular slit diaphragm (SD) serves as a size-selective barrier and is linked to the actin-based cytoskeleton by adaptor proteins, including CD2-associated protein (CD2AP). Therefore, damages to CD2AP affect not only the function of the SD, but also directly disrupt the podocyte cytoskeleton, leading to proteinuria. In addition, CD2AP can facilitate the nephrin-induced phosphoinositide 3-kinase (PI3-K)/Akt signaling, which protects podocytes from apoptosis. Here we found that CD2AP staining was located diffusely but predominantly in the peripheral cytoplasm and CD2AP co-localized with nephrin in mouse podocytes; however, Ang II decreased CD2AP staining diffusely and induced a separation from concentrated nephrin. Ang II notably reduced CD2AP expression in time- and concentration-dependent manners, and this was significantly recovered by losartan. Ang II induced podocyte apoptosis in time- and concentration-dependent manners in TUNEL and FACS assays. LY294002, a PI3-K inhibitor, further reduced CD2AP expression and increased podocyte apoptosis, which was augmented by siRNA for CD2AP. Thus, Ang II induces the relocalization and reduction of CD2AP via AT1R, which would cause podocyte apoptosis by the suppression of CD2AP/PI3-K signaling.     

Class II phosphoinositide 3-kinase C2alpha: what we learned so far

More than fifteen years after the first identification of a class II isoform of phosphoinositide 3-kinase (PI3K) in Drosophila melanoǵaster this subfamily remains the most enigmatic among all PI3Ks. What are the functions of these enzymes? What are their mechanisms of activation? Which downstream effectors are specifically regulated by these isoforms? Are class I and class II PI3Ks redundant or do they control different intracellular processes? And, more important, do class II PI3Ks have a role in human diseases? The recent increased interest on class II PI3Ks has started providing some answers to these questions but still a lot needs to be done to completely uncover the contribution of these enzymes to physiological processes and possibly to pathological conditions. Here we will summarise the recent findings on the alpha isoform of mammalian class II PI3Ks (PI3K-C2α ) and we will discuss the potential involvement of this enzyme in human diseases.     

Discovery of new thienopyrimidine derivatives as potent and orally efficacious phosphoinositide 3-kinase inhibitors

A series of new thienopyrimidine derivatives has been discovered as potent PI3K inhibitors. The systematic SAR studies for these analogues are described. Among them, 8a and 9a exhibit nanomolar enzymatic potencies and sub-micromolar cellular anti-proliferative activities. 8a displays favorable pharmacokinetic profiles, while 9a easily undergoes deacetylation to yield a major metabolite 8a. Furthermore, 8a and 9a potently inhibit tumor growth in a dose-dependent manner in the NCI-H460 xenograft model with an acceptable safety profile.     

Involvement of the Src kinase Lyn in phospholipase C-gamma 2 phosphorylation and phosphatidylinositol 3-kinase activation in Epo signalling

We examined the role of the Src kinase Lyn in phospholipase C-gamma 2 (PLC-gamma 2) and phosphatidylinositol (PI) 3-kinase activation in erythropoietin (Epo)-stimulated FDC-P1 cells transfected with a wild type (WT) Epo-receptor (Epo-R). We showed that two inhibitors of Src kinases, PP1 and PP2, abolish both PLC-gamma 2 tyrosine phosphorylation and PI 3-kinase activity in WT Epo-R FDC-P1 cells. We also demonstrated that Epo-phosphorylated Lyn is associated with tyrosine phosphorylated PLC-gamma 2 and PI 3-kinase in WT Epo-R FDC-P1-stimulated cells. Moreover Epo-activated Lyn phosphorylates in vitro PLC-gamma 2 immunoprecipitated from unstimulated cells. Our results suggest that the Src kinase Lyn is involved in PLC-gamma 2 phosphorylation and PI 3-kinase activation induced by Epo.     

PI3-kinase promotes TRPV2 activity independently of channel translocation to the plasma membrane

Cellular or chemical activators for most transient receptor potential channels of the vanilloid subfamily (TRPV) have been identified in recent years. A remarkable exception to this is TRPV2, for which cellular events leading to channel activation are still a matter of debate. Diverse stimuli such as extreme heat or phosphatidylinositol-3 kinase (PI3-kinase) regulated membrane insertion have been shown to promote TRPV2 channel activity. However, some of these results have proved difficult to reproduce and may underlie different gating mechanisms depending on the cell type in which TRPV2 channels are expressed. Here, we show that expression of recombinant TRPV2 can induce cytotoxicity that is directly related to channel activity since it can be prevented by introducing a charge substitution in the pore-forming domain of the channel, or by reducing extracellular calcium. In stably transfected cells, TRPV2 expression results in an outwardly rectifying current that can be recorded at all potentials, and in an increase of resting intracellular calcium concentration that can be partly prevented by serum starvation. Using cytotoxicity as a read-out of channel activity and direct measurements of cell surface expression of TRPV2, we show that inhibition of the PI3-kinase decreases TRPV2 channel activity but does not affect the trafficking of the channel to the plasma membrane. It is concluded that PI3-kinase induces or modulates the activity of recombinant TRPV2 channels; in contrast to the previously proposed mechanism, activation of TRPV2 channels by PI3-kinase is not due to channel translocation to the plasma membrane.     

Cloning, expression, purification, and characterization of the human Class Ia phosphoinositide 3-kinase isoforms

The Class I phosphoinositide 3-kinases (PI3Ks) are lipid kinases that phosphorylate the 3-hydroxyl group of the inositol ring of phosphatidylinositides. Although closely related, experimental evidence suggests that the four Class I PI3Ks may be functionally distinct. To further study their unique biochemical properties, the three human Class Ia PI3K (alpha, beta, and delta) p110 catalytic domains were cloned and co-expressed with the p85alpha regulatory domain in Sf9 cells. None of the p110 subunits were successfully expressed in the absence of p85alpha. Successful expression and purification of each p85alpha/p110 protein required using an excess of the p110 vector over the p85 vector during co-infection of Sf9 cells. Proteins were purified as the p85alpha/p110 complex by nickel affinity chromatography through an N-terminal His-tag on the p110 subunit using an imidazole gradient. The purification yields were high using the optimized ratio of p85/p110 vector and small culture volumes, with 24mg/L cell culture media for p85alpha/p110alpha, 17.5mg/L for p85alpha/p110delta, and 3.5mg/L for p85alpha/p110beta. The identity of each purified isoform was confirmed by mass spectral analysis and immunoblotting. The activities of the three p85alpha/p110 proteins and the Class Ib p110gamma catalytic domain were investigated using phosphatidylinositol 4,5-bisphosphate (PIP2) as the substrate in a PIP2/phosphatidylserine (PS) liposome. All four enzymes exhibited reaction velocities that were dependent on the surface concentration of PIP2. The surface concentrations that gave maximal activity for each human isoform with 0.5mM PIP2 were 2.5mol% PIP2 for p110gamma, 7.5mol% for p85alpha/p110beta, and 10mol% PIP2 for p85alpha/p110alpha and p85alpha/p110delta. The specific activity of p85alpha/p110alpha was three to five times higher than that of the other human isoforms. These kinetic differences may contribute to the unique roles of these isoforms in cells.     

The α-isoform of class II phosphoinositide 3-kinase is more effectively activated by insulin receptors than IGF receptors, and activation requires receptor NPEY motifs

Little is known about the physiological role and mechanism of activation of class II phosphoinositide 3-kinases (PI3Ks), although it has been shown that the PI3K-C2alpha isoform is activated by insulin. Using chimaeric receptor constructs which can be activated independently of endogenous receptors in transfected cells, we found that PI3K-C2alpha activity was stimulated to a greater extent by insulin receptors than IGF receptors in 3T3-L1 adipocytes. Activation of PI3K-C2alpha required an intact NPEY motif in the receptor juxtamembrane domain. We conclude that PI3K-C2alpha is a candidate for participation in insulin-specific intracellular signalling.     

Constitutive diffuse activation of phosphoinositide 3-kinase at the plasma membrane by v-Src suppresses the chemotactic response to PDGF by abrogating the polarity of PDGF receptor signalling

Cancer cells depend on chemotaxis for invasion and frequently overexpress and/or activate Src. We previously reported that v-Src accelerates motility by promoting phosphoinositide 3-kinase (PI3-K) signalling but abrogates chemotaxis. We here addressed the mechanism of the loss of chemotactic response to platelet-derived growth factor (PDGF) gradients in fibroblasts harbouring a thermosensitive v-Src kinase. At non-permissive temperature, PDGF receptor (PDGFR) signalling, assessed by phosphoY(751)-specific antibodies (a docking site for PI3-K), was not detected without PDGF and showed a concentration-dependent PDGF response. Both immunolabeling of PI3-K (p110) and live cell imaging of its product (phosphatidylinositol 3,4,5 tris-phosphate) showed PI3-K recruitment and activation at lamellipodia polarized towards a PDGF gradient. Centrosomes and PDGFR- and Src-bearing endosomes were also oriented towards this gradient. Upon v-Src thermoactivation, (i) Y(751) phosphorylation was moderately induced without PDGF and synergistically increased with PDGF; (ii) PI3-K was recruited and activated all along the plasma membrane without PDGF and did not polarize in response to a PDGF gradient; and (iii) polarization of centrosomes and of PDGFR-bearing endosomes were also abrogated. Thus, PDGF can further increase PDGFR auto-phosphorylation despite strong Src kinase activity, but diffuse downstream activation of PI3-K by Src abrogates cell polarization and chemotaxis: "signalling requires silence".     

Inhibition of the phosphoinositide 3-kinase pathway decreases innate resistance to lipopolysaccharide toxicity in TLR4 deficient mice

Background

Upon lipopolysaccharide (LPS) stimulation, activation of both the Toll-like receptor 4 (TLR4) and phosphoinositide 3-kinase (PI3K) pathways serves to balance proinflammatory and anti-inflammatory responses. Although the antagonist to TLR4 represents an emerging promising target for the treatment of sepsis; however, the role of the PI3K pathway under TLR4-null conditions is not well understood. This goal of this study was to investigate the effect of inhibition of PI3K on innate resistance to LPS toxicity in a murine model.

Results

The overall survival of the cohorts receiving intraperitoneal injections of 100, 500, or 1000 μg LPS from Escherichia coli serotype 026:B6 after 7 d was 100%, 10%, and 10%, respectively. In contrast, no mortality was noted after 500-μg LPS injection in Tlr4^-/- mice. When the PI3K inhibitor {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002 was injected (1 mg/25 g body weight) 1 h prior to the administration of LPS, the overall survival of the Tlr4^-/- mice was 30%. In the Tlr4^-/- mice, the LPS injection induced no NF-κB activation but an increased Akt phosphorylation in the lung and liver, when compared to that of the C57BL/6 mice. Injection of 500 μg LPS led to a significant induction in O₂^- detected by electron paramagnetic resonance (EPR) spin trapping spectroscopy in the lung and liver at 3 and 6 h in C57BL/6 but not Tlr4^-/- mice. Addition of {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002 only significantly increased the O₂^- level in the lung and liver of the Tlr4^-/- mice but not in the C57BL/6 mice following 500-μg LPS injection. In addition, the serum IL-1β and IL-2 levels were more elevated in C57BL/6 mice than in Tlr4^-/- mice. Notably, IL-1β and IL-2 were significantly increased in Tlr4^-/- mice but not in the C57BL/6 mice when the PI3K pathway was inhibited by {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002 prior to LPS injection.

Conclusions

In this study, we demonstrate that innate resistance to LPS toxicity in Tlr4^-/- mice is impaired by inhibition of the PI3K pathway, with a corresponding increase in mortality and production of tissue O₂^- and inflammatory cytokines.     

PI 3-kinase and MAP kinase regulate bradykinin induced prostaglandin E(2) release in human pulmonary artery by modulating COX-2 activity

Here we studied the role of phosphoinositide 3-kinase (PI 3-kinase) and mitogen activated protein (MAP) kinase in regulating bradykinin (BK) induced prostaglandin E₂ (PGE₂) production in human pulmonary artery smooth muscle cells (HPASMC). BK increased PGE₂ in a three step process involving phospholipase A₂ (PLA₂), cyclooxygenase (COX) and PGE synthase (PGES). BK stimulated PGE₂ release in cultured HPASMC was inhibited by the PI 3-kinase inhibitor LY294002 and the p38 MAP kinase inhibitor SB202190. The inhibitory mechanism used by LY294002 did not involve cytosolic PLA₂ activation or COX-1, COX-2 and PGES protein expression but rather a novel effect on COX enzymatic activity. SB202190 also inhibited COX activity.     

eIF4E binding protein 1 and H-Ras are novel substrates for the protein kinase activity of class-I phosphoinositide 3-kinase

Class-I phosphoinositide 3-kinases (PI 3-kinases) are dual specificity enzymes that possess both lipid and protein kinase activity. While the best characterized property of this protein kinase is as an autokinase activity, there have also been reports it can phosphorylate exogenous substrates including peptides, IRS-1 and PDE-3B. The identification of two novel potential protein substrates of PI 3-kinase is described here. By employing in vitro kinase assays using recombinant proteins as the substrates, it is shown that the translational regulator 4EBP1 becomes phosphorylated by the p110alpha and p110gamma isoforms of class-I PI 3-kinases. The lipid kinase activity of both these isoforms is increased by allosteric binding of H-Ras or betagamma subunits of heterotrimeric G proteins, but we find this is not the case for the protein kinase activity. Surprisingly though, a site on H-Ras is phosphorylated by p110alpha and p110gamma. This raises the possibility that these proteins could serve as physiological substrates for the protein kinase activity of PI 3-kinase and suggests this activity operates in a physiological context by phosphorylating substrates other than the PI 3-kinase itself. This may be particularly important in regulating the interaction of Ras with PI 3-kinase.     

Involvement of phosphoinositide 3-kinase and PTEN protein in mechanism of activation of TRPC6 protein in vascular smooth muscle cells

TRPC6 is a cation channel in the plasma membrane that plays a role in Ca(2+) entry after the stimulation of a G(q)-protein-coupled or tyrosine-kinase receptor. TRPC6 translocates to the plasma membrane upon stimulation and remains there as long as the stimulus is present. However, the mechanism that regulates the trafficking and activation of TRPC6 are unclear. In this study we showed phosphoinositide 3-kinase and its antagonistic phosphatase, PTEN, are involved in the activation of TRPC6. The inhibition of PI3K by PIK-93, LY294002, or wortmannin decreased carbachol-induced translocation of TRPC6 to the plasma membrane and carbachol-induced net Ca(2+) entry into T6.11 cells. Conversely, a reduction of PTEN expression did not affect carbachol-induced externalization of TRPC6 but increased Ca(2+) entry through TRPC6 in T6.11 cells. We also showed that the PI3K/PTEN pathway regulates vasopressin-induced translocation of TRPC6 to the plasma membrane and vasopressin-induced Ca(2+) entry into A7r5 cells, which endogenously express TRPC6. In summary, we provided evidence that the PI3K/PTEN pathway plays an important role in the translocation of TRPC6 to the plasma membrane and may thus have a significant impact on Ca(2+) signaling in cells that endogenously express TRPC6.     

Interleukin-10 and interleukin-13 inhibit proinflammatory cytokine-induced ceramide production through the activation of phosphatidylinositol 3-kinase

Ceramide produced by hydrolysis of plasma membrane sphingomyelin (SM) in different cells including brain cells in response to proinflammatory cytokines [tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta)] plays an important role in coordinating cellular responses to stress, growth suppression, and apoptosis. The present study underlines the importance of IL-10 and IL-13, cytokines with potent antiinflammatory properties, in inhibiting the proinflammatory cytokine (TNF-alpha and IL-1beta)-mediated degradation of SM to ceramide in rat primary astrocytes. Treatment of rat primary astrocytes with TNF-alpha or IL-1beta led to rapid degradation of SM to ceramide, whereas IL-10 and IL-13 by themselves were unable to induce the degradation of SM to ceramide. Interestingly, both IL-10 and IL-13 prevented proinflammatory cytokine-induced degradation of SM to ceramide. Both IL-10 and IL-13 caused rapid activation of phosphatidylinositol (PI) 3-kinase, and inhibition of that kinase activity by wortmannin and LY294002 potently blocked the inhibitory effect of IL-10 and IL-13 on proinflammatory cytokine-mediated induction of ceramide production. This study suggests that the inhibition of proinflammatory cytokine-mediated degradation of SM to ceramide by IL-10 and IL-13 is mediated through the activation of PI 3-kinase. As ceramide induces apoptosis and IL-10 and IL-13 inhibit the induction of ceramide production, we examined the effect of IL-10 and IL-13 on proinflammatory cytokine-mediated apoptosis. Inhibition of TNF-alpha-induced apoptosis by IL-10 and IL-13 suggests that the antiapoptotic nature of IL-10 and IL-13 is probably due to the inhibition of ceramide production.     

Differential regulation of the phosphoinositide 3-kinase and MAP kinase pathways by hepatocyte growth factor vs. insulin-like growth factor-I in myogenic cells

Hepatocyte growth factor (HGF) promotes the proliferation of adult myoblasts and inhibits their differentiation, whereas insulin-like growth factor I (IGF-I) enhances both processes. Recent studies indicate that activation of the phosphoinositide 3'-kinase (PI3K) pathway promotes myoblast differentiation, whereas activation of the mitogen-activated protein kinase/extracellular signal-regulated protein kinase (MAPK/ERK) promotes proliferation and inhibits their differentiation. This simple model is confounded by the fact that both HGF and IGF-I have been shown to activate both pathways. In this study, we have compared the ability of HGF and IGF-I to activate PI3K and MAPK/ERK in i28 myogenic cells. We find that, although the two stimuli result in comparable recruitment of the p85alpha subunit of PI3K into complexes with tyrosine-phosphorylated proteins, the p85beta regulatory subunit and p110alpha catalytic subunit of PI3K are preferentially recruited into these complexes in response to IGF-I. In agreement with this observation, IGF-I is much more potent than HGF in stimulating phosphorylation of Akt/PKB, a protein kinase downstream of PI3K. In contrast, MAPK/ERK phosphorylation was higher in response to HGF and lasted longer, relative to IGF-I. Moreover, the specific PI3K inhibitor, Wortmannin, abolished MAPK/ERK and Elk-1 phosphorylation in HGF-treated cells, suggesting the requirement of PI3K in mediating the HGF-induced MAPK pathway. UO126, a specific MAPK pathway inhibitor, had no effect on PI3K activity or Akt phosphorylation, implying that at least in muscle cells, the MAPK/ERK pathway is not required for HGF-induced PI3K activation. These results provide a biochemical rationale for the previous observations that HGF and IGF-I have opposite effects on myogenic cells, consistent with studies linking PI3K activation to differentiation and MAPK/ERK activation to proliferation in these cells. Moreover, the finding that PI3K activity is required for HGF-induced MAPK activation suggests its additional role in proliferation, rather than exclusively in the differentiation of adult myoblasts.     

蛋白激酶B及其在磷脂酰肌醇3-激酶介导的信号转导中的作用

蛋白激酶Ｂ（ＰＫＢ）是原癌基因ｃ－ａｋｔ的表达产物,它参与由生长因子激活的经磷脂磷肌醇３－激酶（ＰＩ３Ｋ）介导的信号转导过程。与许多蛋白激酶相似,ＰＫＢ分子具有一特殊的ＡＨ／ＰＨ结构域（ＡＨ／ＰＨｄｏｍａｉｎ）,后者能介导信号分子间的相互作用。ＰＫＢ是ＰＩ３Ｋ直接的靶蛋白。ＰＩ３Ｋ产生的脂类第二信使ＰＩ－３,４,Ｐ２和ＰＩ－３,４,５－Ｐ３等均能与ＰＫＢ和磷酸肌醇依赖性蛋白激酶（ＰＤＫ）的ＡＨ／Ｐ     

Inhibitors of phosphoinositide 3-kinase cause defects in the postendocytic sorting of beta2-adrenergic receptors

Phosphatidylinositol 3-kinase inhibitors have been shown to affect endocytosis or subsequent intracellular sorting in various receptor systems. Agonist-activated beta(2)-adrenergic receptors undergo desensitization by mechanisms that include the phosphorylation, endocytosis and degradation of receptors. Following endocytosis, most internalized receptors are sorted to the cell surface, but some proportion is sorted to lysosomes for degradation. It is not known what governs the ratio of receptors that recycle versus receptors that undergo degradation. To determine if phosphatidylinositol 3-kinases regulate beta(2)-adrenergic receptor trafficking, HEK293 cells stably expressing these receptors were treated with the phosphatidylinositol 3-kinase inhibitors LY294002 or wortmannin. We then studied agonist-induced receptor endocytosis and postendocytic sorting, including recycling and degradation of the internalized receptors. Both inhibitors amplified the internalization of receptors after exposure to the beta-agonist isoproterenol, which was attributable to the sorting of a significant fraction of receptors to an intracellular compartment from which receptor recycling did not occur. The initial rate of beta(2)-adrenergic receptor endocytosis and the default rate of receptor recycling were not significantly altered. During prolonged exposure to agonist, LY294002 slowed the degradation rate of beta(2)-adrenergic receptors and caused the accumulation of receptors within rab7-positive vesicles. These results suggest that phosphatidylinositol 3-kinase inhibitors (1) cause a misrouting of beta(2)-adrenergic receptors into vesicles that are neither able to efficiently recycle to the surface nor sort to lysosomes, and (2) delays the movement of receptors from late endosomes to lysosomes.     

Emodin with PPARgamma ligand-binding activity promotes adipocyte differentiation and increases glucose uptake in 3T3-Ll cells

Emodin, one of the main active components in the root and rhizome of Rheum palmatum L, promoted the conversion of 3T3-L1 fibroblasts to adipocytes, as evidenced by increased glycerol-3-phosphate dehydrogenase (GPDH) activity and the expression of adipocyte aP2 mRNA, as well as accelerated triacylglycerol (TG) accumulation, which was associated with increased mRNA expression levels of both C/EBPalpha and PPARgamma2. By using surface plasmon resonance (SPR) experiment, it was showed that emodin exhibited a very high binding affinity to PPARgamma. In differentiated 3T3-L1 adipocytes, emodin induced a time- and dose-dependent increase in glucose uptake as well as GLUT1 and GLUT4 mRNA expression, and the rate of uptake was partly abrogated by wortmannin (phosphoinositide 3-kinase inhibitor). Meanwhile, insulin-stimulated glucose uptake was increased significantly after treatment with low doses of emodin, and the degree of potentiation was decreased thereafter in response to increasing concentrations. Furthermore, 50 microM emodin profoundly inhibited insulin-stimulated glucose uptake by 25%. These data suggest a new role for emodin as a PPARgamma agonist in 3T3-L1 cells. Besides, it is possible that emodin may also possess other properties contribute to glucose utilization in the adipocytes.